my jury members: Pr. Stephen BAGHDIGUIAN and Dr. Anne LAURENCON for accepting to participate in my thesis jury.

Behind this work, there have been tremendous efforts done by Dr. Ounissa AÏT AHMED. I would like to thank her, not only for welcoming me in her laboratory and giving me the opportunity to develop my experience abroad, but also for her continuous motivating encouragement, and her valuable advices. She has provided me immense support, guidance and leadership that not only helped through these four years but also lighten the way of my future.

I cannot forget the effort of Michèle CAPRI and her patience in teaching me different basics and laboratory techniques I used in preparing this thesis.

To accomplish this work, the Institut de Génétique Humaine has been the proper place. I acknowledge Giacomo Cavalli and Philippe Pasero for their consideration. It has been invaluable. I would like to thank all the leaders and members of computing, cell imaging, Drosophila, sterilization and the store facilities of the Institute of Human Genetics (IGH) for their unlimited co-operation during the four years of my stay in the institute. I also acknowledge the Institut de Recherche en Biothérapie (IRB), especially Pr Hamamah and his laboratory, Pr Klein and the administration of the IRB for providing space and access to the IRB facilities to complete the writing of my thesis manuscript in good conditions.

I acknowledge the members of my thesis committee, Robert feil and Olivier Coux for the time they spent in the follow up of my thesis work.

I am also thankful to the members of the postgraduate office in the University Montpellier 1 and the "Ecole Doctorale" CBS2 for their cooperation.

I would like to give my gratitude to everyone who helped and encouraged me through these years to pave the way for this work to see the daylight. I would like to acknowledge the Ministry of Higher Education and Scientific Research, Egypt for the financial support.

And, finally, I would like to thank my family members especially my wife, and my dear friends for their love and support.

Last but not least I thank God for giving me the strength and endurance to complete this work, and my parents for their absolute and sincere support.

Ahmed Algazeery

Abbreviations

AE Axial elements Asf1

Anti-silencing factor

Figures and tables

Figures 3: Localizations and functions of canonical core histones and histone variants in mammals .....

Introduction

General introduction

All living organisms have life cycle-they are born, they grow, and eventually die. In order to prevent themselves from dying out, they reproduce. In most multicellular eukaryotes, reproduction requires two parents-father and mother. Sexual reproduction relies on two main processes, the formation of paternal and maternal reproductive cells (gametes) and fertilization that combines the father's sperm and mother's egg genetic materials through syngamy (Figure 1).

Figure 1: Sexual reproduction cycle

Sexual reproduction involves two main critical processes: meiosis that results in haploid reproductive cells (sperm and egg) and fertilization after which the haploid parental genomes gather in the zygote before they undergo syngamy.

Mitosis and meiosis are the two strategies used by eukaryotes to propagate the genome. Both mitosis and meiosis are preceded by a single round of DNA replication which generates one pair of sister chromatids from each original parent chromosome (Figure 2). Chromosomes are then maintained by sister chromatid cohesion throughout the prophase [START_REF] Nasmyth | Disseminating the genome: joining, resolving, and separating sister chromatids during mitosis and meiosis[END_REF]. The two chromosomes are attached by their kinetochores to the spindle fibers apparatus that aligns the chromosomes on the equatorial plate and then separate them to opposite poles.

Despite similarities between the mitotic and meiotic cell divisions [START_REF] Budd | DNA polymerase I is required for premeiotic DNA replication and sporulation but not for X-ray repair in Saccharomyces cerevisiae[END_REF][START_REF] Forsburg | Mitotic replication initiation proteins are not required for pre-meiotic S phase[END_REF], many characteristic features distinguish the two cell division forms. In the mitotic cell division, the duplicated genome set undergoes only one chromosome segregation round, resulting in diploid daughter cells. Therefore, by mitosis, two daughter cells are produced from a single parent cell, each daughter cell being genetically identical to the original parent cell [START_REF] Cnudde | Meiosis: inducing variation by reduction[END_REF][START_REF] Zickler | The leptotene-zygotene transition of meiosis[END_REF].

In contrast to mitosis, the standard meiotic cell division process is unique in that it reduces the number of chromosomes by half, resulting in the formation of haploid gametes [START_REF] Murakami | Regulating the formation of DNA double-strand breaks in meiosis[END_REF][START_REF] Roeder | Meiotic chromosomes: it takes two to tango[END_REF]. Studies revealed that meiosis is a highly conserved process in sexually reproducing organisms [START_REF] Armstrong | Meiotic cytology and chromosome behaviour in wild-type Arabidopsis thaliana[END_REF]. The reduction of chromosome number is accomplished by following the single round of DNA replication phase by two consecutive rounds of meiotic chromosome segregation, meiosis I and meiosis II (Figure 2). Meiosis II, also called equational division, is most similar to the mitotic division, while genome set reduction is achieved by the first meiotic division (meiosis I) and so called reductional division (Figure 2). The meiotic pathway, at least in the male, ends with the production of four haploid cells different from a single parent cell, each daughter cell carries half the amount of parental genetic material [START_REF] Cnudde | Meiosis: inducing variation by reduction[END_REF][START_REF] Zickler | The leptotene-zygotene transition of meiosis[END_REF].

The prophase of meiosis I was found to be extended compared to the prophase of the second meiotic division or to that of mitotic prophase. The extended duration of prophase I is not surprising given the complexity of events that must occur in order to segregate the homologous chromosomes. The homologous chromosomes must come into proximity, recognize each other as a homolog, intimately associate, and align with one another along their lengths and exchange their genetic information through a process called homologous recombination. At this stage they are called bivalent chromosomes. These major chromosomal events of meiosis, pairing, synapsis, and recombination are largely conserved across eukaryotes. These processes associated with many cytological changes further divide the meiotic prophase I into five substages: leptotene, zygotene, pachytene, diplotene and diakinesis [START_REF] Pawlowski | Coordinating the events of the meiotic prophase[END_REF][START_REF] Petronczki | Un menage a quatre: the molecular biology of chromosome segregation in meiosis[END_REF][START_REF] Zickler | Meiotic chromosomes: integrating structure and function[END_REF].

One critical function of this exchange is that it provides a physical connection between the homologous chromosomes, the chiasma. Chiasmata allow chromosomes to align properly on the spindle and to separate accurately such that they will orient to opposite poles after prophase I is completed. Proper segregation of homologous chromosomes at anaphase I relies on chiasmata dissolution as a consequence of cohesion release from sister chromatid arms [START_REF] Cai | The Arabidopsis SYN1 cohesin protein is required for sister chromatid arm cohesion and homologous chromosome pairing[END_REF]. Subsequent release of cohesion from sister chromatid centromeric regions is a prerequisite for the equational division of chromosomes at anaphase II [START_REF] Chelysheva | AtREC8 and AtSCC3 are essential to the monopolar orientation of the kinetochores during meiosis[END_REF].

Figure 2: Schematic representation of a meiotic cell division as compared to mitosis

Before meiosis and mitosis the genome content of the dividing cell is duplicated by one round of DNA replication. In mitosis a single round of chromosome segregation occurs, whereas meiosis involves two chromosome segregation rounds. At the first meiotic division (reductional), homologous chromosomes are segregated, and at the second meiotic division (equational), sister chromatids are separated [START_REF] Alberts | Molecular biology of the cell[END_REF].

Homologous recombination is initiated principally by the formation of meiotically induced Double Strand Breaks (DSBs). This process is a highly controlled process, in terms of both position and timing. Failing to initiate DSBs or initiating them at the wrong time or in the wrong place has severe consequences on the subsequent chromosome segregations and disastrous consequences on the viability of the meiotic products [START_REF] Lake | The Drosophila zinc finger protein trade embargo is required for double strand break formation in meiosis[END_REF]. If the meiotic divisions were not properly executed, the organism fails to produce proper gametes and if fertilization occurs, it results in aneuploid embryos, an outcome that typically causes spontaneous abortion or birth defects [START_REF] Mehta | Cohesin: A guardian of genome integrity[END_REF].

In eukaryotes, DNA replication and chromosome segregation are temporally separated. For successful cell division, these processes must be carefully coordinated. Several associations have to be established between parental chromosome pairs to keep them together from the time of their generation during S phase till the time they split during metaphase to anaphase transition. Therefore, the identification of the molecular mechanisms of sister chromatids cohesion is of central importance for understanding how the cell governs its meiotic cycle [START_REF] Peters | The cohesin complex and its roles in chromosome biology[END_REF]. Studies of these mechanisms revealed two main protein complexes responsible to ensure this association:

-The molecular 'glue' established during DNA-replication that keeps the sister chromatids together from S-phase to metaphase-anaphase transition is known as "cohesin", a multiprotein complex which is believed to entrap the sister chromatids [START_REF] Mehta | Cohesin: A guardian of genome integrity[END_REF].

-The synaptonemal complex (SC), a tri-partite proteinaceous structure that forms between homologous chromosomes during prophase I [START_REF] Khetani | Regulation of meiotic cohesion and chromosome core morphogenesis during pachytene in Drosophila oocytes[END_REF].

In spite of the similarity of the major meiotic events during both spermatogenesis and oogenesis, each process has its own complexity. Spermatogenesis is a continuous process that occurs throughout most of the life of the male [START_REF] Cooper | Meiotic control of the APC/C: similarities & differences from mitosis[END_REF]. In many animal species, the sperm DNA is packaged with specific non-histone proteins, protamines. At fertilization, these proteins have to be removed and replaced by maternally provided histones.

My thesis work is dedicated to unveil such mechanisms in an organism that provides powerful genetic tools, Drosophila melanogaster. We aimed in the present work to contribute to gain insight into the molecules and mechanisms that are involved in oocyte meiosis and paternal chromatin assembly in the zygote. The discovery and functional analysis of Yemanuclein (also called Yemanuclein-alpha) by Aït-Ahmed and collaborators have provided the basis for this work. Before the presentation of my experimental data, I will provide some bibliographical background on these critical events on which sexual reproduction relies, in the next two chapters.

Chapter 1: Molecular analysis of early meiotic prophase

Essentially, meiosis is a modified mitotic cell division. Despite the similarities between them in many mechanisms, the most notable difference is the two-steps chromosome segregation process that characterizes meiosis [START_REF] Budd | DNA polymerase I is required for premeiotic DNA replication and sporulation but not for X-ray repair in Saccharomyces cerevisiae[END_REF][START_REF] Forsburg | Mitotic replication initiation proteins are not required for pre-meiotic S phase[END_REF].

Chromosomes are first replicated during the period of pre-meiotic S phase that is similar to S phase in mitosis (Figure 3). However there is a key difference between S phase in mitosis and pre-meiotic S phase: pre-meiotic S phase takes much longer (25-30 minutes for mitotic Sphase vs. ~75 minutes for pre-meiotic S-phase) [START_REF] Cha | Progression of meiotic DNA replication is modulated by interchromosomal interaction proteins, negatively by Spo11p and positively by Rec8p[END_REF]. This extra time may be needed for the proper establishment of the additional chromosome cohesion required for the subsequent critical meiotic processes. Indeed sister chromatids must have a specific cohesion that maintains them together until metaphase II [START_REF] Petronczki | Un menage a quatre: the molecular biology of chromosome segregation in meiosis[END_REF].

Prophase of meiosis I also takes much longer. During this lengthy phase, the homologous chromosomes condense into thread-like structures which first align and pair before becoming closely associated along their length by the tripartite proteinaceous structure known as the [START_REF] Watanabe | Modifying sister chromatid cohesion for meiosis[END_REF]. (* found in yeast). synaptonemal complex (SC). This process is known as synapsis. The different stages of prophase I are distinguished based on the progression of synapsis. In leptotene the homologous chromosomes pair along their length before synapsis is initiated. The SC starts to form between paired homologous chromosomes at zygotene; this process is completed at pachytene and the SC starts to disassemble at diplotene. By the final stage of prophase I, diakinesis, the chromosomes begin to condense to achieve their shortest length at metapahase I. Meiotic recombination occurs during prophase I and the synaptonemal complex plays a key role in this critical event [START_REF] Page | The genetics and molecular biology of the synaptonemal complex[END_REF]. Yemanuclein is a key player in these events [START_REF] Meyer | A single mutation results in diploid gamete formation and parthenogenesis in a Drosophila yemanuclein-alpha meiosis I defective mutant[END_REF], this manuscript. Therefore in this chapter I will concentrate on the bibliographical background of these early meiotic events that are required for the formation of the bivalents, a basis for homologous chromosomes segregation at meiosis I.

I. Components of meiotic chromosome cohesion

DNA replication and chromosome segregation of eukaryotes are temporally separated. A remarkable feature of meiosis is a prolonged prophase arrest following formation of bivalent chromosomes especially in female meiosis in mammals. Indeed it lasts from around birth until ovulation, an interval that might be many months in the mouse and several decades in humans [START_REF] Tachibana-Konwalski | Rec8-containing cohesin maintains bivalents without turnover during the growing phase of mouse oocytes[END_REF]. Therefore, it is essential to keep the sister chromatids together from the time of their generation during S phase till the time they split at metaphase to anaphase transition. Without the cohesion between the two sister chromatids they could separate from each other prematurely. If the meiotic divisions are not properly executed, after fertilization aneuploid embryos may form, an outcome that typically causes spontaneous abortion or birth defects [START_REF] Mehta | Cohesin: A guardian of genome integrity[END_REF]. Identification of the molecular mechanisms that underlie sister chromatids cohesion is therefore of central importance [START_REF] Peters | The cohesin complex and its roles in chromosome biology[END_REF].

The identification of the molecules that are involved in this process is a first step in the unveiling of the mechanisms.

A. Structural maintenance of chromosomes (SMC) protein family

The first proteins that are essential for cohesion were identified by genetic screens for mutants that separate sister chromatids precociously before anaphase, during mitosis in the budding yeast Saccharomyces cerevisiae [START_REF] Guacci | A direct link between sister chromatid cohesion and chromosome condensation revealed through the analysis of MCD1 in S. cerevisiae[END_REF][START_REF] Michaelis | Cohesins: chromosomal proteins that prevent premature separation of sister chromatids[END_REF] and during meiosis I in the fruit fly Drosophila melanogaster [START_REF] Davis | Genetic analysis of a meiotic mutant resulting in precocious sister-centromere separation in Drosophila melanogaster[END_REF][START_REF] Kerrebrock | The Drosophila mei-S332 gene promotes sister-chromatid cohesion in meiosis following kinetochore differentiation[END_REF][START_REF] Miyazaki | Sister-chromatid misbehavior in Drosophila ord mutants[END_REF][START_REF] Peters | The cohesin complex and its roles in chromosome biology[END_REF].

The cohesion between the two sister chromatids is achieved by specific protein complexes that have subunits called structural maintenance of chromosome (SMC) proteins [START_REF] Haering | Molecular architecture of SMC proteins and the yeast cohesin complex[END_REF][START_REF] Hirano | SMC protein complexes and higher-order chromosome dynamics[END_REF]. SMC proteins are a family of large coiled-coil proteins, which are conserved in prokaryotes, eukaryotes and archea (Figure 4). In eukaryotes, six different SMC proteins have been discovered that form specific combinations, generating three heterodimers with different functions; Smc1/Smc3 heterodimers make up the structural basis for cohesins, Smc2 and Smc4 complexes forming condensins and a third complex with the two remaining SMC proteins, are Smc5 and Smc6 (Hirano, 2005b;[START_REF] Losada | Dynamic molecular linkers of the genome: the first decade of SMC proteins[END_REF][START_REF] Nasmyth | The structure and function of SMC and kleisin complexes[END_REF]. SMC proteins are large complexes that regulate chromosome architecture and function. They maintain genome integrity by enabling post-replicative DNA repair, shaping chromosomes in preparation for cell division, and holding sister chromatids together to ensure that daughter cells receive a full complement of chromosomes [START_REF] Seitan | Cohesin and chromatin organisation[END_REF].

Figure 4: Architecture of the SMC complexes

A) The core of each SMC complex is formed by two SMC proteins; each contains a hinge domain which has ATPase head and an intramolecular antiparallel coiled coil that connects the two proteins. B) Showing various SMC complexes composed of a specific SMC dimer and several non-SMC subunits: The SMC1/SMC3 cohesin, the condensin I and condensin II and the SMC5/6 complex [START_REF] Wu | The Smc complexes in DNA damage response[END_REF].

The SMC proteins contain about 1,000 amino acids and share similar domain structures. The ATPase domain of each SMC protein is separated into N-and C-terminal halves by a long linker. The two nucleotide-binding Walker A and Walker B motifs reside in the two different ATPase halves. The SMC linker folds into an intra-molecular antiparallel coiled coil and allows the N-terminal ATPase half of an SMC protein to fold back to its C-terminal ATPase half and create a single globular ATPase head (Figure 4). The hinge domain at one end of the coiled coil mediates the heterodimerization of eukaryotic SMC proteins [START_REF] Hirano | At the heart of the chromosome: SMC proteins in action[END_REF][START_REF] Losada | Dynamic molecular linkers of the genome: the first decade of SMC proteins[END_REF][START_REF] Nasmyth | The structure and function of SMC and kleisin complexes[END_REF]. The two ATPase heads at the other end of the coiled coil can transiently interact with each other to bind and hydrolyze ATP. As revealed by electron microscopy, the SMC heterodimers can adopt different conformations, including V-shaped dimer and ring-like structures, possibly depending on the nucleotidebinding states of their ATPase heads [START_REF] Anderson | Condensin and cohesin display different arm conformations with characteristic hinge angles[END_REF][START_REF] Melby | The symmetrical structure of structural maintenance of chromosomes (SMC) and MukB proteins: long, antiparallel coiled coils, folded at a flexible hinge[END_REF].

Condensin complex

Condensin is a pentameric complex, which comprises two members of the SMC family, SMC2 and SMC4, as well as three non-SMC regulatory subunits (Figure 4). In higher eukaryotes two condensin complexes were reported, condensin I and condensin II. Both condensins contain a heterodimer of SMC2 and SMC4 backbone but associate with different regulatory proteins. Condensin I comprises CAP-H/Barren, CAP-D2 and CAP-G whereas condensin II comprises CAP-H2, CAP-D3 and CAP-G2 (Hirano, 2005a;[START_REF] Ono | Differential contributions of condensin I and condensin II to mitotic chromosome architecture in vertebrate cells[END_REF][START_REF] Yeong | Identification of a subunit of a novel Kleisinbeta/SMC complex as a potential substrate of protein phosphatase 2A[END_REF]. Condensin I and II are both present in metazoans, however fungi have a single condensin complex, which is most similar to condensin I of metazoans. In Caenorhabditis elegans, a third condensin-like complex -the dosage compensation complex (DCC)-regulates the expression of X-linked genes [START_REF] Jans | A condensin-like dosage compensation complex acts at a distance to control expression throughout the genome[END_REF].

SMC5/SMC6 complex

The SMC5/SMC6 complex (Figure 4) has a prominent role in the metabolism of meiotic DSBs and recombination but its exact function remains elusive [START_REF] Farmer | The Smc5-Smc6 complex is required to remove chromosome junctions in meiosis[END_REF][START_REF] Lehmann | The role of SMC proteins in the responses to DNA damage[END_REF]. Smc5-Smc6 complex is thought to promote the repair of DNA doublestrand breaks (DSBs) by error-free sister-chromatid recombination (SCR), which makes it essential for genome stability and suppressing the inappropriate non-sister recombination events [START_REF] De Piccoli | Smc5-Smc6 mediate DNA double-strand-break repair by promoting sister-chromatid recombination[END_REF]. Recently, Smc5-Smc6 complex was found to localize to some specific chromosome regions during meiotic prophase I. The absence of Smc5-Smc6 complex in meiotic cells results in many catastrophic meiotic divisions as a consequence of unresolved linkages between homologous chromosomes [START_REF] Farmer | The Smc5-Smc6 complex is required to remove chromosome junctions in meiosis[END_REF].

Cohesin complex

Cohesin is a large multiprotein complex (Figure 5) conserved from yeast to human (Table 1) [START_REF] Haering | Building and breaking bridges between sister chromatids[END_REF][START_REF] Nasmyth | Disseminating the genome: joining, resolving, and separating sister chromatids during mitosis and meiosis[END_REF]. The spatiotemporal regulation of the association and dissociation of the cohesin complex to and from the sister chromatids respectively, is instrumental in faithful segregation of the sister chromatids during both mitosis and meiosis [START_REF] Mehta | Cohesin: A guardian of genome integrity[END_REF]. Cohesin complex localizes to the meiotic axis in many species [START_REF] Cai | The Arabidopsis SYN1 cohesin protein is required for sister chromatid arm cohesion and homologous chromosome pairing[END_REF][START_REF] Eijpe | Meiotic cohesin REC8 marks the axial elements of rat synaptonemal complexes before cohesins SMC1beta and SMC3[END_REF][START_REF] Klein | A central role for cohesins in sister chromatid cohesion, formation of axial elements, and recombination during yeast meiosis[END_REF][START_REF] Page | Chromosome choreography: the meiotic ballet[END_REF][START_REF] Severson | The axial element protein HTP-3 promotes cohesin loading and meiotic axis assembly in C. elegans to implement the meiotic program of chromosome segregation[END_REF]. The major function of cohesin complex, as its name indicates, is to regulate sister chromatids cohesion both their proper cohesion and timely separation. In addition to its canonical function cohesin has been suggested by several lines of investigation in recent years to play additional roles in apoptosis, DNA-damage response, transcriptional regulation and hematopoiesis [START_REF] Panigrahi | A cohesin-RAD21 interactome[END_REF].

Chromatin immunoprecipitation (ChIP) technique of the budding yeast genome revealed that cohesin binds to discrete cohesin attachment regions (CARs) on chromosome arms and to a larger domain surrounding centromeres [START_REF] Blat | Cohesins bind to preferential sites along yeast chromosome III, with differential regulation along arms versus the centric region[END_REF][START_REF] Megee | The centromeric sister chromatid cohesion site directs Mcd1p binding to adjacent sequences[END_REF][START_REF] Peters | The cohesin complex and its roles in chromosome biology[END_REF][START_REF] Tanaka | Identification of cohesin association sites at centromeres and along chromosome arms[END_REF]. Importantly, cohesin was found to associate with chromosomes in budding yeast from late G1 phase until metaphase [START_REF] Michaelis | Cohesins: chromosomal proteins that prevent premature separation of sister chromatids[END_REF].

Figure 5: The cohesin complex and its associated proteins

Cohesin complex is composed of four subunits: a heterodimer of two very long SMC molecules (SMC1 and SMC3), associated with two smaller non-SMC subunits (kleisin family Mcd1/Scc1/Rad21 and stromalin family Scc3/SA1/SA2) proteins. Together these subunits form a large ring capable of topologically encircling DNA strands. Other proteins regulate cohesin's binding to DNA and its residency there. The NIPBL/MAU2 dimer loads cohesin onto DNA, whereas WAPL/PDS5 release cohesin from chromosomes by opening the SMC3-RAD21 interface [START_REF] Horsfield | Diverse developmental disorders from the one ring: distinct molecular pathways underlie the cohesinopathies[END_REF].

Structurally, cohesin complex is composed of four subunits (Figure 5): a heterodimer of two very long SMC protein molecules (SMC1 and SMC3), associated with two smaller non-SMC subunits(kleisin family Mcd1/Scc1/Rad21 and stromalin family Scc3/SA1/SA2) proteins [START_REF] Nasmyth | The structure and function of SMC and kleisin complexes[END_REF][START_REF] Onn | Sister chromatid cohesion: a simple concept with a complex reality[END_REF][START_REF] Shintomi | How are cohesin rings opened and closed?[END_REF].

Kleisins are a superfamily of non SMC protein partners evolutionarily conserved from prokaryotes to humans [START_REF] Schleiffer | Kleisins: a superfamily of bacterial and eukaryotic SMC protein partners[END_REF]. The term kleisin is derived from the Greek word for closure [START_REF] Xiong | Regulators of the cohesin network[END_REF]. The SMC heterodimer and the non-SMC subunits form the complex which entraps the sister chromatids in its tri-partite ring structure (Figure 5) [START_REF] Haering | Molecular architecture of SMC proteins and the yeast cohesin complex[END_REF][START_REF] Losada | Dynamic molecular linkers of the genome: the first decade of SMC proteins[END_REF].

Role in cohesion Saccharomyces Cerevisiae

Schizosaccharomyces pombe

Drosophila melanogaster

Vertebrates

Cohesin complex subunits 2)M is only distantly related to Rec8 and does not fulfill typical Rec8 function [START_REF] Heidmann | The Drosophila meiotic kleisin C(2)M functions before the meiotic divisions[END_REF]. (**) Rad21L is a newly identified kleisin whose properties are reminiscent of C( 2)M [START_REF] Ishiguro | A new meiosis-specific cohesin complex implicated in the cohesin code for homologous pairing[END_REF][START_REF] Lee | RAD21L, a novel cohesin subunit implicated in linking homologous chromosomes in mammalian meiosis[END_REF][START_REF] Llano | Meiotic cohesin complexes are essential for the formation of the axial element in mice[END_REF]. This table is taken from [START_REF] Xiong | Regulators of the cohesin network[END_REF] with modifications, red indicates a meiosis-specific function.

Scc1/Mcd1 Rad21 Rad21 Rad21 Scc3/Irr1 Psc3, Rec11 SA1 SA1, SA2, STAG3 Smc1 Psm1 Smc1 Smc1α, Smc1β Smc3 Psm3 Smc3 (Cap) Smc3 Rec8 Rec8 C (
Cohesin is a highly dynamic structure, the association and dissociation of cohesin complex to and from chromosomes alternate through every cell cycle (Figure 6). Cohesin is loaded onto unreplicated chromosomes in G1, followed by cohesion establishment during DNA replication in S phase and maintenance of cohesion in G2. In M phase sister, chromatids cohesion is resolved, and chromosomes segregate. It is noteworthy that in mammals two meiosis specific kleisins (Rec8 and Rad21L) have been identified that play essential roles in meiosis I specific cohesion and homologous recombination. Interestingly in Drosophila there is no Rad21 paralogous sequence (see Table 1 for more details). Two pairs of sister chromatids, that is, two homologs, synapse thanks to the synaptonemal complex (SC; yellow) formation. Recombination occurs between non-sister chromatids. In anaphase I, cohesin complex in the chromosome arms is dissolved and the homologs segregate with cohesin only at the still intact centromeres. In anaphase II, centromeric cohesion is dissolved, and the sister chromatids segregate. Green arrowheads indicate orientation of kinetochores [START_REF] Revenkova | Keeping sister chromatids together: cohesins in meiosis[END_REF].

i. Loading of cohesin at G1 phase

The loading of the cohesin complex onto chromosomes is mediated by Scc2-Scc4 loading proteins, (Figure 6) [START_REF] Ciosk | Cohesin's binding to chromosomes depends on a separate complex consisting of Scc2 and Scc4 proteins[END_REF][START_REF] Takahashi | Recruitment of Xenopus Scc2 and cohesin to chromatin requires the pre-replication complex[END_REF]. Scc2 was first discovered in budding yeast [START_REF] Ciosk | Cohesin's binding to chromosomes depends on a separate complex consisting of Scc2 and Scc4 proteins[END_REF] and it belongs to a family called chromosomal adherins.

Scc2 homologs have been discovered in fission yeast (Mis4), Drosophila (Nipped-B),

Xenopus and human (NIPBL) (Table 1) [START_REF] Furuya | Faithful anaphase is ensured by Mis4, a sister chromatid cohesion molecule required in S phase and not destroyed in G1 phase[END_REF][START_REF] Gillespie | Scc2 couples replication licensing to sister chromatid cohesion in Xenopus egg extracts[END_REF][START_REF] Rollins | Nipped-B, a Drosophila homologue of chromosomal adherins, participates in activation by remote enhancers in the cut and Ultrabithorax genes[END_REF][START_REF] Tonkin | NIPBL, encoding a homolog of fungal Scc2-type sister chromatid cohesion proteins and fly Nipped-B, is mutated in Cornelia de Lange syndrome[END_REF]. Scc4 protein was found in humans [START_REF] Watrin | Human Scc4 is required for cohesin binding to chromatin, sister-chromatid cohesion, and mitotic progression[END_REF] while its first homolog, Ssl3 was discovered in fission yeast. [START_REF] Bernard | A screen for cohesion mutants uncovers Ssl3, the fission yeast counterpart of the cohesin loading factor Scc4[END_REF]. In C. elegans and Drosophila the Scc4 homologue is Mau-2 protein, which interacts with the Scc2 orthologs (Table 1) [START_REF] Benard | mau-2 acts cell-autonomously to guide axonal migrations in Caenorhabditis elegans[END_REF][START_REF] Seitan | Metazoan Scc4 homologs link sister chromatid cohesion to cell and axon migration guidance[END_REF]. SCC2 and SCC4 and their orthologs are found to be specifically required for sister chromatids cohesion establishment and not its maintenance [START_REF] Skibbens | Unzipped and loaded: the role of DNA helicases and RFC clamp-loading complexes in sister chromatid cohesion[END_REF].

ii. Establishment of cohesion at s-phase

After having been loaded onto unreplicated DNA, the loaded cohesin then becomes cohesive during DNA replication and has been proposed to topologically embrace both sister chromatids inside its ring to establish sister-chromatid cohesion (Figure 6) [START_REF] Cipak | Sister chromatids caught in the cohesin trap[END_REF][START_REF] Uhlmann | A matter of choice: the establishment of sister chromatid cohesion[END_REF]. Cohesion can only be established once DNA has been replicated during S phase. Several lines of evidence point towards the direction that cohesion establishment is tightly coupled to DNA replication. Despite the mechanism by which cohesin is converted to the cohesive state during DNA replication is not completely understood, it is known that it requires the acetylation of Smc3 by the Eco1 family of acetyltransferases (Figure 6) [START_REF] Skibbens | Ctf7p is essential for sister chromatid cohesion and links mitotic chromosome structure to the DNA replication machinery[END_REF]. Eco1 contains a zinc finger in the N-terminal portion and an acetyltransferase domain in the C-terminal portion. Eco1 appears to be a global regulator of cohesion and it may be able to take advantage of the proximity of sisters during DNA replication to promote the cohesion [START_REF] Xiong | Regulators of the cohesin network[END_REF]. In humans, two paralogs of ECO1 were discovered (Table 1), ESCO1 and ESCO2. An acetyltransferase ortholog called Deco needed for centromeric cohesion has been identified in Drosophila [START_REF] Hou | The acetyltransferase activity of San stabilizes the mitotic cohesin at the centromeres in a shugoshin-independent manner[END_REF][START_REF] Williams | Two putative acetyltransferases, san and deco, are required for establishing sister chromatid cohesion in Drosophila[END_REF].

iii. Maintenance of cohesin ring

After cohesion establishment in S phase, sister chromatids are held together by cohesin throughout G2. Independent of the cohesion establishment factors, several proteins (including Pds5, Sororin, and Wapl) (Table 1) were reported to maintain the sister chromatids cohesion following S phase (Figure 6) [START_REF] Skibbens | Ctf7p is essential for sister chromatid cohesion and links mitotic chromosome structure to the DNA replication machinery[END_REF][START_REF] Toth | Yeast cohesin complex requires a conserved protein, Eco1p(Ctf7), to establish cohesion between sister chromatids during DNA replication[END_REF]. In budding yeast, Pds5

is the essential gene required for cohesion maintenance during G2/M phase [START_REF] Hartman | Pds5p is an essential chromosomal protein required for both sister chromatid cohesion and condensation in Saccharomyces cerevisiae[END_REF][START_REF] Panizza | Pds5 cooperates with cohesin in maintaining sister chromatid cohesion[END_REF][START_REF] Stead | Pds5p regulates the maintenance of sister chromatid cohesion and is sumoylated to promote the dissolution of cohesion[END_REF]. In metazoans two PDS5 genes were identified, Pds5A and Pds5B, Both Pds5A and B-deficient mice die at birth [START_REF] Zhang | Dosage effects of cohesin regulatory factor PDS5 on mammalian development: implications for cohesinopathies[END_REF]Zhang et al., 2007). Sororin is a vertebrate-specific component of the cohesin network [START_REF] Rankin | Sororin, a substrate of the anaphase-promoting complex, is required for sister chromatid cohesion in vertebrates[END_REF][START_REF] Schmitz | Sororin is required for stable binding of cohesin to chromatin and for sister chromatid cohesion in interphase[END_REF]. Sororin was also suggested to be important for the DSBs repair in G2 phase [START_REF] Schmitz | Sororin is required for stable binding of cohesin to chromatin and for sister chromatid cohesion in interphase[END_REF]. WPL1/RAD61/Wapl is the evolutionarily conserved gene involved in cohesion maintenance (Table 1). Wapl (wings apart like), first discovered in Drosophila [START_REF] Verni | Genetic and molecular analysis of wings apart-like (wapl), a gene controlling heterochromatin organization in Drosophila melanogaster[END_REF] was found to promote the cohesin dissociation from chromosome arms in mammalian cells during prophase and increases its turnover on chromatin during interphase [START_REF] Gandhi | Human Wapl is a cohesin-binding protein that promotes sisterchromatid resolution in mitotic prophase[END_REF][START_REF] Kueng | Wapl controls the dynamic association of cohesin with chromatin[END_REF].

iv. Dissociation of cohesin at anaphase

At the metaphase-anaphase transition (Figure 6), the proteolytic cleavage of the kleisin subunit by protease enzyme separase/Esp1 allows the cohesin ring to come apart and chromosomes to separate [START_REF] Ciosk | An ESP1/PDS1 complex regulates loss of sister chromatid cohesion at the metaphase to anaphase transition in yeast[END_REF][START_REF] Funabiki | Fission yeast Cut1 and Cut2 are essential for sister chromatid separation, concentrate along the metaphase spindle and form large complexes[END_REF]. In mammalian cells, cohesion between chromosome arms is removed after kleisin Rad21 phosphorylation by Polo kinase permits the cohesion to dissolve [START_REF] Gimenez-Abian | Regulation of sister chromatid cohesion between chromosome arms[END_REF][START_REF] Hauf | Dissociation of cohesin from chromosome arms and loss of arm cohesion during early mitosis depends on phosphorylation of SA2[END_REF]. The presence of Polo kinase phosphorylation is of special importance during meiotic divisions, as it permits the non-destructive dissociation of cohesion between the sister chromatids at meiosis I leaving the centromeric cohesion maintained until meiosis II when it is targeted by separase (Figure 6) [START_REF] Buonomo | Disjunction of homologous chromosomes in meiosis I depends on proteolytic cleavage of the meiotic cohesin Rec8 by separin[END_REF][START_REF] Xiong | Regulators of the cohesin network[END_REF].

B. Synaptonemal complex (SC)

Synaptonemal complex (SC) is a meiosis-specific structure first identified by [START_REF] Fawcett | The fine structure of chromosomes in the meiotic prophase of vertebrate spermatocytes[END_REF]Moses in 1956 (Fawcett, 1956;[START_REF] Moses | Chromosomal structures in crayfish spermatocytes[END_REF]. It is found almost universally in sexually reproducing eukaryotic organisms [START_REF] Anderson | Crossing over in a Case of Attached X Chromosomes in DROSOPHILA MELANOGASTER[END_REF][START_REF] Ashburner | Drosophila : a laboratory handbook[END_REF][START_REF] Bridges | Non-Disjunction as Proof of the Chromosome Theory of Heredity (Concluded)[END_REF].

The ultrastructural analysis of the SC by transmission electron microscopy has revealed a tripartite proteinaceous structure (Figure 7). It comprises two chromosome axes (also called lateral elements -LE), surrounding a central element (CE). The axes of the two homologous chromosomes and the CE are connected along their entire length by fine fibrillar structures, the transverse filaments (TF), generating a zipper-or ladder-like structure. The TF and the CE together form the central region (CR) of the SC [START_REF] Handel | Genetics of mammalian meiosis: regulation, dynamics and impact on fertility[END_REF][START_REF] Kouznetsova | Meiosis in mice without a synaptonemal complex[END_REF]. Synapsis initiates during early prophase I, where each pair of sister chromatids undergoes shortening along their longitudinal axes, resulting in the formation of 'chromosome cores' upon which the axial/lateral elements (AEs/LEs) of the SC assemble [START_REF] Revenkova | Cohesin SMC1 beta is required for meiotic chromosome dynamics, sister chromatid cohesion and DNA recombination[END_REF][START_REF] Stack | A model for chromosome structure during the mitotic and meiotic cell cycles[END_REF]. The cohesion between meiotic sister chromatids was reported by many evidences to play an essential role in assembly of the axial/lateral elements (AEs/LEs) of the SC [START_REF] Cai | The Arabidopsis SYN1 cohesin protein is required for sister chromatid arm cohesion and homologous chromosome pairing[END_REF][START_REF] Klein | A central role for cohesins in sister chromatid cohesion, formation of axial elements, and recombination during yeast meiosis[END_REF][START_REF] Pasierbek | A Caenorhabditis elegans cohesion protein with functions in meiotic chromosome pairing and disjunction[END_REF][START_REF] Webber | The cohesion protein ORD is required for homologue bias during meiotic recombination[END_REF].

During pachytene, SC central element proteins join each set of homologous AEs/LEs along their entire length resulting in synapsis of homologues [START_REF] Page | The genetics and molecular biology of the synaptonemal complex[END_REF].

Synaptonemal complexes were found to stabilize all the counterpart associations from the period when the pairing interactions are lost until the crossovers are formed [START_REF] Zickler | Meiotic chromosomes: integrating structure and function[END_REF]. Despite the complexity and diversity of use of synaptonemal complexes, it appears a number of analogies between different species [START_REF] Hunter | Synaptonemal complexities and commonalities[END_REF]. Proteins constituting synaptonemal complex were isolated by biochemical or genetic screens; they are described in the following paragraph.

II. Regulation of meiotic recombination

A. Initiation of meiotic recombination

Double strand breaks (DSBs) formation and repair

An important aspect that differentiates mitosis from meiosis is that levels of genetic recombination in meiosis are much higher (100-1000 fold) during meiosis than in mitosis.

This difference is largely due to the programmed formation of DNA double stranded breaks (DSBs). Their formation is considered to be a tightly controlled process in terms of position, timing and number. The mechanism(s) that controls the DSB formation is still not completely understood [START_REF] Lake | The molecular control of meiotic chromosomal behavior: events in early meiotic prophase in Drosophila oocytes[END_REF]. However it is known that formation of DSBs at the wrong time or in the wrong place has severe consequences in the viability of meiotic products. A specific feature of meiosis is that DSBs are repaired by using the two homologs (Figure 8). This homologous repair of DSBs can result in two types of recombination outcomes. One type is crossing over (reciprocal exchange) while the other type results in gene conversion [START_REF] Fogel | Meiotic gene conversion in yeast tetrads and the theory of recombination[END_REF][START_REF] Fogel | Recombination in yeast[END_REF][START_REF] Holliday | Molecular aspects of genetic exchange and gene conversion[END_REF][START_REF] Stadler | The Relationship of Gene Conversion to Crossing over in Neurospora[END_REF]. Several models have been proposed to explain both crossing over and gene conversion. Schematic representation shows the major chromosomal events of meiotic prophase I in Saccharomyces cerevisiae [START_REF] Hochwagen | Checking your breaks: surveillance mechanisms of meiotic recombination[END_REF] and Drosophila melanogaster (Joyce and McKim, 2011;[START_REF] Lake | The molecular control of meiotic chromosomal behavior: events in early meiotic prophase in Drosophila oocytes[END_REF]. Prophase I involves three main processes; synapsis, double strand break (DSB) formation and DSB repair. These processes are largely conserved across eukaryotes and are associated with many cytological changes that further divide the meiotic prophase I into five substages: leptotene, zygotene, pachytene, diplotene and diakinesis. Meiotic recombination is achieved via a physical connection between the homologous chromosomes, the chiasma, which allows chromosomes to align properly on the spindle after prophase I is completed at diakinesis. Note the differences in the timing of DSBs formation between Yeast and Drosophila.

The first model of recombination (Holliday Model and Meselson-Radding Model) stipulated that recombination initiates via single-stranded DNA breaks [START_REF] Holliday | The Induction of Mitotic Recombination by Mitomycin C in Ustilago and Saccharomyces[END_REF][START_REF] Meselson | A general model for genetic recombination[END_REF]. In 1981 it was observed that yeast transformation is stimulated up to 3000fold by introducing a DSB into a plasmid [START_REF] Orr-Weaver | Yeast transformation: a model system for the study of recombination[END_REF]. Based on these experiments the double-strand break repair model was proposed to be the actual method for meiotic recombination. Today, the most accepted model for recombination is the double strand break (DSB) repair model [START_REF] Szostak | The double-strand-break repair model for recombination[END_REF].

The DSB model requires breaks to be created in both strands of DNA in one chromatid (Figure 9). These breaks are then resected (exonuclease digested) in a 5' to 3' direction [START_REF] Alani | Analysis of wild-type and rad50 mutants of yeast suggests an intimate relationship between meiotic chromosome synapsis and recombination[END_REF][START_REF] Liu | The location and structure of double-strand DNA breaks induced during yeast meiosis: evidence for a covalently linked DNA-protein intermediate[END_REF]. This results in a single stranded DNA with a free 3' end. This 3' recombinogenic end invades a chromatid on the homologous chromosome, which results in displacement of one of the strands. The displaced strand can then act as a template for DNA polymerase to fill in the gap left after the strand invasion. Each strand is ligated to form double Holliday junctions that appear as cross bridges of single strands of DNA [START_REF] Griffiths | An Introduction to genetic analysis[END_REF]. Another model called SDSA (synthesis-dependent strand annealing) was proposed to account for the variability in gene conversion [START_REF] Allers | Intermediates of yeast meiotic recombination contain heteroduplex DNA[END_REF].

Hotspots and coldspots

In S. cerevisiae, it was found that recombination is not evenly distributed along chromosomes.

Therefore, the DSB events are also not uniformly distributed. The regions that experience high levels of recombination are known as hotspots and are associated with higher levels of DSB formation [START_REF] Gerton | Global mapping of meiotic recombination hotspots and coldspots in the yeast Saccharomyces cerevisiae[END_REF][START_REF] Goldway | A short chromosomal region with major roles in yeast chromosome III meiotic disjunction, recombination and double strand breaks[END_REF][START_REF] Nicolas | An initiation site for meiotic gene conversion in the yeast Saccharomyces cerevisiae[END_REF][START_REF] Sun | Double-strand breaks at an initiation site for meiotic gene conversion[END_REF] while the regions with low levels of recombination (coldspots) show a much lower frequency of DSBs [START_REF] Petes | Meiotic recombination hot spots and cold spots[END_REF]. Despite that all of the factors that determine the location of hotspots are not yet known, some features are common to many hotspots. For example, the incidence of hotspots was found to be correlated with promoters or in other areas of open chromatin, such as those that are transcriptionally active and more sensitive to nucleases [START_REF] Baudat | Chromosome synapsis defects and sexually dimorphic meiotic progression in mice lacking Spo11[END_REF][START_REF] Blat | Physical and functional interactions among basic chromosome organizational features govern early steps of meiotic chiasma formation[END_REF][START_REF] Petes | Meiotic recombination hot spots and cold spots[END_REF][START_REF] Wu | Meiosis-induced double-strand break sites determined by yeast chromatin structure[END_REF]. Few hotspots could be also detected within regions surrounding centromeres, telomeres and other transcriptionally silent areas [START_REF] Blat | Physical and functional interactions among basic chromosome organizational features govern early steps of meiotic chiasma formation[END_REF][START_REF] Gerton | Global mapping of meiotic recombination hotspots and coldspots in the yeast Saccharomyces cerevisiae[END_REF][START_REF] Klein | A central role for cohesins in sister chromatid cohesion, formation of axial elements, and recombination during yeast meiosis[END_REF][START_REF] Lambie | A yeast centromere acts in cis to inhibit meiotic gene conversion of adjacent sequences[END_REF][START_REF] Petes | Meiotic recombination hot spots and cold spots[END_REF]. Recently published evidence indicates that histone modifications such as the tri-methylation of lysine 4 on histone H3 creates the chromatin environment that forms a hotspot [START_REF] Borde | Histone H3 lysine 4 trimethylation marks meiotic recombination initiation sites[END_REF]; this observation, may simply account for the already known association between the hotspots and the transcriptionally active regions [START_REF] Petes | Meiotic recombination hot spots and cold spots[END_REF]. Alternatively, the junction can be dissolved by double Holliday junction dissolution to form a noncrossover [START_REF] Youds | The choice in meiosis -defining the factors that influence crossover or noncrossover formation[END_REF].

Proteins required for DSB formation

Several key proteins have been identified to be required for the formation of DSBs. In Saccharomyces cerevisiae, ten proteins were found to be essential for DSB formation, these proteins include: Mre11, Rad50, andXrs2, Ski8, Rec102, Rec104, Rec114, Mei4, Mer2, and Spo11 [START_REF] Keeney | Mechanism and control of meiotic recombination initiation[END_REF]. Mutations in any one of these ten recombination initiation genes confer similar phenotypes which involve a complete elimination of DSB formation. The four early recombination genes RAD50, XRS2, MRE11, and SKI8 (also known as REC103)

function both in mitosis and meiosis [START_REF] Keeney | Mechanism and control of meiotic recombination initiation[END_REF]. Three of these genes form the MRE11-RAD50-XRS2/ Nbs1 (MRX/N complex) [START_REF] Andrews | MRX (Mre11/Rad50/Xrs2) mutants reveal dual intra-S-phase checkpoint systems in budding yeast[END_REF][START_REF] Chamankhah | Formation of the yeast Mre11-Rad50-Xrs2 complex is correlated with DNA repair and telomere maintenance[END_REF][START_REF] Usui | Functional regulation of the Mre11/Rad50/Xrs2 complex in the processes of recombination and DSB repair[END_REF].

i. MRE11-RAD50-XRS2/ Nbs1 (MRX/N complex)
After DSB formation, the 5' ends of the break are resected to produce 3' single stranded overhangs. Resection is carried out by the MRX/N complex which comprises MRE11 [START_REF] Puizina | Mre11 deficiency in Arabidopsis is associated with chromosomal instability in somatic cells and Spo11-dependent genome fragmentation during meiosis[END_REF], RAD50 [START_REF] Bleuyard | Meiotic defects in the Arabidopsis rad50 mutant point to conservation of the MRX complex function in early stages of meiotic recombination[END_REF] and XRS2/NBS1 [START_REF] Waterworth | NBS1 is involved in DNA repair and plays a synergistic role with ATM in mediating meiotic homologous recombination in plants[END_REF].

ii. Ski8 protein

The fourth gene is Ski8, which was originally characterized for its role in degrading virus dsRNAs. Its loss of function mutation results in increased viral expression, and its name indicates "superkiller". SKI8 is found to be involved in mitotic mRNA translation and stability [START_REF] Brown | The yeast antiviral proteins Ski2p, Ski3p, and Ski8p exist as a complex in vivo[END_REF][START_REF] Searfoss | 3' poly(A) is dispensable for translation[END_REF][START_REF] Wickner | Mutants of the killer plasmid of Saccharomyces cerevisiae dependent on chromosomal diploidy for expression and maintenance[END_REF]. The role of Ski8 in mitotic RNA metabolism involves modulating 3' to 5' exonucleolytic degradation of damaged mRNAs that are not poly-adenylated [START_REF] Frischmeyer | An mRNA surveillance mechanism that eliminates transcripts lacking termination codons[END_REF][START_REF] Searfoss | 3' poly(A) is dispensable for translation[END_REF]. In meiosis, however, it has been proposed to function as a scaffold protein with a role in assembling the DSB initiation complex Spo11-Rec102-Rec104 [START_REF] Araki | Ski7p G protein interacts with the exosome and the Ski complex for 3'-to-5' mRNA decay in yeast[END_REF]. Ski8 migrates from the cytosol to the nucleus during meiosis and it has been found to specifically localize to chromosomes during prophase I [START_REF] Arora | Antiviral protein Ski8 is a direct partner of Spo11 in meiotic DNA break formation, independent of its cytoplasmic role in RNA metabolism[END_REF].

The products of the remaining six early recombination genes (Rec102, Rec104, Rec114, Rec107 [also known as Mer2], Mei4, and Spo11) function specifically in meiosis [START_REF] Keeney | Mechanism and control of meiotic recombination initiation[END_REF][START_REF] Malone | Isolation of mutants defective in early steps of meiotic recombination in the yeast Saccharomyces cerevisiae[END_REF][START_REF] Malone | Recombinationless meiosis in Saccharomyces cerevisiae[END_REF][START_REF] Roeder | Meiotic chromosomes: it takes two to tango[END_REF][START_REF] Weber | A RAD9-dependent checkpoint blocks meiosis of cdc13 yeast cells[END_REF].

iii. SPO11

Spo11 has been shown to be indispensable for meiotic recombination in various species including Schizosaccharomyces pombe [START_REF] Lin | Transient, meiosis-induced expression of the rec6 and rec12 genes of Schizosaccharomyces pombe[END_REF], Arabidopsis thaliana [START_REF] Grelon | AtSPO11-1 is necessary for efficient meiotic recombination in plants[END_REF], Drosophila melanogaster [START_REF] Mckim | mei-W68 in Drosophila melanogaster encodes a Spo11 homolog: evidence that the mechanism for initiating meiotic recombination is conserved[END_REF],

Caenorhabditis elegans [START_REF] Dernburg | Meiotic recombination in C. elegans initiates by a conserved mechanism and is dispensable for homologous chromosome synapsis[END_REF], Mus musculus [START_REF] Baudat | Chromosome synapsis defects and sexually dimorphic meiotic progression in mice lacking Spo11[END_REF] and in a variety of fungi including Neurospora crassa [START_REF] Bowring | Chromosome pairing and meiotic recombination in Neurospora crassa spo11 mutants[END_REF], Sordaria macrospora [START_REF] Storlazzi | Meiotic double-strand breaks at the interface of chromosome movement, chromosome remodeling, and reductional division[END_REF] and Coprinus cinereus [START_REF] Celerin | Multiple roles of Spo11 in meiotic chromosome behavior[END_REF][START_REF] Merino | Replication-dependent early meiotic requirement for Spo11 and Rad50[END_REF].

SPO11 mutations in all these systems result in reduced recombination, sterility and/or reduced viability of gametes. Spo11 was shown to be homologous to topoisomerase VI, a subunit of archaebacterial type II topoisomerase [START_REF] Bergerat | An atypical topoisomerase II from Archaea with implications for meiotic recombination[END_REF]. It was shown to be responsible for the catalysis of DNA breaks [START_REF] Keeney | Meiosis-specific DNA double-strand breaks are catalyzed by Spo11, a member of a widely conserved protein family[END_REF][START_REF] Neale | Endonucleolytic processing of covalent protein-linked DNA doublestrand breaks[END_REF]. The catalytic activity in Saccharomyces Spo11 is presumed to require tyrosine 135 [START_REF] Keeney | Meiosis-specific DNA double-strand breaks are catalyzed by Spo11, a member of a widely conserved protein family[END_REF][START_REF] Prieler | The control of Spo11's interaction with meiotic recombination hotspots[END_REF]. This tyrosine is substituted by a phenylalanine residue in some protists [START_REF] Malik | Protist homologs of the meiotic Spo11 gene and topoisomerase VI reveal an evolutionary history of gene duplication and lineagespecific loss[END_REF][START_REF] Ramesh | A phylogenomic inventory of meiotic genes; evidence for sex in Giardia and an early eukaryotic origin of meiosis[END_REF]. Tyrosine and phenylalanine are both aromatic ringcontaining amino acid residues; it has been proposed that this phenylalanine residue should perform the same biochemical activity as tyrosine [START_REF] Diaz | Identification of residues in yeast Spo11p critical for meiotic DNA double-strand break formation[END_REF][START_REF] Henderson | Tying synaptonemal complex initiation to the formation and programmed repair of DNA double-strand breaks[END_REF].

Although SPO11 encodes the key protein required for making the DSB, it is not sufficient to activate DSBs on its own, as proved by many studies at least 9 additional proteins are needed.

These proteins include Mre11, Rad50, Xrs2, Mer2, Mei2, Rec102, Rec104 and Rec114 [START_REF] Grelon | AtSPO11-1 is necessary for efficient meiotic recombination in plants[END_REF][START_REF] Keeney | Mechanism and control of meiotic recombination initiation[END_REF][START_REF] Pecina | Targeted stimulation of meiotic recombination[END_REF].

iv. Rec104, Rec102, Rec114, Mei4, and Rec107 recombination proteins

Another family of recombination initiation proteins, despite they are absolutely required for DSB formation have specific functions that are largely unknown. This group includes Rec104, Rec102, Rec114, Mei4, and Rec107. Rec102 was found to mediate the Rec104 phosphorylation during meiosis, Rec104 protein having been shown to interact with the SC axial element protein Hop1 and to be required for mature SC formation [START_REF] Hollingsworth | A conditional allele of the Saccharomyces cerevisiae HOP1 gene is suppressed by overexpression of two other meiosis-specific genes: RED1 and REC104[END_REF][START_REF] Kee | Spatial organization and dynamics of the association of Rec102 and Rec104 with meiotic chromosomes[END_REF]). Rec102 itself is phosphorylated but neither the kinase responsible nor the timing for this activation are presently known [START_REF] Shcherbik | A single PXY motif located within the carboxyl terminus of Spt23p and Mga2p mediates a physical and functional interaction with ubiquitin ligase Rsp5p[END_REF].

Rec102 and Rec104 also associate with the DNA on meiotic chromosomes and each requires the presence of the other for the complete Rec102/Rec104 complex loading. The localization of Rec102 and Rec104 to chromatin also requires Spo11 and Ski8 [START_REF] Kee | Spatial organization and dynamics of the association of Rec102 and Rec104 with meiotic chromosomes[END_REF]. Rec107

protein was found to show high abundance during meiotic prophase, in a Cdc7/Dbf4-dependent manner, and phosphorylation on residues Ser11, Ser15, Ser19, Ser22, Ser29 [START_REF] Sasanuma | Cdc7-dependent phosphorylation of Mer2 facilitates initiation of yeast meiotic recombination[END_REF].

Rec114 and Mei4 can associate with DSBs hotspots in a Spo11-independent manner suggesting that they may be the first to bind to the DNA. This binding is mediated by the Cdc7 phosphorylation of Rec107 protein [START_REF] Sasanuma | Cdc7-dependent phosphorylation of Mer2 facilitates initiation of yeast meiotic recombination[END_REF]. Rec114 was suggested to have a role in attracting the late recombination factors [START_REF] Bishop | High copy number suppression of the meiotic arrest caused by a dmc1 mutation: REC114 imposes an early recombination block and RAD54 promotes a DMC1-independent DSB repair pathway[END_REF]. Dmc1 is a homolog of the E. coli protein RecA that is involved in strand invasion during the later events of recombination [START_REF] Bishop | DMC1: a meiosis-specific yeast homolog of E. coli recA required for recombination, synaptonemal complex formation, and cell cycle progression[END_REF].

v. Role of DDK complex in meiotic progression

The catalytic component of the DDK (Dbf4-dependent Cdc7 kinase) complex is Cdc7 which phosphorylates and activates pre-replication components such as MCM helicase proteins during mitosis and meiosis [START_REF] Sclafani | Cdc7p-Dbf4p becomes famous in the cell cycle[END_REF]. A possible link was suggested between DDK function and recombination initiation. Yeast mutants expressing only 15% of wild type levels of DDK display DSBs abolishment, despite they produced live, diploid, dyad spores [START_REF] Matos | Dbf4-dependent CDC7 kinase links DNA replication to the segregation of homologous chromosomes in meiosis I[END_REF]. Similar phenotypes have been observed in spo11 spo13 and rec104 spo13 double mutant cells (Klapholz and Esposito, 1980a;Klapholz and Esposito, 1980b;[START_REF] Malone | Isolation of mutants defective in early steps of meiotic recombination in the yeast Saccharomyces cerevisiae[END_REF]. This suggests DDK has another function after pre-meiotic replication to enable the formation of DSBs and to establish the monopolar spindle attachment required for the reductional division [START_REF] Matos | Dbf4-dependent CDC7 kinase links DNA replication to the segregation of homologous chromosomes in meiosis I[END_REF][START_REF] Rabitsch | Kinetochore recruitment of two nucleolar proteins is required for homolog segregation in meiosis I[END_REF][START_REF] Toth | Functional genomics identifies monopolin: a kinetochore protein required for segregation of homologs during meiosis i[END_REF]. These experiments suggest that there is coordination between meiotic S-phase and recombination, but the exact mechanism, which coordinates these two critical events, is still not understood.

vi. Chromosome cohesion and homologous recombination

Many studies suggest the presence of a strong relationship between axis formation and recombination in many systems [START_REF] Kleckner | Chiasma formation: chromatin/axis interplay and the role(s) of the synaptonemal complex[END_REF]. Many recombination complexes are physically associated to the axis [START_REF] Blat | Physical and functional interactions among basic chromosome organizational features govern early steps of meiotic chiasma formation[END_REF][START_REF] Carpenter | Electron microscopy of meiosis in Drosophila melanogaster females: II. The recombination nodule--a recombination-associated structure at pachytene[END_REF][START_REF] Moens | Initiation and resolution of interhomolog connections: crossover and non-crossover sites along mouse synaptonemal complexes[END_REF][START_REF] Tesse | Localization and roles of Ski8p protein in Sordaria meiosis and delineation of three mechanistically distinct steps of meiotic homolog juxtaposition[END_REF]. This physical association is thought to allow error-free coordination of chromosome recombination events. As prophase I progresses from early to mid pachytene, crossover complexes retain SC association whereas non-crossover complexes are released from the SC [START_REF] Moens | Initiation and resolution of interhomolog connections: crossover and non-crossover sites along mouse synaptonemal complexes[END_REF][START_REF] Terasawa | Meiotic recombination-related DNA synthesis and its implications for cross-over and non-cross-over recombinant formation[END_REF]. Moreover, in Arabidopsis for example, the DSB formation takes place only when the axes are formed, suggesting a temporal link between these two processes [START_REF] Sanchez-Moran | ASY1 mediates AtDMC1-dependent interhomolog recombination during meiosis in Arabidopsis[END_REF].

vii. Synaptonemal complex proteins mediate meiotic recombination

During prophase I, homologous recombination manifests in form of developing recombination protein complexes (known as recombination nodules) that are closely associated with AEs of meiocytes [START_REF] Carpenter | Electron microscopy of meiosis in Drosophila melanogaster females: II. The recombination nodule--a recombination-associated structure at pachytene[END_REF][START_REF] Moens | Initiation and resolution of interhomolog connections: crossover and non-crossover sites along mouse synaptonemal complexes[END_REF]. In mammals, about 250-300 early recombination nodules (ENs) are associated with the AEs before synapsis is initiated [START_REF] Fraune | The mammalian synaptonemal complex: protein components, assembly and role in meiotic recombination[END_REF]. This observation gives a clear evidence for the close interdependency and the tight temporal correlation between the process of homologous recombination and the assembly of synaptonemal complex.

In budding yeast, mutations of the meiosis specific axis components Red1, Hop1 or the Rec8 cohesin subunit, result in an altered distribution of DSBs implying their essential role in the initiation and positioning of DSBs [START_REF] Blat | Physical and functional interactions among basic chromosome organizational features govern early steps of meiotic chiasma formation[END_REF][START_REF] Glynn | Genome-wide mapping of the cohesin complex in the yeast Saccharomyces cerevisiae[END_REF]. In yeast red1 mutants, AEs are not detectable and recombination is severely defective [START_REF] Rockmill | Meiosis in asynaptic yeast[END_REF][START_REF] Xu | Meiotic cells monitor the status of the interhomolog recombination complex[END_REF]. The recombination which does occur in these mutants is biased towards the sister chromatid, implying that Red1 mediates homologue bias in recombination [START_REF] Xu | Meiotic cells monitor the status of the interhomolog recombination complex[END_REF]. The Mek1 kinase localizes to the binding sites of Red1 and Hop1 and phosphorylates Red1 [START_REF] Bailis | Synaptonemal complex morphogenesis and sister-chromatid cohesion require Mek1-dependent phosphorylation of a meiotic chromosomal protein[END_REF]. It has recently been proposed that the role of Red1/Mek1 (and presumably Hop1) in establishing inter-homologues bias maybe to counteract the inter-sisters bias imposed by Rec8 and its role in sister chromatid cohesion [START_REF] Kim | Sister cohesion and structural axis components mediate homolog bias of meiotic recombination[END_REF].

B. DSBs repair

SPO11 induced DSBs trigger the meiotic equivalent DNA-damage response, which involves sensing of breaks, recruitment of repair proteins and processing of recombination intermediates (Figure 9). Because DSB repair is a fundamental process in both somatic and meiotic cells, homologous recombination repair (Hrr) proteins are the most conserved group of meiotic recombination proteins [START_REF] Handel | Genetics of mammalian meiosis: regulation, dynamics and impact on fertility[END_REF]. DSBs repair is initiated by the removal of Spo11 from meiotic DSB ends. In yeast, the process is mediated by Sae2 and Mre11-Rad50-Xrs2-dependent endonucleolysis that releases Spo11 bound to a short nucleotide sequence [START_REF] Neale | Clarifying the mechanics of DNA strand exchange in meiotic recombination[END_REF][START_REF] Neale | Endonucleolytic processing of covalent protein-linked DNA doublestrand breaks[END_REF]. Once Spo11 is removed, the 5′-ending strands are degraded to expose the 3′-ending single-stranded DNA (ssDNA).

These tails, invading an intact homologous DNA duplex, are used as primers for DNA synthesis (La [START_REF] Volpe | Meiotic double strand breaks repair in sexually reproducing eukaryotes: we are not all equal[END_REF]. Strand invasion of homologous chromosome by the 3′ single strand is mediated by RAD51 and/or the meiosis-specific DMC1 [START_REF] Masson | The meiosis-specific recombinase hDmc1 forms ring structures and interacts with hRad51[END_REF][START_REF] Masson | The Rad51 and Dmc1 recombinases: a non-identical twin relationship[END_REF]; the ssDNA-binding (SSB) protein, replication protein A (RPA), is an essential cofactor in this process [START_REF] Kantake | The recombination-deficient mutant RPA (rfa1-t11) is displaced slowly from single-stranded DNA by Rad51 protein[END_REF][START_REF] Wang | Role of Saccharomyces single-stranded DNA-binding protein RPA in the strand invasion step of double-strand break repair[END_REF]. RPA is a heterotrimeric protein involved in numerous DNA metabolic pathways including replication, repair, and recombination [START_REF] Anciano Granadillo | Targeting the OB-Folds of Replication Protein A with Small Molecules[END_REF]. It binds single-stranded DNA (ssDNA) during homologous recombination through interactions with a series of oligonucleotide/oligosaccharide binding-folds (OB-folds) that display a high affinity for ssDNA. This prevents DNA from winding back on itself allowing repair and prophase to occur properly [START_REF] Bochkarev | From RPA to BRCA2: lessons from single-stranded DNA binding by the OB-fold[END_REF]Joyce and McKim, 2009). Rad51 was reported to facilitate the formation of a physical filament connection between the invading DNA substrate and homologous duplex DNA template, leading to the generation of heteroduplex DNA (D-loop). After DNA is synthesized using the invading end as a primer, Rad51 dissociates from dsDNA to ensure DNA synthesis by exposing the 3'OH [START_REF] Krejci | Homologous recombination and its regulation[END_REF].

III. Drosophila melanogaster as a model for meiotic studies

It is important to mention that in Drosophila, males and females use very different strategies to segregate their chromosomes, as meiotic recombination is specific for female meiosis.

Drosophila female meiosis is an attractive model system to study the early events of meiosis because of powerful tools in genetics and cytology that can be used to identify and characterize the genes required for meiotic recombination [START_REF] Mehrotra | Synapsis, double-strand breaks, and domains of crossover control in drosophila females[END_REF]. Moreover, the ovary is arranged in accordance with developmental stage (Figure 10). The females reproductive system is represented by a pair of ovaries, each consisting of 14-16 reproductive units called ovarioles; each ovariole contains a chain of developing egg chambers that become more mature as they reach the posterior end of the ovary. At the anterior tip of each ovariole is the germarium which is divided morphologically into four regions: region 1, region 2A, region 2B and region 3 [START_REF] King | Ovarian development in Drosophila melanogaster[END_REF]. The germline stem cells reside in the anterior-most part of region 1, each stem cell divides asymmetrically to generate a cystoblast. The cystoblast undergoes four rounds of synchronized mitotic cell divisions with incomplete cytokinesis to produce a 16-cell interconnected cyst. All 16 cells of the cyst share the same cytoplasm through bridge structures called ring canals which ensure the exchange of intercellular material between the nutritive cells and the oocyte [START_REF] Robinson | Morphogenesis of Drosophila ovarian ring canals[END_REF].

The events of early meiotic prophase can be observed cytologically in context of oocyte development within the germarium (Figure 11). Region 1 is concerned with the pre-meiotic DNA replication phase, after this, prophase and recombination are initiated [START_REF] King | Ovarian development in Drosophila melanogaster[END_REF][START_REF] Mehrotra | Temporal analysis of meiotic DNA double-strand break formation and repair in Drosophila females[END_REF][START_REF] Mehrotra | Synapsis, double-strand breaks, and domains of crossover control in drosophila females[END_REF].

Prophase is initiated by pachytene. Early pachytene involves two processes: the full-length assembly of SC and DSB formation while DSBs repair occurs at mid-pachytene. Therefore, the three pachytene stages could be assigned to the three regions of the germarium: 2A, 2B, and 3 [START_REF] Mehrotra | Synapsis, double-strand breaks, and domains of crossover control in drosophila females[END_REF]. In region 2A, up to four cells within the cyst initiate assembly of the SC. However, within the cyst, only two pro-oocytes have the potential to develop into a mature oocyte and enter into pachytene and assemble mature SC. Eventually, as pachytene progresses, one pro-oocyte of the two cells disassembles the SC and becomes a nurse cell.

The proocyte that is selected to become the oocyte localizes at the posterior end of the developing egg chamber. Throughout regions 2A and 2B most of the oocytes are in pachytene, with the SC assembled between homologs along their entire lengths.

Figure 11: Oogenesis in Drosophila

The females reproductive system in Drosophila is represented by a pair of ovaries, each consisting of 14-16 reproductive units called ovarioles; each ovariole contains a chain of developing egg chambers that become more mature as they reach the posterior end of the ovary. At the anterior tip of each ovariole is the germarium where the events of early meiotic prophase can be observed. The remainder of the ovariole (stages 2-14) is termed the vitellarium. At the anterior tip of the germarium, a germline stem cell divides to produce a cystoblast (premeiotic region 1). The cystoblast undergoes four rounds of mitotic divisions to produce a 16-cell germline cyst. Within the cyst, up to four cells can initiate the formation of the synaptonemal complex (SC) (red) in zygotene, but only two prooocytes continue into meiosis and form a full-length SC in early pachytene (region 2A). At this stage programmed meiotic DSBs are induced and can be detected with an antibody to a phosphorylated form of a histone H2A variant, H2AV, at serine 137 (γ-H2AV) (blue circles denote DSBs detected with a γ-H2AV antibody). By early/midpachytene (region 2B), fewer DSBs (γ-H2AV foci) can be detected within the pro-oocytes. By mid-pachytene (region 3), one cell has been selected to become the oocyte [START_REF] Lake | The molecular control of meiotic chromosomal behavior: events in early meiotic prophase in Drosophila oocytes[END_REF].

By region 3, oocyte determination has been completed since a single oocyte is usually evident and positioned toward the posterior of the cyst [START_REF] Mehrotra | Synapsis, double-strand breaks, and domains of crossover control in drosophila females[END_REF].

A. Pairing of homologous chromosomes (synapsis) in Drosophila

In Drosophila, a pair of pro-oocytes enters in prophase at region 2A. Only one of them is selected by region 3 and defined by the presence of SC which is detected by antibodies against C(3)G transverse element (Figure 11) [START_REF] Page | c(3)G encodes a Drosophila synaptonemal complex protein[END_REF]. C(3)G is a coiled-coil protein similar to proteins in budding yeast (ZIP1), C. elegans (SYP-1, SYP-2), and mammals (SYCP1) [START_REF] Page | The genetics and molecular biology of the synaptonemal complex[END_REF][START_REF] Watts | SUMO meets meiosis: an encounter at the synaptonemal complex: SUMO chains and sumoylated proteins suggest that heterogeneous and complex interactions lie at the centre of the synaptonemal complex[END_REF]. Zygotene pro-oocytes were identified by their patchy C(3)G staining, as opposed to the thread-like staining typical of pachytene (Figure 11). SC initiation was found to be linked strongly with cohesion proteins and has interesting parallels with synapsis initiation in budding yeast [START_REF] Tanneti | A pathway for synapsis initiation during zygotene in Drosophila oocytes[END_REF].

The cohesion proteins components SMC1 and SMC3 were found to interact with C(2)M [START_REF] Heidmann | The Drosophila meiotic kleisin C(2)M functions before the meiotic divisions[END_REF]. C(2)M protein is a distantly related member of the kleisin family that includes Rec8 and Rad21 paralogues [START_REF] Schleiffer | Kleisins: a superfamily of bacterial and eukaryotic SMC protein partners[END_REF]. It was shown to be a lateral element component of the synaptonemal complex [START_REF] Anderson | Juxtaposition of C(2)M and the transverse filament protein C(3)G within the central region of Drosophila synaptonemal complex[END_REF]. In wild-type, C(2)M was found to co-localize with C(3)G in most locations to the exception of the centromeres [START_REF] Khetani | Regulation of meiotic cohesion and chromosome core morphogenesis during pachytene in Drosophila oocytes[END_REF][START_REF] Tanneti | A pathway for synapsis initiation during zygotene in Drosophila oocytes[END_REF].

Ord is a meiosis-specific protein that localizes to chromosome arms and centromeres. It is required for chromosome cohesion and crossover formation [START_REF] Anderson | Juxtaposition of C(2)M and the transverse filament protein C(3)G within the central region of Drosophila synaptonemal complex[END_REF][START_REF] Balicky | Meiotic cohesion requires accumulation of ORD on chromosomes before condensation[END_REF][START_REF] Khetani | Regulation of meiotic cohesion and chromosome core morphogenesis during pachytene in Drosophila oocytes[END_REF][START_REF] Webber | The cohesion protein ORD is required for homologue bias during meiotic recombination[END_REF]. In ord mutant oocytes, the centromeres clustering is defective and the association of SC proteins with the centromeres is disrupted [START_REF] Tanneti | A pathway for synapsis initiation during zygotene in Drosophila oocytes[END_REF]. Although Ord is found to be required for the cohesin accumulation onto centromeres, it is not essential for the initial loading of the cohesin subunits Smc1 and Smc3 to chromosome arms [START_REF] Khetani | Regulation of meiotic cohesion and chromosome core morphogenesis during pachytene in Drosophila oocytes[END_REF][START_REF] Lake | The molecular control of meiotic chromosomal behavior: events in early meiotic prophase in Drosophila oocytes[END_REF].

B. Initiation of double strand breaks in Drosophila

The phosphorylated form of the H2Av variant (γ-H2Av) forms foci that have been used as markers for DSBs. By using an antibody that recognizes γ-H2AV at DSB sites, Mehrotra &

McKim showed that γ-H2AV foci first appear in early pachytene after SC formation [START_REF] Mehrotra | Temporal analysis of meiotic DNA double-strand break formation and repair in Drosophila females[END_REF]. By counting the number of γ-H2AV foci, a gradual increase in the number of DSBs has been noticed as cysts mature in region 2A (Figure 11). This increase is followed by a decline in number as the cyst progresses from early to mid-pachytene (region 2B). The γ-H2AV foci virtually disappear in the oocyte by the time the cyst reaches midpachytene (region 3). Because the gradual decrease in the number of γ-H2AV foci coincides with progression through pachytene, this decrease likely represents the process of DSB repair [START_REF] Lake | The molecular control of meiotic chromosomal behavior: events in early meiotic prophase in Drosophila oocytes[END_REF]. Whether the removal of γ-H2AV indicates that repair is complete is still unknown.

In Drosophila, in addition to the Spo11 ortholog Mei-W68 [START_REF] Mckim | mei-W68 in Drosophila melanogaster encodes a Spo11 homolog: evidence that the mechanism for initiating meiotic recombination is conserved[END_REF], Mei-P22 is reported to be required for DSB formation (Figure 12).

Immunofluorescence analysis of an epitope-tagged Mei-P22 showed that during early pachytene after SC formation, Mei-P22 localizes to discrete foci prior to the time DSBs can be detected [START_REF] Liu | mei-P22 encodes a chromosome-associated protein required for the initiation of meiotic recombination in Drosophila melanogaster[END_REF][START_REF] Mehrotra | Temporal analysis of meiotic DNA double-strand break formation and repair in Drosophila females[END_REF]. The percentage of Mei-P22 foci showed a partial overlap with the γ-H2AV marker [START_REF] Mehrotra | Temporal analysis of meiotic DNA double-strand break formation and repair in Drosophila females[END_REF].

Interestingly, in the absence of mei-W68, Mei-P22 is able to load onto the chromosomes, which indicates that Mei-P22 localization is independent of DSB formation [START_REF] Liu | mei-P22 encodes a chromosome-associated protein required for the initiation of meiotic recombination in Drosophila melanogaster[END_REF].

A third gene required for the induction of meiotically induced DSBs is trem [START_REF] Lake | The Drosophila zinc finger protein trade embargo is required for double strand break formation in meiosis[END_REF]. Trade embargo (trem), a 5-C2H2 zinc finger protein is expressed in early zygotene, localizes to chromatin in a thread-like fashion and is required for normal DSB levels. A point mutation in trem (tremF9) failed to localize Mei-P22 to discrete foci during pachytene.

Because Mei-P22 can associate with chromatin in the absence of DSBs, these data suggest that Trem acts upstream of both Mei-P22 and Mei-W68 in the initiation of DSBs. The mechanism by which trem localizes Mei-P22 to discrete foci on the chromatin is still unknown [START_REF] Lake | The molecular control of meiotic chromosomal behavior: events in early meiotic prophase in Drosophila oocytes[END_REF].

C. Repair of double strand breaks in Drosophila

In Drosophila, the DSBs repair genes process the recombination intermediates that can be resolved into either non-crossover (gene conversion) or crossover products [START_REF] Haber | Recombination: a frank view of exchanges and vice versa[END_REF]. In order to make crossovers, two classes of genes named (crossover-specific genes) are required (Joyce and McKim, 2009).

The first class is precondition genes (Figure 12) that are responsible for determining which DSBs become crossovers and their distribution such as (mei-218 and rec) [START_REF] Bhagat | Studies on crossover-specific mutants and the distribution of crossing over in Drosophila females[END_REF]Joyce and McKim, 2009;[START_REF] Liu | Two genes required for meiotic recombination in Drosophila are expressed from a dicistronic message[END_REF]. The other class is called exchange genes, which produce crossovers (Figure 12). Four proteins have been identified in the exchange class: MEI-9, ERCC1, MUS312 and HDM (Joyce et al., 2009). Mutations in mei-9, Ercc1, mus312, and hdm result in a reduced and randomly distributed crossing over along chromosomes (Joyce et al., 2009). Mei-9 is the Drosophila homolog of human and yeast nucleotide excision repair (NER) [START_REF] Sekelsky | The Drosophila meiotic recombination gene mei-9 encodes a homologue of the yeast excision repair protein Rad1[END_REF][START_REF] Sijbers | Xeroderma pigmentosum group F caused by a defect in a structure-specific DNA repair endonuclease[END_REF]. Mei-9 is required for 90% of all meiotic crossovers [START_REF] Boyd | The mei-9 alpha mutant of Drosophila melanogaster increases mutagen sensitivity and decreases excision repair[END_REF]. The HDM protein encodes a RPA-like protein (Joyce et al., 2009).

Drosophila has two checkpoint pathways that are activated to monitor delays with different meiotic events. The first is the ATR/MEI-41-dependent DSB repair checkpoint, which works when there is a defect in repairing DSBs (Joyce et al., 2011). When there are mutations in any of the DSB repair such as precondition genes or exchange genes, pachytene checkpoint delays the chromatin remodeling response to DSBs (Joyce and McKim, 2009).

The first part of my thesis work is dedicated to the characterization of Yemanuclein as a new actor in Drosophila meiotic recombination. We show that Yemanuclein is associated to the SC and that it interacts in vivo with cohesin complex components. Interestingly yemanuclein was not recovered in the numerous genetic screens that were undertaken to identify mutations that affect meiotic recombination.

Chapter 2: Chromatin reprogramming in sexual reproduction

The genetic information of a eukaryotic cell is encoded by long linear duplex polymers of deoxyribonucleic acid (DNA). DNA comprises huge information ranging from 10 6 to 10 11 base pairs (bp). This "database" is confined in about "2 meter length library" occupying only 5-20 µm in the cell nucleus [START_REF] Widom | Structure, dynamics, and function of chromatin in vitro[END_REF]. In order to fit this small space, the DNA filaments undergo tremendous compaction by wrapping around highly basic proteins called histones. The resulting DNA-histone complexes are nucleosomes, which are the building blocks of the macro-nucleoprotein structure called chromatin, which controls not only genetic inheritance, but also the activity of genes [START_REF] Kornberg | Twenty-five years of the nucleosome, fundamental particle of the eukaryote chromosome[END_REF].

I. Chromatin structure A. Chromatin organization and genome dynamics

The nucleosomes constitute the first level of chromatin compaction and this contributes a relatively small fraction of the condensation needed to fit the typical genome into an interphase nucleus indicating that there are additional "higher order" levels of chromatin condensation (Figure 13). 'Higher-order' structures are poorly understood, but are known to rely on the interactions between nucleosomes and include additional assemblage of nucleosomes that assumes a reproducible conformation in 3D space [START_REF] Woodcock | Chromatin higher-order structure and dynamics[END_REF].

Chromatin organization has a major effect on DNA readout, since the packaged DNA has to be made accessible for transcription, replication and DNA repair to occur properly. Moreover, some regions of the genome are differently accessible than others. Such differences in compaction state, at the same time, provide an opportunity for differential regulation of gene expression in programming different cell types that have to be remembered by a cell through multiple divisions.

DNA undergoes multiple levels of compaction that change over the course of the cell cycle (Figure 13). In interphase, chromosomes have a decondensed appearance, and occupy spatially distinct regions in the nucleus termed 'chromosome territories' [START_REF] Cremer | Chromosome territories, nuclear architecture and gene regulation in mammalian cells[END_REF]. The use of DNA coloration dyes revealed that chromatin in interphase can be visualized at microscopic level into two relatively distinct forms, euchromatin and heterochromatin [START_REF] Fransz | Heterochromatin in interphase nuclei of Arabidopsis thaliana[END_REF]. Euchromatin is characterized by a loose decondensed chromatin. The DNA is less densely packed during interphase and here the genes are in a so-called permissive state for transcription. While, heterochromatins are initially referred as those chromatin regions that remain densely stained and are highly condensed.

Heterochromatin can be either constitutive or facultative. Constitutive heterochromatin consists of non-coding and repetitive DNA sequences such as those present in the centromeric, pericentromeric and sub-telomeric regions. They persist in silenced state through cell divisions [START_REF] Probst | Epigenetic inheritance during the cell cycle[END_REF]. Facultative heterochromatin on the other hand is not a permanent feature but is seen in some cells at certain developmental stages. It may result from transformation of euchromatin via specific epigenetic alterations at certain times, and contains loci activated at specific stages while repressed at others [START_REF] Brown | Genomes[END_REF].

The concentration of DNA in different nuclear regions was found to be inversely correlated with the level of transcription, leading to the idea that gene expression is suppressed in heterochromatin. Recently, heterochromatin studies have led to a dramatic advance in Nucleosomes are connected to one another by short stretches of linker DNA. At the next level of organization the string of nucleosomes is folded into a fibre about 30 nm in diameter, and these fibers are then further folded into higher-order structures. At levels of structure beyond the nucleosome the details of folding are still uncertain [START_REF] Felsenfeld | Controlling the double helix[END_REF]. B) Chromatin organization within the nucleus. An interphase nucleus seen by electron microscopy; one may distinguish euchromatin (light, transcriptionally active) and heterochromatin (dark, highly condensed, transcriptionally inactive); image taken from [START_REF] Ross | Introduction to molecular medicine[END_REF].

understanding epigenetic control of gene activity [START_REF] Bernstein | RNA meets chromatin[END_REF][START_REF] Lund | Epigenetics and cancer[END_REF]. Nevertheless, the precise molecular interactions and structural changes at chromatin higher-order folding remain barely known [START_REF] Grigoryev | The end adjusts the means: heterochromatin remodelling during terminal cell differentiation[END_REF]. Differential gene expression could be explained by understanding the effect of heterochromatin regions.

Studies in Drosophila revealed a phenomenon known as PEV for "position effect variegation" [START_REF] Reuter | Mutants affecting position-effect heterochromatinization in Drosophila melanogaster[END_REF][START_REF] Reuter | Isolation of dominant suppressor mutations for position-effect variegation in Drosophila melanogaster[END_REF]. PEV is observed when a euchromatic gene is artificially relocated next to heterochromatin, which confers to the gene that comes in close proximity to heterochromatin, variegated expression. Silencing of certain genes may lead to phenotypic variegation in tissues. While the genetic information required for gene activation is present in all cells, the gene is active in a differential sporadic manner, being fully active in some cells, and completely inactive in others. This 'spreading' of repressive features along the chromosome results in a variegated expression pattern that can appear to be clonal or randomly speckled [START_REF] Schotta | SU(VAR)3-9 is a conserved key function in heterochromatic gene silencing[END_REF]. It is worth mentioning that transcriptionally active and silent regions may be positioned adjacent to each other; therefore, boundaries must exist to prevent two neighboring regions from influencing each other. These borders are called insulators and are DNA elements that prevent stimulation of transcription or silencing from one region to another [START_REF] West | Insulators: many functions, many mechanisms[END_REF].

Position-effect variegation is an excellent genetic tool to screen gene functions, many genes now known to be involved in chromatin dynamics have been successfully identified by using the phenomenon of PEV, such as suppressor of variegation 2-5, encoding the heterochromatin protein 1 (HP1) [START_REF] James | Identification of a nonhistone chromosomal protein associated with heterochromatin in Drosophila melanogaster and its gene[END_REF]. It was found that the physical tethering of HP1 to the heterochromatin regions ensures the chromatin condensation [START_REF] Greil | Distinct HP1 and Su(var)3-9 complexes bind to sets of developmentally coexpressed genes depending on chromosomal location[END_REF][START_REF] Verschure | In vivo HP1 targeting causes large-scale chromatin condensation and enhanced histone lysine methylation[END_REF], while paradoxically, via a potential interaction with RNA, HP1 has also been found to localize to some active genes [START_REF] Cryderman | Role of Drosophila HP1 in euchromatic gene expression[END_REF][START_REF] Piacentini | Heterochromatin protein 1 (HP1) is associated with induced gene expression in Drosophila euchromatin[END_REF]. Therefore HP1 may be required for the expression of both its euchromatic and heterochromatic target genes. This broad distribution of HP1 makes it essential for various functions ranging from repressing transcription to long-range chromatin interactions.

B. Nucleosome assembly

1. Histones as the basic components DNA and histones have very high and non-specific affinity to one another. Therefore, the proper assembly of nucleosomes requires that they have to be built in a stepwise fashion. This involves multiple intermediate stages and auxiliary factors (Figure 14). The DNA is first wrapped around two dimers of the histones H3 and H4 that act as a scaffold, it gives the central 80 bp (H3/H4) 2 tetramer of the 147 bp of nucleosomal DNA that wraps into a unit known as tetramer or the tetrasome. The resulting tetrasome structure is then sandwiched between two dimers of histones H2A and H2B and wraps the additional 67 bp of DNA to give rise to the complete nucleosome core particle [START_REF] Luger | Crystal structure of the nucleosome core particle at 2.8 A resolution[END_REF]. Nucleosomes are spaced apart by a stretch of linker DNA that has an average length of ~50 bp. The linker region is bound by the non-core H1 histone, an interaction that contributes to overall compaction of the nucleosomal array. Histones interact with the phosphate backbone of DNA and therefore have no sequence specificity for their binding. The proper spacing of the nucleosomes therefore requires the activity of ATP-driven auxiliary factors known as 'chromatin remodelers' that move the DNA with respect to the histone octamer [START_REF] Cairns | The logic of chromatin architecture and remodelling at promoters[END_REF].

Figure 14: Schematic representation of stepwise assembly of the nucleosome core particle

The core histones are color coded (yellow = H2A, red = H2B, blue = H3, green = H4) and cylinders represent helices [START_REF] Dutnall | Nucleosome assembly: more than electric in the making? Structure[END_REF].

Histones of the nucleosome unit present similar structures containing 2 distinct domains: the histone fold domain formed by three α-helices connected by two loops, and the N-terminal tail domain consisting of 15-30 basic amino acids and protruding out from nucleosome surface [START_REF] Khorasanizadeh | The nucleosome: from genomic organization to genomic regulation[END_REF]. More than 120 direct atomic interactions between histones and DNA backbone, and nearly equal number of water-mediated interactions are distributed at the 14 super-helix locations at the interface of these sub-complexes, while multiple electrostatic and hydrophobic effects and hydrogen bonds are required for nucleosome formation [START_REF] Kamakaka | Histone variants: deviants?[END_REF][START_REF] Luger | Structure and dynamic behavior of nucleosomes[END_REF].

As basic components of the nucleosome, histones have critical contribution in nucleosome structure. It is certain that modification on histone residues or alteration in primary sequence would affect nucleosome dynamics possibly via causing subtle structural variation or recruiting different chromatin-modifying factors, and hence influence the DNA accessibility and thereafter higher architecture.

Nucleosome assembly pathways

Histones assembly onto nucleosomes can be divided into two main pathways, a pathway coupled to DNA synthesis, which includes DNA replication and repair, called replicationcoupled (RC) assembly and a pathway independent of DNA synthesis called replicationindependent (RI) assembly.

i. Replication-coupled assembly

Ahead of the replication fork, nucleosomes must be temporarily disassembled or remodeled in order for the DNA replication machinery to gain access to the DNA. During DNA replication DNA unwinding is accompanied by transient disruption of parental nucleosomes. And following DNA replication during S-phase, the naked DNA is an available target for every DNA binding protein in the cell. Many studies reported that the assembly of replicated DNA into nucleosomes is coupled to the on-going DNA replication [START_REF] Mcknight | Electron microscopic analysis of chromatin replication in the cellular blastoderm Drosophila melanogaster embryo[END_REF][START_REF] Stillman | Chromatin assembly during SV40 DNA replication in vitro[END_REF].

During S phase of the cell cycle, bulk DNA is replicated through a highly orchestrated multiple-component process that results in two copies of the genome. The newly synthesized core essential histones are strictly incorporated into DNA; these histones are so called canonical histones. Canonical histone proteins are encoded by replication-dependent genes and must rapidly reach high levels of expression during S phase. The majority of histone deposition takes place during S-phase and mainly involves the canonical histones. Thus, coupling nucleosome assembly to DNA replication within the cell is a vital process to establish a silent, closed chromatin conformation that helps to prevent aberrant gene expression [START_REF] Loyola | Histone chaperones, a supporting role in the limelight[END_REF]. It ensures proper inheritance of chromatin structure, propagation of epigenetic marks on histones to daughter cells and maintenance of genome integrity [START_REF] Li | All roads lead to chromatin: Multiple pathways for histone deposition[END_REF].

ii. Replication-independent assembly

Independently to the DNA replication and under specific developmental conditions such as DNA repair, meiotic recombination or transcription initiation and termination, large families of histone variants are incorporated to the nucleosomes and alter their structure and dynamics.

The histones that are already present in the chromatin are replaced by the histone variants through the replication-independent assembly pathway. Thus, the presence of such variants in chromatin adds a level of complexity to the regulation of chromatin structure [START_REF] Talbert | Histone variants--ancient wrap artists of the epigenome[END_REF]

II. Regulation of chromatin structure

Chromatin is a highly dynamic structure that must keep the balance between being folded as much as needed and being accessible whenever necessary. The functional state of chromatin is ultimately regulated by the structural changes that make the underlying DNA molecule more or less accessible to the different biological processes such as DNA replication, transcription and DNA repair via its degree of compaction. DNA compaction depends on the status of its associated histones. The deposition of histones on DNA is the fundamental basis for epigenetic inheritance. It ensures that certain chromatin states are maintained inside a single cell and/or transmitted throughout cell divisions to daughter cells. This requires not only proteins that deposit histones on DNA but also proteins or complexes and biochemical processes mediated by many specific enzymes in order to maintain the chromatin modification status. The functional status of chromatin can be modulated through several interconnected regulatory mechanisms:

-incorporation of histone variants -post-translational modifications of N-terminal tails that emerge from the nucleosome -ATP-dependent chromatin remodeling -histone chaperones

A. Histone Variants

Eukaryotes, beside canonical histones, have evolved various histone variants (Figure 15). In general, canonical histone genes have multiple copies that are highly similar in sequence.

Moreover they are intron-less and they are expressed during S-phase. These histones are devoted to the DNA-synthesis process, or so called Replication Coupled (RC) chromatin assembly. They are highly conserved during evolution and are synthesized to provide the main supply of histones at DNA replication. In contrast histone variants or "replacement" histones have evolved from the corresponding canonical histones and differ from their canonical paralogues in primary sequence and the presence of introns in their genes (Zhu et al., 2012). Histone variants expression timing and deposition mechanisms are also different from canonical histones (Table 2). Histone variants are usually expressed throughout the cell cycle and are thus available, at least theoretically, in nucleosome assembly pathways that occur in a replication independent (RI) manner [START_REF] Henikoff | Assembly of variant histones into chromatin[END_REF][START_REF] Sarma | Histone variants meet their match[END_REF]. Histone variants may therefore be incorporated into nucleosomes during the entire cell cycle and can impart unique properties to the nucleosomes they occupy [START_REF] Kamakaka | Histone variants: deviants?[END_REF]. The incorporation of these histone variants is essential for nucleosomes modulation. This variability is important to determine the properties of the chromatin fiber at a local and regional level, with respect to essential aspects of DNA metabolism, such as replication, transcription, heterochromatin formation, DNA repair, condensation or kinetochore formation.

Histone variants may preferentially or specifically be expressed in certain tissues, for instance, the testis specific histone H3 variant [START_REF] Witt | Testis-specific expression of a novel human H3 histone gene[END_REF]. Some others are enriched at specific chromosomes or locations. As an example, CENP-A is an H3 variant that specifically localizes to centromeres [START_REF] Palmer | Purification of the centromere-specific protein CENP-A and demonstration that it is a distinctive histone[END_REF]. Another such variant is macroH2A, which is enriched on the inactive human X-chromosome [START_REF] Chadwick | Cell cycle-dependent localization of macroH2A in chromatin of the inactive X chromosome[END_REF].

Histone variants were discovered on the basis of differences at the level of their primary sequence. Variants have been identified for all histone classes with the exception of histone H4 [START_REF] Brown | Histone variants: are they functionally heterogeneous?[END_REF].

H2A variants

The H2A family (Table 2) contains a plethora of 'universal variants' found in almost all organisms. In mammals four H2A variants have been reported (H2A.Z, H2A.X, marcoH2A and H2A.Bbd) [START_REF] Malik | Phylogenomics of the nucleosome[END_REF].

i. H2A.Z variant

H2A.Z is a highly conserved histone variant; it was found to have a single evolutionary origin different from all other H2A variants [START_REF] Malik | Phylogenomics of the nucleosome[END_REF]. H2A.Z differs from canonical H2A and other H2A variants mainly in its "docking" domain in the C-terminus and in the L1 loop where two H2A molecules contact each other [START_REF] Arents | The nucleosomal core histone octamer at 3.1 A resolution: a tripartite protein assembly and a left-handed superhelix[END_REF][START_REF] Luger | Crystal structure of the nucleosome core particle at 2.8 A resolution[END_REF][START_REF] Suto | Crystal structure of a nucleosome core particle containing the variant histone H2A[END_REF]. Genome-wide nucleosome occupancy studies in yeast (Mavrich et al., 2008a;[START_REF] Yuan | Genome-scale identification of nucleosome positions in S. cerevisiae[END_REF] and flies (Mavrich et al., 2008b) showed a specific localization pattern of H2A.Z nucleosomes near the transcription start site. This has been proposed to be an epigenetic marker for directing or regulating the positioning of downstream nucleosomes.

Therefore, ensuring the correct deposition of H2A.Z nucleosomes is crucial for the maintenance of epigenetic modifications at the transcription start site. [START_REF] Gupta | Predicting human nucleosome occupancy from primary sequence[END_REF][START_REF] Rando | Rules and regulation in the primary structure of chromatin[END_REF]. H2AZ has also been shown to be essential in early embryonic development. Cell differentiation of murine inner cell mass (ICM) revealed that H2A.Z is first enriched at pericentromeric heterochromatin and subsequently enriched at other chromatin regions.

Figure 15: Schematic representation of histone variants

A) Structural domains of canonical histones and histone variants, the regions that differ in histone variants are shown in red (Allis et al., 2007b). B) Genomic locations of some histone variants as identified by immunofluorescence and genome-wide ChIP studies [START_REF] Banaszynski | Histone variants in metazoan development[END_REF].

H2A.Z depletion in mice causes genome instability and disruption of HP1α localization at the pericentromeric regions, which suggests that HP1α function and pericentromeric heterochromatin identity are regulated by H2A.Z during early embryonic development [START_REF] Rangasamy | Pericentric heterochromatin becomes enriched with H2A.Z during early mammalian development[END_REF][START_REF] Rangasamy | RNA interference demonstrates a novel role for H2A.Z in chromosome segregation[END_REF]. Interestingly, the only Drosophila H2A variant, known as H2Av, is paralogous to H2AZ [START_REF] Talbert | A unified phylogeny-based nomenclature for histone variants[END_REF].

ii. H2A.X variant H2A.X has a histone fold domain that is similar to the canonical H2A and differs from it only in having a unique C-terminal motif hydrophobic residue [START_REF] Malik | Phylogenomics of the nucleosome[END_REF][START_REF] Talbert | Histone variants--ancient wrap artists of the epigenome[END_REF]. Despite that H2A.X has been implicated to play a role in meiosis, growth, tumor suppression and immune receptor rearrangements, its role in DNA doublestrand breaks (DSB) repair makes it viewed as the "histone guardian of the genome" [START_REF] Fernandez-Capetillo | H2AX: the histone guardian of the genome[END_REF]. The H2A.X serine residue at the γ-position of the C terminus can be phosphorylated (termed γ-H2A.X) as a result of DSB formation. Because DSB can potentially occur anywhere in the genome, H2A.X distributes randomly as well [START_REF] Downs | A role for Saccharomyces cerevisiae histone H2A in DNA repair[END_REF][START_REF] Rogakou | Initiation of DNA fragmentation during apoptosis induces phosphorylation of H2AX histone at serine 139[END_REF]. During spermatogenesis (Figure 16), the pachytene stage is accompanied by γ-H2A.X enrichment at the sex chromosomes to initiate meiotic sex chromosome inactivation (MSCI) and X and Y chromosome condensation, which leads to the formation of a partially paired sex body. Interestingly, as a result of the knockout of H2A.X in mice, this entire process was impaired, resulting in sterility [START_REF] Fernandez-Capetillo | H2AX is required for chromatin remodeling and inactivation of sex chromosomes in male mouse meiosis[END_REF].

iii. macroH2A variant

MacroH2A is a vertebrate-specific H2A variant that differs from its corresponding canonical H2A in having a large C terminal domain (200 residues), termed "the macro domain", that shares no sequence similarity with any other histone [START_REF] Malik | Phylogenomics of the nucleosome[END_REF]. In mammals, macroH2A is enriched on the inactive X chromosomes in females [START_REF] Costanzi | Histone macroH2A1 is concentrated in the inactive X chromosome of female preimplantation mouse embryos[END_REF]. Therefore it is thought to contribute to the maintenance of an inactive X chromosome. Some studies suggest that the C-terminal macro domain of macroH2A interferes with the binding of transcription factors but also with the N-terminal domain which impedes chromatin remodeling via SWI/SNF pathway [START_REF] Angelov | The histone variant macroH2A interferes with transcription factor binding and SWI/SNF nucleosome remodeling[END_REF]. In addition, macroH2A is enriched at developmental genes in human pluripotent cells in males and regulates the timing of HoxA activation. The distribution pattern of macroH2A at the Hox loci reveals its overlapping with PRC2 and therefore, marcoH2A is thought to cooperate with PRC2 and potentially acts as an epigenetic regulator of key developmental genes [START_REF] Buschbeck | The histone variant macroH2A is an epigenetic regulator of key developmental genes[END_REF]. MacroH2A can also mediate gene silencing by inhibiting the catalytic activity and substrate binding capacity of PARP1, which is a nuclear enzyme that is involved in gene activation [START_REF] Buschbeck | The histone variant macroH2A is an epigenetic regulator of key developmental genes[END_REF][START_REF] Ouararhni | The histone variant mH2A1.1 interferes with transcription by down-regulating PARP-1 enzymatic activity[END_REF].

iv. H2A.Bbd variant

This histone variant was named because of its unique genomic distribution where it is deficient in inactive X chromosomes (bar body deficient) (Table 2 and figure 16). The unique subnuclear localization and key features of H2A.Bbd suggest that it is involved in gene activation. H2A.Bbd nucleosome has a truncated C-terminal docking domain [START_REF] Chadwick | A novel chromatin protein, distantly related to histone H2A, is largely excluded from the inactive X chromosome[END_REF]. As a result it binds only 116 base pairs of DNA making the H2A.Bbd nucleosome less stable than the canonical nucleosome [START_REF] Bao | Nucleosomes containing the histone variant H2A.Bbd organize only 118 base pairs of DNA[END_REF][START_REF] Gautier | Histone variant H2ABbd confers lower stability to the nucleosome[END_REF].

H2A.Bbd nucleosome not only lacks a small acidic region on the surface that is involved in transcriptional repression but also also K119, a residue that is often ubiquitinated on canonical H2A [START_REF] Zhou | The nucleosome surface regulates chromatin compaction and couples it with transcriptional repression[END_REF].

v. H2AL variant

H2AL variant is a H2A.Bbd-like histone. It was recently identified in mouse spermatids. It has truncated C-terminal region and specifically marks pericentric regions [START_REF] Ferguson | Two novel mouse genes mapped to chromosome Yp are expressed specifically in spermatids[END_REF][START_REF] Govin | Pericentric heterochromatin reprogramming by new histone variants during mouse spermiogenesis[END_REF]. H2AL is also ectopically expressed in somatic cells and interact with the testis-specific H2B variant TH2B forming more labile nucleosomes than canonical H2A/H2B dimers [START_REF] Govin | Pericentric heterochromatin reprogramming by new histone variants during mouse spermiogenesis[END_REF][START_REF] Van Roijen | Immunoexpression of testis-specific histone 2B in human spermatozoa and testis tissue[END_REF][START_REF] Zalensky | Human testis/sperm-specific histone H2B (hTSH2B). Molecular cloning and characterization[END_REF].

vi. TH2A variant

TH2A is a testis-specific histone variant expressed and incorporated within spermatocytes chromatin during pachytene (Figure 16) and disappears in round spermatids but its exact function is still unknown [START_REF] Meistrich | Histone variants in rat spermatogonia and primary spermatocytes[END_REF][START_REF] Rao | Structural organization of the meiotic prophase chromatin in the rat testis[END_REF].

H2B variants

In contrast to H3 and H2A histones, H2B isoforms are expressed in few tissues (Table 2), including sperm specific H2B variant (spH2B). In human, two variants of histone H2B specifically expressed in the testis have been described: TH2B and H2BFWT (Table 2) [START_REF] Gineitis | Human sperm telomerebinding complex involves histone H2B and secures telomere membrane attachment[END_REF]. Interestingly, TH2B was proposed to be a platform in specifying pericentric heterochromatin during late spermatogenesis (Table 2 and figure 16) [START_REF] Govin | Pericentric heterochromatin reprogramming by new histone variants during mouse spermiogenesis[END_REF]. On the other hand, H2BFWT appears to be enriched at telomere interstitial blocks, and it has been proposed that it may serve as an epigenetic marker of telomeric identity in testis (Table 2 and figure 16) [START_REF] Churikov | Novel human testis-specific histone H2B encoded by the interrupted gene on the X chromosome[END_REF].

H3 Histone variants

In human, eight histone H3 proteins were identified [START_REF] Hamiche | Chaperoning the histone H3 family[END_REF]. They have been arranged on the basis of their incorporation onto chromatin, in two different groups: two replication-dependent canonical H3 histones (H3.1 and H3.2) and six replication-independent histone H3 variants (H3t, H3.3, CENP-A, H3.X, H3.Y and H3.5). It is noteworthy that H3.3, CENP-A, H3.X, and H3.Y are somatic histone variants, while H3t and H3.5 are testis specific variants [START_REF] Hamiche | Chaperoning the histone H3 family[END_REF]. In yeast, a single H3 isoform has been detected equivalent to the mammalian H3.3 that can be deposited by both replication dependent and replication independent pathways [START_REF] Hamiche | Chaperoning the histone H3 family[END_REF]. Thus, H3.3 gene is suggested to be the common ancestor which gave rise to all other non-centromeric major H3 variants [START_REF] Postberg | The evolutionary history of histone H3 suggests a deep eukaryotic root of chromatin modifying mechanisms[END_REF]. Drosophila H3.2 variant would be the first derivative, followed by H3.1 and H3t that appeared on mammals [START_REF] Malik | Phylogenomics of the nucleosome[END_REF][START_REF] Postberg | The evolutionary history of histone H3 suggests a deep eukaryotic root of chromatin modifying mechanisms[END_REF].

H3.X and H3.Y were later detected in primates [START_REF] Wiedemann | Identification and characterization of two novel primate-specific histone H3 variants, H3.X and H3[END_REF]. The amino acids differences between the different H3 histones may provide the specificity for their differential chromatin assembly and regulation.

i. CENP-A

CenH3 or CENP-A (Table 2) was identified as an H3 variant during co-purification with other core histones [START_REF] Palmer | A 17-kD centromere protein (CENP-A) copurifies with nucleosome core particles and with histones[END_REF][START_REF] Sullivan | Human CENP-A contains a histone H3 related histone fold domain that is required for targeting to the centromere[END_REF]. The other H3 variants, such as H3.1, H3.2 and H3.3, are at the same molecular weight and differ by only four to five amino acid residues [START_REF] Palmer | A 17-kD centromere protein (CENP-A) copurifies with nucleosome core particles and with histones[END_REF]. In contrast, CENP-A has a variable N-terminus, with no sequence similarity to the N-terminal region of the other H3 variants [START_REF] Sarma | Histone variants meet their match[END_REF]. Moreover, CENP-A shares only 50% identity in the histone fold domain with the other H3 histones [START_REF] Malik | Phylogenomics of the nucleosome[END_REF]. CENP-A is the first histone variant that was shown to identify specific chromatin regions, namely the centromeres [START_REF] Henikoff | Histone variants, nucleosome assembly and epigenetic inheritance[END_REF]. CENP-A is essential for kinetochore formation and chromosome segregation [START_REF] Allshire | Epigenetic regulation of centromeric chromatin: old dogs, new tricks?[END_REF][START_REF] Henikoff | Assembly of variant histones into chromatin[END_REF][START_REF] Sarma | Histone variants meet their match[END_REF].

Table 2: Localizations and functions of canonical core histones and histone variants in mammals

Canonical histones in bold, N.D.-not determined, RC-replication-coupled, RI-replicationindependent, Xi-inactive X chromosome.

Histone Localization Function Reference

H2A

Genome-wide canonical histone H2A.Z Regulatory elements, promoter, pericentric repeats transcriptional activation, nucleosome instability [START_REF] Gupta | Predicting human nucleosome occupancy from primary sequence[END_REF][START_REF] Jin | H3.3/H2A.Z double variantcontaining nucleosomes mark 'nucleosome-free regions' of active promoters and other regulatory regions[END_REF] H2A.X XY Body, sites of double-strand DNA breaks marker of DNA lesions [START_REF] Fernandez-Capetillo | H2AX: the histone guardian of the genome[END_REF].

MacroH2A

Xi chromosome, promoters X inactivation [START_REF] Costanzi | Histone macroH2A1 is concentrated in the inactive X chromosome of female preimplantation mouse embryos[END_REF][START_REF] Pehrson | MacroH2A, a core histone containing a large nonhistone region[END_REF] H2A.Bbd Excluded from Xi destabilizing protein-DNA interaction [START_REF] Chadwick | A novel chromatin protein, distantly related to histone H2A, is largely excluded from the inactive X chromosome[END_REF] H2A.L Pericentric repeats of spermatids N.D. [START_REF] Govin | Pericentric heterochromatin reprogramming by new histone variants during mouse spermiogenesis[END_REF] TH2A Testis N.D. [START_REF] Meistrich | Histone variants in rat spermatogonia and primary spermatocytes[END_REF][START_REF] Rao | Structural organization of the meiotic prophase chromatin in the rat testis[END_REF])

H2B

Genome-wide canonical histone TH2B Testis pericentric heterochromatin. [START_REF] Govin | Pericentric heterochromatin reprogramming by new histone variants during mouse spermiogenesis[END_REF] H2BFWT Testis telomeric identity [START_REF] Churikov | Novel human testis-specific histone H2B encoded by the interrupted gene on the X chromosome[END_REF].

H3.1

Genome-wide canonical histone (RC assembly) [START_REF] Tagami | Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis[END_REF] H3.2 Genome-wide canonical histone (RC assembly) H3.3 a) euchromatin, promoter, gene bodies transcriptional activation (RI assembly) [START_REF] Ahmad | The histone variant H3.3 marks active chromatin by replication-independent nucleosome assembly[END_REF][START_REF] Jin | H3.3/H2A.Z double variantcontaining nucleosomes mark 'nucleosome-free regions' of active promoters and other regulatory regions[END_REF] b) telomeres, pericentric repeats N.D. [START_REF] Goldberg | Distinct factors control histone variant H3.3 localization at specific genomic regions[END_REF] CENP-A Centromere kinetochore assembly [START_REF] Palmer | Purification of the centromere-specific protein CENP-A and demonstration that it is a distinctive histone[END_REF][START_REF] Sullivan | Human CENP-A contains a histone H3 related histone fold domain that is required for targeting to the centromere[END_REF] H3T Testis N.D. [START_REF] Albig | A solitary human H3 histone gene on chromosome 1[END_REF][START_REF] Witt | Testis-specific expression of a novel human H3 histone gene[END_REF] TH3 variant Spermatogonia-specific N.D. [START_REF] Meistrich | Histone variants in rat spermatogonia and primary spermatocytes[END_REF][START_REF] Trostle-Weige | Isolation and characterization of TH3, a germ cell-specific variant of histone 3 in rat testis[END_REF].

H3.X and H3.Y

Primates-specific N.D. [START_REF] Wiedemann | Identification and characterization of two novel primate-specific histone H3 variants, H3.X and H3[END_REF] H3.5 variant Hominid testis-specific N.D. [START_REF] Schenk | H3.5 is a novel hominid-specific histone H3 variant that is specifically expressed in the seminiferous tubules of human testes[END_REF] ii. Histone H3t variant

The testis specific variant H3t (Table 2) has four amino acid substitutions (A24V, V71M, A98S, A111V) that distinguish it from its canonical H3.1 variant [START_REF] Hamiche | Chaperoning the histone H3 family[END_REF]. The H3t variant has been identified as a human variant that is specifically expressed in the testis spermatocytes stage [START_REF] Albig | A solitary human H3 histone gene on chromosome 1[END_REF][START_REF] Witt | Testis-specific expression of a novel human H3 histone gene[END_REF]. H3t variant was reported recently to be present in HeLa cells [START_REF] Govin | Testis-specific histone H3 expression in somatic cells[END_REF], in the brain and in embryos [START_REF] Govin | Pericentric heterochromatin reprogramming by new histone variants during mouse spermiogenesis[END_REF]; its exact function remains unknown.

iii. Histone TH3 variant

TH3 is specific to the rat and was strongly detected in spermatogonia and weakly in spermatocytes and round spermatids unlike other testis specific histones such as TH2A, H1T

and TH2B [START_REF] Meistrich | Histone variants in rat spermatogonia and primary spermatocytes[END_REF][START_REF] Trostle-Weige | Isolation and characterization of TH3, a germ cell-specific variant of histone 3 in rat testis[END_REF].

iv. Histone H3.X and H3.Y variants H3.X and H3.Y (Table 2), the recently identified histones in primates, display interesting changes in amino acids that are known to be modified in H3. 1, H3.2, and H3.3 (Wiedemann et al., 2010).

v. Histone H3.5 variant

Another newly identified hominid-specific histone H3 variant, H3.5 is specifically expressed in testis (Table 2) and was shown to be associated with actively transcribed genes [START_REF] Schenk | H3.5 is a novel hominid-specific histone H3 variant that is specifically expressed in the seminiferous tubules of human testes[END_REF].

vi. Histone variant H3.3

A specific chapter will be dedicated to H3.3 and its protein chaperones.

In conclusion we can say that the use of histone variants is an essential tool in the control of nucleosome and chromatin structure. The differential use of these variants has very important biological impacts. To illustrate this complex use of histone variants an interesting example is their use in the formation of the mammalian male germ cells where they are essential to regulate the complex events such as meiosis and histone replacements. These variants are with no doubt indispensable for male fertility.

Figure 16: Major histone variants involved during spermatogenesis

The early meiotic events in spermatogenesis involve a number of epigenetic changes and differential incorporation of histone variants into paternal chromatin [START_REF] Banaszynski | Histone variants in metazoan development[END_REF].

B. Post-translational histone modifications

In addition to the diversity in nucleosome structure that comes from the usage of histone variants which have specialized roles in chromosome biology, chemical modifications of histones play essential roles. The histones of the nucleosomal core share structural histone fold domains but have unstructured N-terminal tails. Amino acid residues on these tails serve as substrates for the chemical modifications such as acetylation, methylation, ubiquitination, sumoylation, phosphorylation, ADP-ribosylation etc (Figure 17). [START_REF] Graff | Epigenetic codes in cognition and behaviour[END_REF].

These modifications are believed to act combinatorially as a marking system termed 'the histone code' [START_REF] Strahl | The language of covalent histone modifications[END_REF]. They act through the recruitment of a class of factors known as 'chromatin readers' that alter chromatin in various ways. They occur mainly at the N-terminal tails. These short tails (less than 40 amino acids long) that emerge from the nucleosome are subject to multiple posttranslational modifications (PTMs) [START_REF] Campos | Histones: annotating chromatin[END_REF]. PTMs can also be localized to the globular domain or the C-terminal (Figure 17). These changes are put in place and removed by many specific enzymes (Allis et al., 2007a). An overview of the most classical modifications is given here.

Histone acetylation

Histone acetylation was first discovered in the 1960's [START_REF] Allfrey | Acetylation and Methylation of Histones and Their Possible Role in the Regulation of Rna Synthesis[END_REF]. This led to the discovery of the link between the acetylation state of the histone and transcriptional activation. Histone acetylation within the histone tail neutralizes the tail's positive charge, allowing the chromatin to relax which provides space for the transcriptional machinery to access the DNA. The acetylation reaction includes the transfer of an acetyl group from acetyl coenzyme A to amino group of a lysine residue on the histone [START_REF] Roth | Histone acetyltransferases[END_REF]. This reaction is catalyzed by a class of acetyltransferases; the first acetyltransferase to be discovered was HAT1 [START_REF] Kleff | Identification of a gene encoding a yeast histone H4 acetyltransferase[END_REF]. Although this enzyme was first found to localize to the cytoplasm, some studies confirmed its presence in the nucleus [START_REF] Kelly | Type B histone acetyltransferase Hat1p participates in telomeric silencing[END_REF]. Two classes of enzymes reversibly regulate the acetylation state of the histone tail: histone acetyltransferases (HATs) and histone deacetylases (HDACs) [START_REF] Archer | Histone acetylation and cancer[END_REF].

Histone deacetylases catalyze the removal of acetyl groups from the lysine on the tails of histones, which leads to chromatin condensation and transcriptional repression [START_REF] Taunton | A mammalian histone deacetylase related to the yeast transcriptional regulator Rpd3p[END_REF].

Histone methylation i. Overview

Methylation of histones can occur on lysine (K) or arginine (R) residues. The lysines can be modified by the addition of 1, 2 or 3 methyl group (s). This modification is implemented through the activity of histone lysine methyltransferases (KMTs) and removed by histone lysine demethylases (KDMs). The methylation does not affect the charge of histones and therefore does not have a direct effect on the structure of the nucleosome. The relationship between histone methylation and transcriptional state is complex and depends on the modified residue [START_REF] Bannister | Regulation of chromatin by histone modifications[END_REF]. Methyltransferases are grouped in three distinct protein families: the PRMT1, protein arginine methyltransferase 1 family (catalyzing arginine methylation), the SET-domain family and the non-SET domain proteins DOT1/DOT1L [START_REF] Ng | Lysine methylation within the globular domain of histone H3 by Dot1 is important for telomeric silencing and Sir protein association[END_REF]. The SET domain was first identified in three Drosophila proteins: Suppressor of position effect variegation 3-9 (SU(VAR)3-9), Enhancer of zeste (E(Z) and Trithorax (Trx) [START_REF] Jones | The Drosophila Polycomb-group gene Enhancer of zeste contains a region with sequence similarity to trithorax[END_REF][START_REF] Stassen | The Drosophila trithorax proteins contain a novel variant of the nuclear receptor type DNA binding domain and an ancient conserved motif found in other chromosomal proteins[END_REF][START_REF] Tschiersch | The protein encoded by the Drosophila position-effect variegation suppressor gene Su(var)3-9 combines domains of antagonistic regulators of homeotic gene complexes[END_REF]. SUV39H lysine methyltransferase that specifically methylates lysine 9 on the N-terminal tail of H3 was the first to be discovered [START_REF] Rea | Regulation of chromatin structure by site-specific histone H3 methyltransferases[END_REF]. By now several methyltransferases and their sites of modification have been identified [START_REF] Kouzarides | Chromatin modifications and their function[END_REF]. In all cases the methyl donor is S-Adenosyl methionine (SAM) that is converted during the methylation reaction to S-Adenosyl homocysteine (SAH). There are six well characterized methylation sites on histones: H3 (K4, K9, K27, K36, K79) and H4K20. Methylation at H3K4, K36 and K79 is mostly coupled with active transcription [START_REF] Lee | A site to remember: H3K36 methylation a mark for histone deacetylation[END_REF][START_REF] Ruthenburg | Methylation of lysine 4 on histone H3: intricacy of writing and reading a single epigenetic mark[END_REF][START_REF] Steger | DOT1L/KMT4 recruitment and H3K79 methylation are ubiquitously coupled with gene transcription in mammalian cells[END_REF] whereas the H3K9, K27 and H4K20 trimethylation is linked to transcriptional repression [START_REF] Ebert | Histone modification and the control of heterochromatic gene silencing in Drosophila[END_REF]. Among these modifications H3K9 methylation is probably one of the best documented.

ii. H3K9 methylation and HP1 recruitment

Methylation of H3K9 is mostly associated with transcriptionally inactive chromatin (Peters et al., 2003;[START_REF] Schotta | Central role of Drosophila SU(VAR)3-9 in histone H3-K9 methylation and heterochromatic gene silencing[END_REF] but it is also found in transcribed regions of some active genes [START_REF] Vakoc | Histone H3 lysine 9 methylation and HP1gamma are associated with transcription elongation through mammalian chromatin[END_REF]. Different methylation states (me1, me2 or me3) are found within different regions of the genome suggesting various functions of H3K9 methylation [START_REF] Ebert | Su(var) genes regulate the balance between euchromatin and heterochromatin in Drosophila[END_REF]. H3K9 di and trimethylation is involved in pericentromeric heterochromatin formation [START_REF] Ebert | Histone modification and the control of heterochromatic gene silencing in Drosophila[END_REF]. For this function, additional proteins are needed, first SUV39H

methylating histone H3 at K9 creating a binding site for HP1 (heterochromatin protein 1) (Figure 18). HP1 binds via its chromodomain to di-and trimethylated H3K9 [START_REF] Bannister | Selective recognition of methylated lysine 9 on histone H3 by the HP1 chromo domain[END_REF][START_REF] Lachner | Methylation of histone H3 lysine 9 creates a binding site for HP1 proteins[END_REF]. After the initiation site has been introduced, heterochromatin can spread by the binding of SUV39H to HP1 that in return leads to more methylation of H3K9 [START_REF] Almouzni | HP1 and the dynamics of heterochromatin maintenance[END_REF]. By H3K9 methylation tethering to HP1 (Figure 18), various effector proteins are recruited, which in turn regulate various chromosomal processes.

Through the Swi6/HP1 pathway it is possible to carry out a dual function mediating both transcriptional silencing and activation of target loci [START_REF] Cryderman | Role of Drosophila HP1 in euchromatic gene expression[END_REF][START_REF] Huisinga | The contradictory definitions of heterochromatin: transcription and silencing[END_REF][START_REF] Lu | Heterochromatin protein 1 is required for the normal expression of two heterochromatin genes in Drosophila[END_REF][START_REF] Vakoc | Histone H3 lysine 9 methylation and HP1gamma are associated with transcription elongation through mammalian chromatin[END_REF][START_REF] Yasuhara | Oxymoron no more: the expanding world of heterochromatic genes[END_REF]. For example, HP1

recruitment by H3K9 methylation allows the RNAi machinery to spread and operate across large chromosomal domains. Once in place, it provides a mechanism to monitor, detect and remove inappropriate, repeat-derived transcripts [START_REF] Cam | Comprehensive analysis of heterochromatin-and RNAi-mediated epigenetic control of the fission yeast genome[END_REF][START_REF] Noma | RITS acts in cis to promote RNA interference-mediated transcriptional and post-transcriptional silencing[END_REF][START_REF] Sugiyama | RNA-dependent RNA polymerase is an essential component of a self-enforcing loop coupling heterochromatin assembly to siRNA production[END_REF]. The defects in HP1 recruitment, especially those affecting the pericentromeric regions, result in mis-segregation of chromosomes [START_REF] Allshire | Mutations derepressing silent centromeric domains in fission yeast disrupt chromosome segregation[END_REF][START_REF] Grewal | Histone deacetylase homologs regulate epigenetic inheritance of transcriptional silencing and chromosome segregation in fission yeast[END_REF][START_REF] Kellum | Heterochromatin protein 1 is required for correct chromosome segregation in Drosophila embryos[END_REF][START_REF] Peters | Loss of the Suv39h histone methyltransferases impairs mammalian heterochromatin and genome stability[END_REF]. In D. melanogaster, centromeric heterochromatin can facilitate the achiasmate (non-exchange) segregation of chromosomes during meiosis [START_REF] Karpen | Centric heterochromatin and the efficiency of achiasmate disjunction in Drosophila female meiosis[END_REF]. Through Swi6, heterochromatin can directly recruit factors such as cohesin complex, which is essential for sister-chromatid cohesion [START_REF] Bernard | Requirement of heterochromatin for cohesion at centromeres[END_REF][START_REF] Nonaka | Recruitment of cohesin to heterochromatic regions by Swi6/HP1 in fission yeast[END_REF].

Histone ubiquitination and sumoylation

Protein ubiquitination is a key mechanism for the regulation of various cellular processes.

Ubiquitin group is covalently attached to lysine residues. Ubiquitination involves three successive major classes of enzymes called E1 (ubiquitin-activating enzyme), E2 (ubiquitinconjugating enzyme) and E3 (ubiquitin ligase enzyme). The E1 enzyme mainly activates ubiquitin in a process dependent on ATP. The activated ubiquitin is then transferred to E2 enzyme via a thioester bridge. The E3 ubiquitin ligase complex then interacts with the E2 enzyme via the thioester bridge and transfers ubiquitin to the lysines of the protein [START_REF] Deshaies | RING domain E3 ubiquitin ligases[END_REF]. Histones H2A and H2B are subjects to mono-ubiquitination mainly on the lysines 119 and 120 in mammals. The relationship between ubiquitination of histones and transcriptional state depends on the modified residue. In fact ubiquitination of H2A (H2AK119ub) on the body of genes is correlated with repression of transcription, whereas The Heterochromatin HP1 tethering by H3K9 methylation represents a platform for the recruitment of effectors across various extended domains which in turn regulate various chromosomal processes [START_REF] Grewal | Heterochromatin revisited[END_REF].

ubiquitination of H2B (H2BK120ub) at promoters and coding regions is associated with transcriptionally active chromatin [START_REF] Weake | Histone ubiquitination: triggering gene activity[END_REF]. Furthermore, ubiquitination of histones H2A and H2AX on lysine 63 is a key response to DNA damage [START_REF] Srivastava | Role of H2AX in DNA damage response and human cancers[END_REF]. Sumoylation is a post-translational modification leading to a covalent bond of a small ubiquitin-related modifier (SUMO) on a target protein. The process of sumoylation is biochemically very similar to that of ubiquitination. Like ubiquitination, sumoylation is a reversible and dynamic process [START_REF] Sarge | SUMO and its role in human diseases[END_REF].

Histone Phosphorylation

Histones can be phosphorylated on serine (S), threonine (T) and tyrosine (Y). The level of phosphorylation of histones is controlled by the activity of kinases and phosphatases enzymes.

Phosphorylation of serine 10 of histone H3 (H3S10) is extensively studied and is found at high levels in active genes. Mitotic chromosomes, which are very condensed and transcriptionally inactive structures, are also rich in H3S10P [START_REF] Johansen | Regulation of chromatin structure by histone H3S10 phosphorylation[END_REF].

Phosphorylation of histone variant H2AX at serine 139, named γH2AX, is implemented very quickly after inducing double strand breaks in DNA (DSBs). It is involved in signaling the damage and is commonly used as a marker of DSB. Moreover, γH2AX was found to be required also for the accumulation of many subsequent DNA damage response (DDR) proteins at DSBs sites [START_REF] Bonner | GammaH2AX and cancer[END_REF]. Thus γH2AX is believed to be the principal signaling protein involved in DDR and may play an important role in DNA repair [START_REF] Yuan | Focus on histone variant H2AX: to be or not to be[END_REF].

C. ATP-dependent Chromatin Remodelers

One question arising is how cells manage different chemical modifications and incorporate histone variants into the dense structure of chromatin. Furthermore, processes like transcription require access to DNA. By default, the access to DNA is virtually blocked by nucleosomes. In order to render DNA accessible, multiprotein machineries exist that utilize ATP hydrolysis to mobilize nucleosomes [START_REF] Becker | ATP-dependent nucleosome remodeling[END_REF]. Histones may then be moved to a new location or replaced by histone variants [START_REF] Langst | Nucleosome remodeling: one mechanism, many phenomena?[END_REF][START_REF] Tomar | A novel mechanism of antagonism between ATP-dependent chromatin remodeling complexes regulates RNR3 expression[END_REF][START_REF] Workman | Nucleosome displacement in transcription[END_REF].

ATP-dependent chromatin remodeling enzymes are large (>1 MDa) multi-component complexes (consisting of 4-17 subunits) that are highly conserved within eukaryotes [START_REF] Tang | Structure and function of SWI/SNF chromatin remodeling complexes and mechanistic implications for transcription[END_REF]. They are characterized by the presence of an ATPase subunit belonging to the superfamily II helicase-related proteins (Figure 19) [START_REF] Singleton | Modularity and specialization in superfamily 1 and 2 helicases[END_REF]. The first complex to be described was the yeast SWI/SNF which was originally identified in two independent screens for mutants affecting either mating type switching or growth on sucrose [START_REF] Sudarsanam | The Swi/Snf family nucleosome-remodeling complexes and transcriptional control[END_REF][START_REF] Workman | Alteration of nucleosome structure as a mechanism of transcriptional regulation[END_REF]. Hence, the names Switching defective (SWI) and Sucrose non-fermenting (Snf) [START_REF] Mohrmann | Composition and functional specificity of SWI2/SNF2 class chromatin remodeling complexes[END_REF]. The Snf2 helicase domain is conserved in these complexes and is capable of binding and hydrolyzing ATP [START_REF] Eisen | Evolution of the SNF2 family of proteins: subfamilies with distinct sequences and functions[END_REF]. Four main families have been described: Swi/Snf, Iswi, Ino80/SWR1, CHD [START_REF] Hargreaves | ATP-dependent chromatin remodeling: genetics, genomics and mechanisms[END_REF].

SWI/SNF family

Drosophila brahma (BRM), mammalian Brahma related gene 1 (BRG1), and yeast SNF2 are examples of proteins, which are categorized into the Swi/Snf family of proteins. In addition to the Snf2 helicase domain, all of these proteins possess a bromodomain, which has been reported to bind acetylated histone tails [START_REF] Marfella | The Chd family of chromatin remodelers[END_REF][START_REF] Wang | Chromatin remodeling and cancer, Part II: ATP-dependent chromatin remodeling[END_REF].

Swi/Snf complexes are involved in various cellular processes such as DNA replication, repair and transcription [START_REF] Wang | Chromatin remodeling and cancer, Part II: ATP-dependent chromatin remodeling[END_REF] 2. ISWI family Mammalian SNF2H and yeast Isw1 are examples of enzymes, which are categorized as Iswi (imitation switch) remodeling enzymes. In addition to the Snf2 helicase domain, proteins within this family possess a SANT (SWI3, ADA2, NCOR, TFIIIB) domain. This domain is reported to have the ability to bind histone tails [START_REF] Marfella | The Chd family of chromatin remodelers[END_REF][START_REF] Wang | Chromatin remodeling and cancer, Part II: ATP-dependent chromatin remodeling[END_REF]. The chromatin accessibility complex (CHRAC) [START_REF] Varga-Weisz | Chromatin-remodelling factor CHRAC contains the ATPases ISWI and topoisomerase II[END_REF], nucleosome remodeling factor (NURF) complex (Tsukiyama et al., 1995;Tsukiyama and Wu, 1995), and ATP-utilizing chromatin assembly and remodeling factor complex (ACF) [START_REF] Ito | ACF, an ISWI-containing and ATPutilizing chromatin assembly and remodeling factor[END_REF] are examples of complexes which are classified as ISWI complexes. The Iswi protein within each complex acts as the ATPase subunit of the given complex [START_REF] Elfring | Identification and characterization of Drosophila relatives of the yeast transcriptional activator SNF2/SWI2[END_REF]. While the Iswi family was initially identified in Drosophila, paralogues of the Drosophila Iswi proteins have been found in complexes in Yeast [START_REF] Mcconnell | Histone fold protein Dls1p is required for Isw2dependent chromatin remodeling in vivo[END_REF][START_REF] Tsukiyama | Characterization of the imitation switch subfamily of ATP-dependent chromatin-remodeling factors in Saccharomyces cerevisiae[END_REF][START_REF] Vary | Yeast Isw1p forms two separable complexes in vivo[END_REF], Xenopus [START_REF] Guschin | Multiple ISWI ATPase complexes from xenopus laevis. Functional conservation of an ACF/CHRAC homolog[END_REF][START_REF] Maccallum | ISWI remodeling complexes in Xenopus egg extracts: identification as major chromosomal components that are regulated by INCENP-aurora B[END_REF], and humans [START_REF] Barak | Isolation of human NURF: a regulator of Engrailed gene expression[END_REF][START_REF] Bochar | A family of chromatin remodeling factors related to Williams syndrome transcription factor[END_REF][START_REF] Hakimi | A chromatin remodelling complex that loads cohesin onto human chromosomes[END_REF][START_REF] Langst | Nucleosome remodeling: one mechanism, many phenomena?[END_REF][START_REF] Leroy | Requirement of RSF and FACT for transcription of chromatin templates in vitro[END_REF][START_REF] Poot | HuCHRAC, a human ISWI chromatin remodelling complex contains hACF1 and two novel histone-fold proteins[END_REF][START_REF] Strohner | NoRC-a novel member of mammalian ISWI-containing chromatin remodeling machines[END_REF][START_REF] Yasui | SATB1 targets chromatin remodelling to regulate genes over long distances[END_REF]. 4) [START_REF] Mohrmann | Composition and functional specificity of SWI2/SNF2 class chromatin remodeling complexes[END_REF].

CHD family

The CHD (Chromodomain helicase DNA binding) proteins are another example of ATPdependent chromatin remodeling family. Like all ATP-dependent remodeling enzymes, members of this family possess a conserved Snf2 helicase domain. In the CHDs, this domain is C-terminal to tandem chromodomains believed to function in histone binding. The CHD family of proteins can be further divided into subfamilies based on the presence of additional domains. In addition to the double chromodomains and Snf2 helicase domain, CHD1 has a DNA-binding domain near its C-terminus [START_REF] Marfella | The Chd family of chromatin remodelers[END_REF]. CHD1 has been reported to be required for H3.3 deposition on the paternal pronucleus [START_REF] Konev | CHD1 motor protein is required for deposition of histone variant H3.3 into chromatin in vivo[END_REF].

However these data are questioned by our recently published work. We believe that it is required for nucleosome organization on the newly assembled chromatin (this manuscript).

CHD1 has been shown to play essential roles in transcription not only in yeast [START_REF] Alen | A role for chromatin remodeling in transcriptional termination by RNA polymerase II[END_REF][START_REF] Krogan | RNA polymerase II elongation factors of Saccharomyces cerevisiae: a targeted proteomics approach[END_REF][START_REF] Simic | Chromatin remodeling protein Chd1 interacts with transcription elongation factors and localizes to transcribed genes[END_REF] but also in Drosophila [START_REF] Mcdaniel | Investigations of CHD1 function in transcription and development of Drosophila melanogaster[END_REF].

Both human and yeast CHD1 are reported to bind histones methylated on H3 lysine 4, a hallmark of active transcription [START_REF] Marfella | The Chd family of chromatin remodelers[END_REF]. CHD1 is reported to exhibit ATPase activity in yeast and Drosophila but Mouse CHD1 may also bear histone deacetylase activity [START_REF] Marfella | The Chd family of chromatin remodelers[END_REF].

INO80/SWR1family

SRCAP (SNF2-related CREB-activator protein) and p400 are examples of proteins of this family categorized as Ino80 (inositol requiring 80) remodeling proteins that have a split ATPase domain [START_REF] Bao | INO80 subfamily of chromatin remodeling complexes[END_REF][START_REF] Wang | Chromatin remodeling and cancer, Part II: ATP-dependent chromatin remodeling[END_REF]. The Ino80 complex was first identified in yeast. This complex is reported to remodel chromatin, facilitate in vitro transcription, and exhibit DNA helicase activity [START_REF] Grewal | Heterochromatin revisited[END_REF]. The DNA helicase activity of the complex has been attributed to the presence of RuvB proteins. The Ino80 complex is thought to be involved in both transcriptional regulation and DNA repair [START_REF] Bao | INO80 subfamily of chromatin remodeling complexes[END_REF][START_REF] Marfella | The Chd family of chromatin remodelers[END_REF][START_REF] Shen | A chromatin remodelling complex involved in transcription and DNA processing[END_REF]. Some recent studies revealed that INO80 complex is closely related to another ATP-dependent SWR1 complex which was the first identified H2A.Z chaperone [START_REF] Kobor | A protein complex containing the conserved Swi2/Snf2-related ATPase Swr1p deposits histone variant H2A.Z into euchromatin[END_REF][START_REF] Luk | Stepwise histone replacement by SWR1 requires dual activation with histone H2A.Z and canonical nucleosome[END_REF][START_REF] Mizuguchi | ATP-driven exchange of histone H2AZ variant catalyzed by SWR1 chromatin remodeling complex[END_REF][START_REF] Papamichos-Chronakis | Global regulation of H2A.Z localization by the INO80 chromatin-remodeling enzyme is essential for genome integrity[END_REF].

D. Histone chaperones

Histones deposition in chromatin is achieved by histone chaperones; the term "molecular chaperone" was first used by Ron Laskey to describe nuclear proteins in the extracts of frog oocytes that prevent incorrect interactions between histones and DNA [START_REF] Laskey | Nucleosomes are assembled by an acidic protein which binds histones and transfers them to DNA[END_REF].

Histone chaperones shield nonspecific interactions between the negatively charged DNA and the positively charged histones and promote specific interactions that lead to nucleosome assembly by forming an insoluble precipitate [START_REF] Laskey | Nucleosome assembly[END_REF][START_REF] Laskey | Nucleosomes are assembled by an acidic protein which binds histones and transfers them to DNA[END_REF]. The function of histone chaperones is intimately coupled to the action of ATPdependent chromatin remodeling machines to allow the ordered formation of the nucleosome structure in the chromatin assembly process [START_REF] Eitoku | Histone chaperones: 30 years from isolation to elucidation of the mechanisms of nucleosome assembly and disassembly[END_REF][START_REF] Haushalter | Chromatin assembly by DNA-translocating motors[END_REF][START_REF] Liu | Histone transfer among chaperones[END_REF][START_REF] Park | Histone chaperones in nucleosome eviction and histone exchange[END_REF]. Most histone chaperones are conserved in yeast, plants and animals [START_REF] Hamiche | Chaperoning the histone H3 family[END_REF][START_REF] Polo | Chromatin assembly: a basic recipe with various flavours[END_REF][START_REF] Zhu | Histone variants and chromatin assembly in plant abiotic stress responses[END_REF]. Assembly of nucleosomes being a stepwise process that starts with H3-H4 deposition first onto DNA, followed by incorporation of two H2A-H2B dimers.

Chaperones of the canonical histones are classified in two groups according to their affinity either toward H2A-H2B or H3-H4 [START_REF] Gruss | Disruption of the nucleosomes at the replication fork[END_REF][START_REF] Worcel | Assembly of newly replicated chromatin[END_REF]).

An overview is given here for the well characterized chaperones of canonical histones.

CENP-A chaperones are not considered here as they are highly specific for the centromeric region of the chromosome [START_REF] Hamiche | Chaperoning the histone H3 family[END_REF]. A specific chapter is dedicated to H3.3 chaperones and the H3.3 variant as they have been part of my research work.

H2A-H2B histone chaperones

The main histone chaperones of H2A-H2B are nucleoplasmin and NAP1 (Nucleosome Assembly Protein 1). Nucleoplasmin is the typical Xenopus chaperone involved in histone storage. Drosophila NAP1 protein was found associated with histone H2A and H2B in a crude whole-embryo extract by co-immunoprecipitation assay [START_REF] Ito | Drosophila NAP-1 is a core histone chaperone that functions in ATP-facilitated assembly of regularly spaced nucleosomal arrays[END_REF]. Only one NAP1 gene is present in yeast, however, higher eukaryotes, from Drosophila to human, have evolved NAP1 families comprising multiple members [START_REF] Park | The structure of nucleosome assembly protein 1[END_REF]. Other than binding with histones, NAP1 family proteins bind with other basic proteins. During spermatogenesis in Xenopus, histones, especially H2A-H2B, are largely replaced with various SP (Sperm-specific basic Protein). Incubation of NAP1 family members results in its interaction with different SP proteins and their release, leading to the final decondensation of sperm chromatin [START_REF] Matsumoto | Sperm chromatin decondensation by template activating factor I through direct interaction with basic proteins[END_REF].

H3-H4 histone chaperones

The analysis of the stepwise processes of nucleosome assembly and disassembly highlights the importance of the H3/H4 chaperones [START_REF] Liu | Histone transfer among chaperones[END_REF]. The chaperone Asf1 is the first chaperone identified to play a key role in supplying histones H3-H4 to the downstream chaperones, such as CAF-1 (Chromatin Assembly Factor 1) for nucleosome assembly (Figure 20) [START_REF] Green | Replication-independent histone deposition by the HIR complex and Asf1[END_REF][START_REF] Hamiche | Chaperoning the histone H3 family[END_REF][START_REF] Tyler | The RCAF complex mediates chromatin assembly during DNA replication and repair[END_REF]. The assembly of (H3-H4) 2 tetramers into nucleosomes is believed to be the rate limiting step of nucleosome assembly and is the key step in the inheritance of epigenetic information and maintenance of genome integrity [START_REF] Li | All roads lead to chromatin: Multiple pathways for histone deposition[END_REF].

i. CAF-1 (Chromatin Assembly Factor-1)

CAF-1 was initially identified by biochemical fractionation of extracts derived from human HeLa cells [START_REF] Smith | Purification and characterization of CAF-I, a human cell factor required for chromatin assembly during DNA replication in vitro[END_REF][START_REF] Verreault | Nucleosome assembly by a complex of CAF-1 and acetylated histones H3/H4[END_REF]. CAF-1 is the best-documented well-studied chaperone in divergent species, and is considered to be involved in the RC (Replication Coupled) chromatin assembly, DNA repair and heterochromatin silencing. It is an evolutionarily conserved factor, and in most species, it consists of three components: the large, mid and small subunits. In humans, the subunits correspond to p150, p60 and p48, in budding yeast to Cac1, Cac2 and Cac3, and in Arabidopsis to FAS1, FAS2 and MSI1 [START_REF] Ridgway | CAF-1 and the inheritance of chromatin states: at the crossroads of DNA replication and repair[END_REF]. The largest subunit of CAF-1, p150, interacts directly with the accessory factor in DNA polymerase complex, PCNA (Proliferating Cell Nuclear Antigen) (Figure 20). This provides insight into the molecular mechanism whereby CAF-1 is coupled to replication fork, and provides the first molecular link between nucleosome assembly and DNA replication [START_REF] Shibahara | Replication-dependent marking of DNA by PCNA facilitates CAF-1coupled inheritance of chromatin[END_REF].

ii. ASF-1 (Anti-Silencing Factor 1)

The analysis of crude Drosophila embryo extract revealed that DNA replication-coupled chromatin assembly by CAF-1 requires a complex that comprises histones H3 and H4 and a factor homologous to yeast ASF1 [START_REF] Tyler | The RCAF complex mediates chromatin assembly during DNA replication and repair[END_REF]. Asf1 is the central H3/H4 chaperone that accompanies H3/H4 to the nucleus [START_REF] Campos | The program for processing newly synthesized histones H3.1 and H4[END_REF]. It also acts as a histone 'sink' by buffering the majority of non-nucleosomal H3/H4 [START_REF] Groth | Human Asf1 regulates the flow of S phase histones during replicational stress[END_REF][START_REF] Tagami | Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis[END_REF].

The eukaryotic ASF1 proteins are highly conserved through evolution in structure and function. Yeast and Drosophila encode one ASF1 protein, while human genome contains two Asf1 paralogues, Asf1a and Asf1b [START_REF] Tamburini | Functional conservation and specialization among eukaryotic anti-silencing function 1 histone chaperones[END_REF]. In contrast to CAF-1 complex, ASF1 is unable on its own to promote RC nucleosome deposition (Loyola and Almouzni, B) The alignment of amino acid sequences corresponding to human H3 variants; sequences are compared with the "ancestral" variant H3.3 (amino acid differences are highlighted) and the position numbers of amino acids that are different between H3.3 and H3.1/2 are indicated [START_REF] Szenker | The double face of the histone variant H3.3[END_REF].

recognizes histone-modification such as H3K9me3 (Figure 22) [START_REF] Maze | Histone Regulation in the CNS: Basic Principles of Epigenetic Plasticity[END_REF]. ATRX chromatin remodeler with the H3.3 specific DAXX chaperone forms the DAXX-ATRX complex [START_REF] Drane | The death-associated protein DAXX is a novel histone chaperone involved in the replication-independent deposition of H3.3[END_REF]. It was found to regulate replication-independent histone H3.3 chromatin assembly at telomeres and pericentric heterochromatin (Figure 22) [START_REF] Drane | The death-associated protein DAXX is a novel histone chaperone involved in the replication-independent deposition of H3.3[END_REF][START_REF] Goldberg | Distinct factors control histone variant H3.3 localization at specific genomic regions[END_REF][START_REF] Lewis | Daxx is an H3.3-specific histone chaperone and cooperates with ATRX in replication-independent chromatin assembly at telomeres[END_REF][START_REF] Newhart | Single cell analysis of Daxx and ATRX-dependent transcriptional repression[END_REF]. In Drosophila, homologues have been identified for DAXX, DAXX-like protein (DLP) and ATRX, X-linked nuclear protein (XNP or dATRX). Unlike the mammalian ATRX, Drosophila XNP is found to mark the active transcribed genes with a major focus near heterochromatin of the X chromosome and interestingly colocalizes with H3.3 throughout the chromatin of somatic cells [START_REF] Schneiderman | The XNP remodeler targets dynamic chromatin in Drosophila[END_REF]. The ATP-dependent chromatin remodeler ATRX recognizes the H3K4me0 and H3K9me3 histone marks and forms a complex with the H3.3 specific DAXX chaperone. The system deposits the replication-independent histone variant H3.3 at telomeric and pericentromeric heterochromatic regions of the genome [START_REF] Maze | Histone Regulation in the CNS: Basic Principles of Epigenetic Plasticity[END_REF].

HIRA

The isolation of H3.3 pre-deposition complexes identified a distinct factor, the histone regulator A, HIRA [START_REF] Tagami | Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis[END_REF]. HIRA is a member of the HIR (Histone cell cycle Regulation Defective) proteins that were initially identified in yeast. In S. cerevisiae, the genes encoding Hir1p, Hir2p, Hir3p and Hpc2p were first identified as repressors of histone gene expression that are recruited to histone gene regulatory regions [START_REF] Fillingham | Two-color cell array screen reveals interdependent roles for histone chaperones and a chromatin boundary regulator in histone gene repression[END_REF][START_REF] Sherwood | Histone regulatory (hir) mutations suppress delta insertion alleles in Saccharomyces cerevisiae[END_REF][START_REF] Sherwood | Characterization of HIR1 and HIR2, two genes required for regulation of histone gene transcription in Saccharomyces cerevisiae[END_REF][START_REF] Xu | Identification of a new set of cell cycle-regulatory genes that regulate S-phase transcription of histone genes in Saccharomyces cerevisiae[END_REF]. In Schizosaccharomyces pombe, the HIRA orthologs Hip1 and Slm9 are involved in silencing some genes, long terminal repeat (LTR) retrotransposons, and transcription from cryptic promoters [START_REF] Anderson | Silencing mediated by the Schizosaccharomyces pombe HIRA complex is dependent upon the Hpc2-like protein, Hip4[END_REF][START_REF] Anderson | The fission yeast HIRA histone chaperone is required for promoter silencing and the suppression of cryptic antisense transcripts[END_REF][START_REF] Yamane | Asf1/HIRA facilitate global histone deacetylation and associate with HP1 to promote nucleosome occupancy at heterochromatic loci[END_REF]. Indeed, Hip1/Asf1 contributes to heterochromatinization by promoting histone deacetylation and promotion of HP1 binding [START_REF] Yamane | Asf1/HIRA facilitate global histone deacetylation and associate with HP1 to promote nucleosome occupancy at heterochromatic loci[END_REF]. The HIRA complex is conserved through evolution both at the structural and functional level (Figure 23) [START_REF] Amin | A global requirement for the HIR complex in the assembly of chromatin[END_REF].

Figure 23: The evolutionary conservation of the HIR/HIRA complex subunits Schematic representation shows the conservation of HIR complex subunits from yeasts to human. In S. cerevisiae HIR complex is composed of Hir1, Hir2, Hir3 and Hpc2. The human HIRA complex is composed of the proteins HIRA, Ubinuclein1 and Cabin1. In D. melanogaster, HIRA partners are less well characterized. One of the partners has been identified by our laboratory in the course of my thesis work (see the Results section). Drawing after [START_REF] Amin | A global requirement for the HIR complex in the assembly of chromatin[END_REF] with modifications.

In vertebrates, HIRA deposits H3.3 in a multicomponent complex manner that contains UBN1, CABIN1, and Anti-Silencing Factor 1a (ASF1a) [START_REF] Goldberg | Distinct factors control histone variant H3.3 localization at specific genomic regions[END_REF][START_REF] Tagami | Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis[END_REF]. Moreover, despite S. cerevisiae has only a single H3 that resembles H3.3, its deposition also involves a multicomponent complex that comprises Hir1p, Hir2p, Hir3p, Hpc2p and Asf1p [START_REF] Green | Replication-independent histone deposition by the HIR complex and Asf1[END_REF][START_REF] Prochasson | The HIR corepressor complex binds to nucleosomes generating a distinct protein/DNA complex resistant to remodeling by SWI/SNF[END_REF]. Depending on species, there are 4 or 5 subunits in the core complex. Hir1p and Hir2p both display sequence similarities to metazoan HIRA; in fact Hir1p and Hir2p are encoded by genes that eventually merged as a single gene in metazoans [START_REF] Green | Replication-independent histone deposition by the HIR complex and Asf1[END_REF][START_REF] Lorain | Structural Organization of the WD repeat protein-encoding gene HIRA in the DiGeorge syndrome critical region of human chromosome 22[END_REF][START_REF] Prochasson | The HIR corepressor complex binds to nucleosomes generating a distinct protein/DNA complex resistant to remodeling by SWI/SNF[END_REF].

Hpc2p is orthologous to two related proteins in mammals, UBN1 and UBN2 [START_REF] Balaji | HPC2 and ubinuclein define a novel family of histone chaperones conserved throughout eukaryotes[END_REF][START_REF] Banumathy | Human UBN1 is an ortholog of yeast Hpc2p and has an essential role in the HIRA/ASF1a chromatin-remodeling pathway in senescent cells[END_REF]. The first member of the HPC2/UBN was identified in Drosophila [START_REF] Ait-Ahmed | The yemanuclein-alpha: a new Drosophila DNA binding protein specific for the oocyte nucleus[END_REF]. A proteomic analysis identified Yemanuclein in a HIRA complex as well as Asf1 [START_REF] Moshkin | Histone chaperones ASF1 and NAP1 differentially modulate removal of active histone marks by LID-RPD3 complexes during NOTCH silencing[END_REF]. The Yemanuclein/HIRA complex has been confirmed by our work (this manuscript). However no evidence was provided that Asf1 is present in the complex that contains Yem as the reciprocal experiment was not done in the work reported by [START_REF] Moshkin | Histone chaperones ASF1 and NAP1 differentially modulate removal of active histone marks by LID-RPD3 complexes during NOTCH silencing[END_REF]. Asf1p also has two counterparts in mammals, ASF1a and ASF1b [START_REF] Sillje | Identification of human Asf1 chromatin assembly factors as substrates of Tousled-like kinases[END_REF] of which only ASF1a is included in the HIRAcontaining complex [START_REF] Tagami | Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis[END_REF][START_REF] Zhang | Formation of MacroH2A-containing senescence-associated heterochromatin foci and senescence driven by ASF1a and HIRA[END_REF]. Whether Asf1 is in the HIRA core complex is still unclear. What is clearly established is that Human HIRA core complex is composed of at least three subunits, including HIRA, Ubinuclein 1 (UBN1) and Calcineurin-binding protein 1 (CABIN1). CABIN1 is the yeast Hir3p similar sequence which, interestingly, is not found in the Drosophila genome [START_REF] Amin | A global requirement for the HIR complex in the assembly of chromatin[END_REF]. How the different subunits work and which one is the genuine histone H3.3 chaperone within the complex are real issues to be addressed in the near future.

C. Chromatin Remodeling of the Paternal Genome

At fertilization, sperm entry into the oocyte triggers the cell cycle of the dormant mature oocyte arrested at metaphase II to complete its second meiotic division. In Drosophila, ovulation provides the triggering signal for meiosis resumption of the metaphase I arrested oocyte. Once activated, the egg initiates crucial chromatin reprogramming events through a multi-step remodeling of the compacted and transcriptionally inactive sperm chromatin to form the male pronucleus. As soon as maternal and paternal haploid pronuclei are formed they fuse to yield the diploid zygote containing one functional copy of each parent genome opening the early window of embryonic development.

Compaction of paternal genome during spermatogenesis

The chromatin of mature mammalian spermatozoa differs markedly in composition and structure from somatic chromatin. It has an extremely specialized architecture; its genome is packaged into a volume that is typically 10% or less that of a somatic cell nucleus. To achieve this remarkable level of compaction, dramatic chromatin structural rearrangements occur during the late stages of spermatogenesis. As a result the genome is complexed with spermspecific highly basic proteins called protamines [START_REF] Braun | Packaging paternal chromosomes with protamine[END_REF]. Protamines are the major sperm nuclear basic proteins (SNBPs) [START_REF] Ausio | Histone H1 and evolution of sperm nuclear basic proteins[END_REF][START_REF] Balhorn | The protamine family of sperm nuclear proteins[END_REF]. Protamines can be defined as relatively small proteins of up to 100 amino acids with a highly basic amino acid composition consisting predominantly of arginine residues [START_REF] Ausio | Histone H1 and evolution of sperm nuclear basic proteins[END_REF][START_REF] Eirin-Lopez | Origin and evolution of chromosomal sperm proteins[END_REF]. 90%-95% of sperm chromatin histones are replaced by protamines which not only facilitate the nuclear compaction necessary for motility but also protect the genome.

Moreover it precludes the transcriptional activity [START_REF] Carrell | Epigenetics of the male gamete[END_REF][START_REF] Oliva | Protamines and male infertility[END_REF]. The structural transition from a highly dynamic canonical nucleohistone to a highly stable and compact nucleoprotamine is still poorly understood, but recent studies revealed that it is a multistep process assisted by chromatin remodeling complexes, highly germ line specific histone variants and post-translational modifications (PTMs) of the chromosomal proteins involved (Figure 24) [START_REF] Churikov | Novel human testis-specific histone H2B encoded by the interrupted gene on the X chromosome[END_REF][START_REF] Gaucher | From meiosis to postmeiotic events: the secrets of histone disappearance[END_REF][START_REF] Ishibashi | H2A.Bbd: an X-chromosome-encoded histone involved in mammalian spermiogenesis[END_REF].

Figure 24: Histone disappearance events during late spermatogenesis stages Global histone hyperacetylation is essential for the structural transition from a highly dynamic canonical nucleohistone to a highly stable and compact nucleoprotamine [START_REF] Gaucher | From meiosis to postmeiotic events: the secrets of histone disappearance[END_REF].

In both vertebrate and invertebrate organisms the process starts with an increase in the acetylation of sperm histones which relaxes chromatin structure. As a result histones replacement is enhanced with moderately basic transition proteins 1 and 2 (TP1 and TP2) and therefore, triggers the subsequent cascade of events which ends by protamine replacement [START_REF] Rousseaux | Molecular models for post-meiotic male genome reprogramming[END_REF][START_REF] Song | Immunohistochemical Analysis of Histone H3 Modifications in Germ Cells during Mouse Spermatogenesis[END_REF]. In addition to histone hyperacetylation, the histones of vertebrates were also found to be ubiquitinated prior to the protamine displacement. Histone ubiquitination has been recently shown to regulate nucleosome removal [START_REF] Baarends | Histone ubiquitination and chromatin remodeling in mouse spermatogenesis[END_REF][START_REF] Lu | RNF8-dependent histone modifications regulate nucleosome removal during spermatogenesis[END_REF]. In addition proper deposition of protamines onto the DNA template is mediated by its phosphorylation [START_REF] Lewis | A walk though vertebrate and invertebrate protamines[END_REF].

Reprogramming of paternal chromatin at fertilization

Before fertilization, the functional state of either the oocyte or the sperm genome is not yet realized due to two main raisons; first, as mentioned, the sperm genome is tightly condensed and is inaccessible for the different cellular processes. On the other hand the oocyte itself not completed its meiotic cycle. For these two raisons both genomes undergo a series of reprogramming events. At fertilization, the entry of the sperm into the oocyte cytoplasm leads to completion of maternal meiosis, resulting in formation of the polar body and the haploid maternal pronucleus. The situation is slightly different in Drosophila where female meiosis is arrested at metaphase I, its resumption depending on ovulation and not fertilization. At the same time as the maternal pronucleus forms, the paternal chromatin undergoes some drastic reorganization. Notably, the major modification lies in the decondensation of the highly compacted protamine-containing sperm chromatin to produce an accessible haploid genome.

The protamine/histone exchange must take place for DNA replication to take place [START_REF] Piatti | ATP-Dependent Chromatin Remodeling Factors and Their Roles in Affecting Nucleosome Fiber Composition[END_REF][START_REF] Wu | Epigenetic processes implemented during spermatogenesis distinguish the paternal pronucleus in the embryo[END_REF]. This implies the stripping of SNBPs followed by de novo assembly of paternal nucleosomes by the deposition of maternally provided histones. This process is called sperm chromatin remodeling (SCR); it involves different asymmetric epigenetic changes, including differential DNA methylation, posttranslational modification of histone proteins and incorporation of histone variants [START_REF] Banaszynski | Histone variants in metazoan development[END_REF][START_REF] Surani | Genetic and epigenetic regulators of pluripotency[END_REF]. One of the first steps achieved is the reduction of the protamine disulphide bonds to allow protamine removal and subsequent organization of the DNA in a nucleosomal structure [START_REF] Kempisty | Structure and role of protamines 1 and 2 in spermatogenesis and male infertility[END_REF][START_REF] Oliva | Protamines and male infertility[END_REF].

Once protamines are removed, a global reassembly of nucleosomes must occur in order to repackage the genome with maternally stored histones. Many recent studies in worms, flies and mice reported the replication-independent global histone incorporation of the histone H3.3 variant into paternal DNA during decondensation prior to the first replication cycle [START_REF] Loppin | The histone H3.3 chaperone HIRA is essential for chromatin assembly in the male pronucleus[END_REF][START_REF] Ooi | Histone H3.3 variant dynamics in the germline of Caenorhabditis elegans[END_REF][START_REF] Piatti | ATP-Dependent Chromatin Remodeling Factors and Their Roles in Affecting Nucleosome Fiber Composition[END_REF][START_REF] Van Der Heijden | Asymmetry in histone H3 variants and lysine methylation between paternal and maternal chromatin of the early mouse zygote[END_REF].

Extensive studies in Drosophila showed that the de novo incorporation of histone H3.3 into paternal DNA during decondensation was mediated by HIRA [START_REF] Loppin | The histone H3.3 chaperone HIRA is essential for chromatin assembly in the male pronucleus[END_REF]. showing the events that take place from fertilization until the first mitotic division of the zygote. The oocyte that is arrested at metaphase I at the end of oogenesis resumes meiosis upon ovulation. Whether or not it is fertilized it completes its meiotic divisions, which yields to the formation of the female pronucleus. When the egg is fertilized, simultaneously to the completion of female meiosis, the paternal genome undergoes crucial chromatin reprogramming. HIRA complex mediates de no assembly of nucleosomes with maternally provided histone H3.3 variant. This occurs after the protamines were removed by an unknown mechanism.

The point mutation changing the lysine residue at position 225 to an arginine residue in HIRA (named 'sesame' or 'ssm') leads to sterility in females as a result of defects of HIRA in the assembly of paternal chromatin at fertilization [START_REF] Bonnefoy | The essential role of Drosophila HIRA for de novo assembly of paternal chromatin at fertilization[END_REF][START_REF] Loppin | The histone H3.3 chaperone HIRA is essential for chromatin assembly in the male pronucleus[END_REF].

The role of HIRA is probably conserved in mammals, as mouse HIRA associates exclusively with the male pronucleus prior to pronuclear fusion [START_REF] Van Der Heijden | Asymmetry in histone H3 variants and lysine methylation between paternal and maternal chromatin of the early mouse zygote[END_REF]. In human cells, HIRA deposits H3.3 in a multicomponent complex manner that contains UBN1, CABIN1, and Anti-Silencing Factor 1a (ASF1a) [START_REF] Goldberg | Distinct factors control histone variant H3.3 localization at specific genomic regions[END_REF][START_REF] Tagami | Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis[END_REF].

Therefore other factors are expected to play a role in the deposition of H3.3 on paternal chromatin. As described above the Drosophila homologue for UBN1 is Yemanuclein (also called Yemanuclein-alpha or Yem-alpha). Yemanuclein was reported to be a DNA binding protein required for both oocyte meiosis and in the zygote [START_REF] Ait-Ahmed | The yemanuclein-alpha: a new Drosophila DNA binding protein specific for the oocyte nucleus[END_REF][START_REF] Meyer | A single mutation results in diploid gamete formation and parthenogenesis in a Drosophila yemanuclein-alpha meiosis I defective mutant[END_REF].

The second part of my thesis work is dedicated to the characterization of Yemanuclein as a HIRA partner in the assembly of paternal chromatin in the Drosophila zygote.

We show in the present work that Yemanuclein is indeed a HIRA partner in paternal chromatin reprogramming (this manuscript). Surprisingly, ASF1 was reported not to be required in Drosophila, for the deposition of H3.3 during male pronucleus formation [START_REF] Bonnefoy | The essential role of Drosophila HIRA for de novo assembly of paternal chromatin at fertilization[END_REF]. This conclusion requires to be verified. Interestingly a physical interaction with the chromatin remodeling factor CHD1 has been implicated in aiding Drosophila HIRA in the deposition of this histone variant [START_REF] Konev | CHD1 motor protein is required for deposition of histone variant H3.3 into chromatin in vivo[END_REF]. Whereas CHD1 results in a phenotype similar to mutations in HIRA, in our experiments CHD1 is not found in the HIRA deposition complex (this manuscript). The experimental part of my thesis work is dedicated to the functional analysis of Yemanuclein (formerly called Yemanuclein-alpha). Yemanuclein is encoded by a gene that was overlooked in screens for actors of Drosophila female meiosis. We reported its molecular characterization in an earlier work [START_REF] Ait-Ahmed | The yemanuclein-alpha: a new Drosophila DNA binding protein specific for the oocyte nucleus[END_REF]. and we showed that yemanuclein RNA is concentrated in the oocyte throughout meiosis I from the earliest germarial stages [START_REF] Capri | Implication of a 5 coding sequence in targeting maternal mRNA to the Drosophila oocyte[END_REF]. Accordingly we show in the present work that Yem protein was detected in the oocyte nucleus throughout meiosis I. From its specificity for the oocyte nucleus and its affinity for DNA we assumed a role in female meiosis but lack of genetic tools eluded its function for two decades. The first mutation on yemanuclein gene was reported recently by our laboratory [START_REF] Meyer | A single mutation results in diploid gamete formation and parthenogenesis in a Drosophila yemanuclein-alpha meiosis I defective mutant[END_REF]. This point mutation that substitutes a Glutamic acid by a

Valine in a relatively conserved domain (V478E) dramatically affects female fertility. More surprising is the production of exceptional progeny from eggs laid by females homozygous for the yem 1 mutation as a result of parthenogenetic development [START_REF] Meyer | A single mutation results in diploid gamete formation and parthenogenesis in a Drosophila yemanuclein-alpha meiosis I defective mutant[END_REF]. Yemanuclein that link the formation of the female pronucleus during meiosis and the reprogramming of the male pronucleus in the zygote. To our knowledge these observations have no precedent and highlight the exquisite coordination of events that lead to the formation of the zygotic nucleus.

Yemanuclein meiotic functions

Meiosis is a complex process that relies on two chromosome segregation steps. The specific step is meiosis I that results in the segregation of the homologous chromosomes. To achieve this end meiosis I relies on a specific organization of the chromosomes that is supported by highly specialized protein complexes: the synaptonemal complex and the cohesin complex.

Whereas the overall organization and the composition of the synaptonemal complex are specific for recombination proficient meiosis, the cohesin complex is a versatile structure whose overall composition is similar in mitotic, recombination proficient and non-proficient meiotic chromosomes. Interestingly, Yemanuclein interacts in vivo with the two key complexes on which segregation of meiotic chromosome relies. The situation of Yemanuclein is unique in that it is dispensable for the formation of the Synaptonemal Complex. Moreover mutations on Yemanuclein including loss of function alleles do not seem to affect the SC morphology. Either Yemanuclein is loosely associated to the SC with no particular function for its structure or there might be defects that would be detectable at the ultrastructural level only. The functional implication of the interaction of Yemanuclein with the Cohesin Complex also remains to be elucidated. We showed in our previous work that the yem 1 mutation does not affect chromosome cohesion at meiosis I but obviously does affect monopolar orientation of the sister kinetochores. In S. Pombe, the same cohesin subunit, the Rec8 kleisin that is paralogous to Rad21, fulfills both functions, cohesion and monopolar orientation whereas in S. Cerevisiae, Rec8 is required only for cohesion [START_REF] Petronczki | Un menage a quatre: the molecular biology of chromosome segregation in meiosis[END_REF]. There is no Rec8 in Drosophila and in insects in general. How Drosophila ensures these meiosis specific functions is still elusive. The absence of effect of yem 1 mutation on cohesion may be obscured by possible functional redundancy. In contrast, Yemanuclein critical role at early meiosis is clearly demonstrated. Yem is required for the timely formation of the double strand breaks, a result supported by the evidence of a genetic interaction between yem and mei-w68, the Drosophila homologue of spo11. The latter encodes the topoisomerase responsible for the formation of the double strand breaks. Therefore Yemanuclein function is essential for the initiation of the recombination process. Whether Yemanuclein role on DSBs formation depends on its association to the SC and the Cohesin Complex remains to be investigated.

Yemanuclein zygotic function

The following observations prompted us to investigate a possible role of Yemanuclein in the zygote:

-The presence of the Yemanuclein human orthologue, Ubinuclein, in the HIRA complex that supports H3.3 containing nucleosomes assembly [START_REF] Tagami | Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis[END_REF].

-The only known role for HIRA in Drosophila is precisely its requirement for chromatin assembly of the male pronucleus [START_REF] Loppin | The histone H3.3 chaperone HIRA is essential for chromatin assembly in the male pronucleus[END_REF].

yem 1 homozygous mothers give rise to parthenogenetic progeny, a phenomenon that could be explained by defective reprogramming of the paternal pronucleus.

Our experimental data validate a role of Yemanuclein in the zygote. This role is very similar to that of HIRA in the assembly of H3.3-containing nucleosomes on the paternal pronucleus.

Accordingly we were able to show an interaction between HIRA and Yemanuclein.

Future Directions

The originality of our work on Yemanuclein is its contribution to the unveiling of interesting features of sexual reproduction. To our knowledge, it is the first time that a gene whose specialization for sexual reproduction is so exquisite links two critical aspects: formation of a maternal pronucleus through meiosis and the paternal pronucleus through de novo chromatin assembly on the paternal genome. This ensures an interdependency of chromosome number reduction during female meiosis and diploidy restoration after the paternal pronucleus was The biochemical characterization of Yemanuclein family of proteins is at its beginning. The evidence that Yemanuclein and its orthologues in Yeast (HPC2) and Humans (UBN1) are part of the HIRA histone chaperone complex is very recent [START_REF] Balaji | HPC2 and ubinuclein define a novel family of histone chaperones conserved throughout eukaryotes[END_REF][START_REF] Banumathy | Human UBN1 is an ortholog of yeast Hpc2p and has an essential role in the HIRA/ASF1a chromatin-remodeling pathway in senescent cells[END_REF][START_REF] Moshkin | Histone chaperones ASF1 and NAP1 differentially modulate removal of active histone marks by LID-RPD3 complexes during NOTCH silencing[END_REF][START_REF] Tagami | Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis[END_REF]. Yemanuclein was also shown to have DNA binding properties and to be associated to the kinetochores of chromosomes at metaphase I, a stage that is crucial for the reductional division [START_REF] Ait-Ahmed | The yemanuclein-alpha: a new Drosophila DNA binding protein specific for the oocyte nucleus[END_REF][START_REF] Meyer | A single mutation results in diploid gamete formation and parthenogenesis in a Drosophila yemanuclein-alpha meiosis I defective mutant[END_REF].

On the one hand Yemanuclein is highly specialized for sexual reproduction; on the other hand it is involved in various biological complexes and functions. An important issue is the biochemical mechanisms that underlie the various functions. How are they linked? Can the various functions be assigned to specific domains of Yemanuclein? Our work paves the way to future investigations on Yemanuclein and evolution of sexual reproduction, a costly but well conserved strategy.

La Yemanucléine de Drosophile est nécessaire à la méiose ovocytaire et l'assemblage de la chromatine paternelle dans le zygote Mots clés : Drosophile,ovocyte,méiose,complexe synaptonémal,cohésine,recombinaison,zygote,chromatine,H3.3, histone chaperone, HIRA La reproduction sexuée repose sur deux processus fondamentaux : la méiose qui permet la formation des gamètes dont le génome est haploïde et la syngamie qui permet, après fécondation, de restaurer la diploïdie par fusion des deux noyaux parentaux haploïdes. Alors que la méiose repose respectivement sur le génome maternel pour l'ovocyte et paternel pour le spermatozoïde, la restauration de la diploïdie dans le zygote repose exclusivement sur le génome maternel. Si un pronucleus maternel compétent pour la réplication est formé au terme de la méiose ovocytaire, le génome paternel quant à lui, n'acquiert cette compétence que sous l'influence de facteurs maternels. En effet, à la fin de la méiose, le génome paternel est « empaqueté » avec des protamines qui le rendent inactif pour toute fonction biologique, en particulier la réplication. L'éviction des protamines et leur remplacement par des histones maternelles sont des étapes indispensables à l'acquisition par le génome paternel de sa compétence à la réplication, préalable à la syngamie. Tous ces événements doivent être extrêmement coordonnés afin de permettre à un premier noyau zygotique comportant les deux lots de chromosomes parentaux de se former et d'entrer dans le premier cycle mitotique.

Notre laboratoire a identifié yemanuclein-alpha, aussi appelé yemanuclein (yem) dans un crible moléculaire pour des gènes exprimés spécifiquement dans la lignée germinale femelle, et son premier allèle muté yem 1 . Cette mutation ponctuelle (V478E) a été identifiée dans un crible génétique de « stérilité femelle ». Une descendance exceptionnelle observée chez les femelles yem 1 , présente la propriété inattendue d'être parthénogénétique. Cette propriété révèle un double défaut chez le mutant : dans le processus de méiose ovocytaire qui conduit à la formation d'un pronucleus maternel haploïde mais aussi dans la formation d'un pronucleus paternel compétent pour la syngamie.

Mes travaux de thèse ont porté sur les deux aspects de la fonction de la yemanucléine. En conjuguant des méthodes de génétique, de biochimie, et de biologie cellulaire, nous avons pu mettre en évidence des fonctions essentielles de la yemanucléine dans les étapes initiales de la prophase méiotique de l'ovocyte de drosophile. Nous avons pu montrer que la yemanucléine joue un rôle clé dans la recombinaison méiotique et plus particulièrement dans la fréquence et la cinétique d'apparition des cassures double brin. Son association au complexe synaptonémal et au complexe cohésine, tous deux connus comme étant nécessaires à la ségrégation chromosomique, est un élément clé de cette fonction.

Outre cette fonction méiotique, la yemanucléine, facteur maternel, est aussi requise pour l'assemblage de la chromatine du pronucleus paternel. Nous montrons dans ce manuscrit qu'elle joue ce rôle à travers son action dans un troisième complexe, en partenariat avec la protéine HIRA. Le complexe multiprotéique contenant la protéine HIRA est connu pour sa fonction de chaperon du variant de l'histone H3.3 et son rôle dans l'assemblage de la chromatine du pronucleus paternel. La yemanucléine est le premier membre de la famille HPC2/UBN1 caractérisé. Son rôle dans l'assemblage des nucléosomes découplé de la réplication est décrit pour la première fois dans ce manuscrit. C'est aussi la première fois qu'une protéine spécifique de la reproduction est décrite pour son implication à deux étapes clés de ce processus.

Drosophila Yemanuclein is required for meiosis in the oocyte and paternal chromatin assembly in the zygote

Key words: Drosophila,oocyte,meiosis,synaptonemal complex,cohesin,recombination,zygote,chromatin,H3.3,histone chaperone,HIRA Sexual reproduction relies on two key events: formation of cells with a haploid genome through meiosis and restoration of diploidy through syngamy in the zygote. Meiosis completion is supported exclusively by the maternal genome for the oocyte and the paternal genome for the sperm cell. In contrast diploidy restoration in the zygote is entirely dependent on maternal factors. At the end of meiosis the maternal pronucleus is competent for replication, whereas the paternal genome is packed with protamines. These proteins need to be removed in the zygote and replaced by maternally provided histones before the paternal genome acquires competence for replication, a prerequisite for syngamy. All these events must be highly coordinated to allow the first zygotic nucleus to form with the two sets of parental chromosomes and enter the first mitotic cycle.

Our laboratory has identified yemanuclein-alpha, also called yemanuclein (yem) in a molecular screen for genes specifically expressed in the female germ line and its first mutant allele yem 1 , in a female sterile screen. The role played by yem not only in the meiotic process through which a haploid maternal pronucleus is formed but also in the zygotic process that makes a paternal pronucleus competent for syngamy, is underscored by the obtention of exceptional parthenogenetic progeny from yem 1 mothers.

My thesis work is precisely dedicated to the analysis of both aspects of Yemanuclein function: in the oocyte and the zygote. Using genetic, biochemical and cell biology methods we were able to uncover essential functions of Yemanuclein in early meiotic prophase in the Drosophila oocyte. Using yem 1 allele (V478E), we could show its requirement for meiotic recombination especially for the frequency and timing of the double strand breaks formation. Yemanuclein association with two protein complexes, the Synaptonemal Complex (SC) and the Cohesin complex known to be required for proper chromosome segregation, supports these findings. Beyond its meiotic function, Yemanuclein is also required in the zygote for assembly of paternal pronucleus chromatin. This is achieved through a third complex that acts as histone H3.3 chaperone. In the present manuscript we identify Yemanuclein as a partner of HIRA in its role in H3.3 nucleosome assembly and deposition on the paternal pronucleus. Interestingly Yemanuclein is the first member of the HPC2/UBN1 protein family ever characterized. The role of Yem/ HPC2/ UBN1 in replication independent chromatin remodeling remained elusive until very recently. Our work is original in that it is the first to report on a role of one member of this family in oocyte meiosis and paternal chromatin assembly in the zygote. Institut de Génétique Humaine (IGH) -CNRS UPR 1142 141, rue de la cardonille -343960 MONTPELLIER CEDEX 5 France Cette thèse a été préparée grâce à une bourse du Gouvernement Egyptien lors de mon détachement de l'Université de Zagazig où j'exerce en qualité de maître de conférence

Figure 1 :Figure 2 :Figure 4 :TablesTable 2 :

 1242 Figure 1: Sexual reproduction cycle ........................................................................................................Figure 2: Schematic representation of a meiotic cell division as compared to mitosis ...........................Figure 3: Chromosome segregation in mitosis and meiosis .................................................................. Figure 4: Architecture of the SMC complexes ...................................................................................... Figure 5: The cohesin complex and its associated proteins................................................................... Figure 6: The cohesin cycle in mitosis and meiosis. .............................................................................Figure 7: Model of synaptonemal complex structure. ........................................................................... Figure 8: Major molecular events in meiotic prophase I. ...................................................................... Figure 9: Double Holliday junction model for meiotic crossover or non-crossover formation. ........... Figure 10: Organization of the Drosophila germarium. ........................................................................ Figure 11: Oogenesis in Drosophila ...................................................................................................... Figure 12: Major molecular events in meiotic recombination in Drosophila. ....................................... Figure 13: Chromatin organization in eukaryotes ................................................................................. Figure 14: Schematic representation of stepwise assembly of the nucleosome core particle ............... Figure 15: Schematic representation of histone variants ....................................................................... Figure 16: Major histone variants involved during spermatogenesis .................................................... Figure 17: Schematic representation of post-translational histone modifications .................................Figure 18: H3K9 methylation is essential for various biological processes and mediated by HP1 recruitment ............................................................................................................................................Figure 19: ATP-dependent chromatin remodeling enzymes .................................................................Figure 20: The de novo histones H3-H4 assembly through the Asf1-CAF-1 pathway ......................... Figure 21: Schematic representation of histone H3 variants family......................................................Figure 22: H3.3 deposition through DAXX-ATRX pathway ............................................................... Figure 23: The evolutionary conservation of the HIR/HIRA complex subunits ...................................Figure 24: Histone disappearance events during late spermatogenesis stages ...................................... Figure 25: Biological events in the Drosophila fertilized egg ...............................................................

Figure

  Figure 3: Chromosome segregation in mitosis and meiosis (A) During replication phase (S phase or MEIS), cohesion is established by a complex comprising either Rad21/Scc1 (blue) during mitosis or meiosisspecific variant Rec8 (purple). Meiotic recombination between the two counterparts generates reciprocal exchange leading to chiasma formation. Chiasmata connect the two chromosomes of a bivalent and can be visualized by microscopy. (B) During anaphase of mitosis, cohesion is lost along the chromosome. During meiosis, substitution of Rad21/Scc1 by Rec8 induces a progressive loss of cohesion. First along the chromosomes in meiosis I and then near the centromeres at meiosis II[START_REF] Watanabe | Modifying sister chromatid cohesion for meiosis[END_REF]. (* found in yeast).

Figure 6 :

 6 Figure 6: The cohesin cycle in mitosis and meiosis. A) At G1 phase of the cell cycle, the Scc2/Scc4 ensures the cohesin complex loading onto chromatin. Cohesion is established between sister chromatids by acetyltransferase Eco1. Upon entry into G2/M phase, cohesion is maintained by the proteins Pds5, sororin, and Wpl1. In mammalian cells, cohesin along chromosome arms is removed at the transition between prophase and metaphase via phosphorylation of Rad21 by Polo-like kinase 1. Centromeric cohesion is protected from removal by shugoshin and PP2A. At the onset of anaphase, a protease called separase cleaves the Mcd1/Scc1/Rad21 subunit, leading to the opening of the cohesin ring and the separation of sister chromatids (Xiong and Gerton, 2010). B) Unique features of cohesin during meiosis: Cohesin (black bars) connects the two sister chromatids.Two pairs of sister chromatids, that is, two homologs, synapse thanks to the synaptonemal complex (SC; yellow) formation. Recombination occurs between non-sister chromatids. In anaphase I, cohesin complex in the chromosome arms is dissolved and the homologs segregate with cohesin only at the still intact centromeres. In anaphase II, centromeric cohesion is dissolved, and the sister chromatids segregate. Green arrowheads indicate orientation of kinetochores[START_REF] Revenkova | Keeping sister chromatids together: cohesins in meiosis[END_REF].

Figure 7 :

 7 Figure 7: Model of synaptonemal complex structure. A) Representative diagram showing a cross section of a segment of the SC with lateral elements (LE), transverse filaments (TF), central element (CE), and central region. Are also shown cohesins/condensins (blue ovals) and other LE proteins (green ovals) along the LEs (Page and Hawley, 2004). B) schematic view showing the orientation of different proteins that form SC in Drosophila, orientation disruptor (Ord), the axial element (AE)/LE component C(2)M and C(3)G (Lake and Hawley, 2012).

Figure 8 :

 8 Figure 8: Major molecular events in meiotic prophase I.Schematic representation shows the major chromosomal events of meiotic prophase I in Saccharomyces cerevisiae[START_REF] Hochwagen | Checking your breaks: surveillance mechanisms of meiotic recombination[END_REF] and Drosophila melanogaster(Joyce and McKim, 2011;[START_REF] Lake | The molecular control of meiotic chromosomal behavior: events in early meiotic prophase in Drosophila oocytes[END_REF]. Prophase I involves three main processes; synapsis, double strand break (DSB) formation and DSB repair. These processes are largely conserved across eukaryotes and are associated with many cytological changes that further divide the meiotic prophase I into five substages: leptotene, zygotene, pachytene, diplotene and diakinesis. Meiotic recombination is achieved via a physical connection between the homologous chromosomes, the chiasma, which allows chromosomes to align properly on the spindle after prophase I is completed at diakinesis. Note the differences in the timing of DSBs formation between Yeast and Drosophila.

Figure 9 :

 9 Figure 9: Double Holliday junction model for meiotic crossover or non-crossover formation. Double strand breaks are generated and their 5' ends are resected to generate a 3' overhang. A strand invasion event then generates a single-end invasion D loop intermediate. If the second end of the original DSB also engages with the homologue, a double Holliday junction is formed (shown on the left). The double Holliday junction can be resolved to form either a crossover or a non-crossover.Alternatively, the junction can be dissolved by double Holliday junction dissolution to form a noncrossover[START_REF] Youds | The choice in meiosis -defining the factors that influence crossover or noncrossover formation[END_REF].

Figure 10 :

 10 Figure 10: Organization of the Drosophila germarium.At the anterior tip of the germarium, a germline stem cell divides to produce a cystoblast (premeiotic region 1). The cystoblast undergoes four rounds of mitotic divisions to produce a 16-cell germline cyst. Within the cyst, up to four cells can initiate the formation of the synaptonemal complex (SC) (red) in zygotene, but only two prooocytes continue into meiosis and form a full-length SC in early pachytene (region 2A). At this stage programmed meiotic DSBs are induced and can be detected with an antibody to a phosphorylated form of a histone H2A variant, H2AV, at serine 137 (γ-H2AV) (blue circles denote DSBs detected with a γ-H2AV antibody). By early/midpachytene (region 2B), fewer DSBs (γ-H2AV foci) can be detected within the pro-oocytes. By mid-pachytene (region 3), one cell has been selected to become the oocyte[START_REF] Lake | The molecular control of meiotic chromosomal behavior: events in early meiotic prophase in Drosophila oocytes[END_REF].

Figure 12 :

 12 Figure 12: Major molecular events in meiotic recombination in Drosophila. Homologous recombination HR involves three main processes: pairing of homologous chromosomes via synaptonemal complex, double strand break DSBs fomation and DSBs repair. The SC is shown in green, the Drosophila genes names are shown along with human homologs in parentheses (Joyce and McKim, 2011).

Figure 13 :

 13 Figure 13: Chromatin organization in eukaryotesA) The organization of DNA within the chromatin structure: the lowest level of organization is the nucleosome in which two superhelical turns of DNA are wound around the outside of a histone octamer. Nucleosomes are connected to one another by short stretches of linker DNA. At the next level of organization the string of nucleosomes is folded into a fibre about 30 nm in diameter, and these fibers are then further folded into higher-order structures. At levels of structure beyond the nucleosome the details of folding are still uncertain[START_REF] Felsenfeld | Controlling the double helix[END_REF]. B) Chromatin organization within the nucleus. An interphase nucleus seen by electron microscopy; one may distinguish euchromatin (light, transcriptionally active) and heterochromatin (dark, highly condensed, transcriptionally inactive); image taken from[START_REF] Ross | Introduction to molecular medicine[END_REF].

Figure 17 :

 17 Figure 17: Schematic representation of post-translational histone modificationsPrevalent post-translational histone modifications of the N-and C-termini of the core histones and their residue-specific epigenetic modifications include methylation (green M boxes) of lysine and arginine, acetylation (yellow A pentagon) of lysine, phosphorylation (red P sphere) of threonine or serine and ubiquitination (blue U triangle) of lysine residues[START_REF] Graff | Epigenetic codes in cognition and behaviour[END_REF].

Figure 18 :

 18 Figure 18: H3K9 methylation is essential for various biological processes and mediated by HP1 recruitmentThe Heterochromatin HP1 tethering by H3K9 methylation represents a platform for the recruitment of effectors across various extended domains which in turn regulate various chromosomal processes[START_REF] Grewal | Heterochromatin revisited[END_REF].

Figure 19 :

 19 Figure 19: ATP-dependent chromatin remodeling enzymes A) Showing the domain structure of the four major classes of ATP-dependent chromatin remodeling complexes. Members of ATP-dependent remodeling enzyme share a conserved Snf2 helicase domain capable of binding and hydrolyzing ATP, image taken from[START_REF] Jerzmanowski | SWI/SNF chromatin remodeling and linker histones in plants[END_REF]. B) Schematic representation of possible action mechanisms of ATP-dependent chromatin remodeling enzymes that alter chromatin structure by utilizing the energy of ATP hydrolysis. These enzymes can alter chromatin structure by moving histones to a new location on the same piece of DNA (1), disrupting DNA histone contacts(2), moving histones to new DNA (3), or replacing histones with histone variants (4)[START_REF] Mohrmann | Composition and functional specificity of SWI2/SNF2 class chromatin remodeling complexes[END_REF].

Figure 21 :

 21 Figure 21: Schematic representation of histone H3 variants family A) Showing the evolutionary conservation of histone variant H3.3 which is suggested to be the ancestor of all other H3 variants.B) The alignment of amino acid sequences corresponding to human H3 variants; sequences are compared with the "ancestral" variant H3.3 (amino acid differences are highlighted) and the position numbers of amino acids that are different between H3.3 and H3.1/2 are indicated[START_REF] Szenker | The double face of the histone variant H3.3[END_REF].

Figure 22 :

 22 Figure 22: H3.3 deposition through DAXX-ATRX pathwayThe ATP-dependent chromatin remodeler ATRX recognizes the H3K4me0 and H3K9me3 histone marks and forms a complex with the H3.3 specific DAXX chaperone. The system deposits the replication-independent histone variant H3.3 at telomeric and pericentromeric heterochromatic regions of the genome[START_REF] Maze | Histone Regulation in the CNS: Basic Principles of Epigenetic Plasticity[END_REF].

Figure 25 :

 25 Figure 25: Biological events in the Drosophila fertilized egg Schematic diagram of a Drosophila fertilized eggshowing the events that take place from fertilization until the first mitotic division of the zygote. The oocyte that is arrested at metaphase I at the end of oogenesis resumes meiosis upon ovulation. Whether or not it is fertilized it completes its meiotic divisions, which yields to the formation of the female pronucleus. When the egg is fertilized, simultaneously to the completion of female meiosis, the paternal genome undergoes crucial chromatin reprogramming. HIRA complex mediates de no assembly of nucleosomes with maternally provided histone H3.3 variant. This occurs after the protamines were removed by an unknown mechanism.

Figure 1 Meyer,

 1 Figure 1

Figure 2 Figure 3 Figure 4 Figure 5

 2345 Figure 2

  Such a result supposes at least two defective steps in the sexual reproduction process of the mutant mothers. In other words the parthenogenetic adults developed from eggs in which the outcome of female meiosis was a diploid pronucleus. Moreover this diploid maternal pronucleus entered the first zygotic mitosis in absence of syngamy with the paternal counterpart. These considerations were the basis for the two directions of our research to gain insight into the functions affected in the mutant 1) during female meiosis, 2) during the process that results in the biological competence of the paternal pronucleus. Added to the previously published work, the present work contributes to unveil new functions for

  reprogrammed and made competent for biological functions. Investigating the role of Yemanuclein in other organisms is one of the directions that might help to shed light on the mechanisms that underly its functions. Preliminary data on orthopterans allowed us to show a functional conservation of Yemanuclein during meiosis (collaboration with Julio Rufas, Madrid). More interestingly we aimed to investigate the conservation of Yemanuclein function in aphids, insects that are alternatively sexual or parthenogenetic depending on environmental conditions (collaboration with Denis Tagu, Rennes). The analysis of genes such as yem on these insects should provide critical information on the mechanisms that underlie the shift between parthenogenesis and sexual reproduction. Experimental analyses combined to the phylogeny (collaboration with Christophe Terzian, Lyon) will provide invaluable information on the structural and functional evolution of Yemanuclein family of proteins. The specificity acquired by Drosophila Yemanuclein for maternal functions is intriguing. A possible interpretation is that it might have been acquired from the adaptive use of the Drosophila protein for specific needs of sexual reproduction[START_REF] Meyer | A single mutation results in diploid gamete formation and parthenogenesis in a Drosophila yemanuclein-alpha meiosis I defective mutant[END_REF]. It is of special interest to investigate how widespread this specialization is, especially among insects (collaboration with Emmanuelle d'Alençon and Philippe Fournier).

Table 1 : Sister chromatid cohesion proteins

 1

	2)M*	Rec8
		Rad21L**

Meyer, Algazeery et al 2013

ACKNOWLEDGEMENTS

I am grateful to Dr. Ali HAMICHE and Pr. Christophe TERZIAN for accepting to spend their precious time in reviewing my thesis manuscript and giving me their valuable suggestions. I am thankful to all

2004

). ASF1 is proposed to transfer nascent histones to CAF-1, the specialized histone deposition factor complex (Figure 20). Thus it represents the prototype of a histone donor chaperone that could effectively ensure a constant supply of histones at sites of nucleosome assembly [START_REF] Loyola | Histone chaperones, a supporting role in the limelight[END_REF].

Figure 20: The de novo histones H3-H4 assembly through the Asf1-CAF-1 pathway [START_REF] Jasencakova | Broken silence restored--remodeling primes for deacetylation at replication forks[END_REF]. See text for further details.

III. Histone variant H3.3, its chaperones and paternal chromatin assembly

A. Histone variant H3.3

Histone variant H3.3 differs from the canonical H3 by only four amino acid residues (Figure 21); three of these residues are clustered in the α2 helix of the histone fold domain and the other residue is in the N-terminal tail [START_REF] Hake | Histone H3 variants and their potential role in indexing mammalian genomes: the "H3 barcode hypothesis[END_REF][START_REF] Yuan | Histone variants and epigenetic inheritance[END_REF]. In spite of the high sequence similarity between H3.3 and H3, these specific residues have been proposed to account for particular properties of histone H3.3 [START_REF] Szenker | The double face of the histone variant H3.3[END_REF]. In the vertebrates and Drosophila, residues 87, 89 and 90 are S, V and M in H3, and A, I and G in H3.3 (Figure 21). The amino acid residues found at these positions in H3 and H3.3 vary between species but always distinguish H3 from H3.3. It is noteworthy that H3.3 histone variant is encoded by two intron-containing genes, h3.3A and h3.3B, the transcripts of which are regulated through polyadenylation and whose translation results in identical protein products [START_REF] Wellman | Characterization of mouse H3.3-like histone genes[END_REF]. H3.3 histones are enriched at transcriptionally active regions [START_REF] Ahmad | The histone variant H3.3 marks active chromatin by replication-independent nucleosome assembly[END_REF][START_REF] Henikoff | Genome-wide profiling of salt fractions maps physical properties of chromatin[END_REF][START_REF] Mito | Genome-scale profiling of histone H3.3 replacement patterns[END_REF] as well as telomeres and pericentromeric regions [START_REF] Drane | The death-associated protein DAXX is a novel histone chaperone involved in the replication-independent deposition of H3.3[END_REF][START_REF] Goldberg | Distinct factors control histone variant H3.3 localization at specific genomic regions[END_REF][START_REF] Wong | ATRX interacts with H3.3 in maintaining telomere structural integrity in pluripotent embryonic stem cells[END_REF][START_REF] Wong | Histone H3.3 incorporation provides a unique and functionally essential telomeric chromatin in embryonic stem cells[END_REF]. These studies indicate that histone variant H3.3 is incorporated in both transcriptionally active and inactive chromatin sites but it is still not known how these sites are functionally linked by its deposition [START_REF] Szenker | The double face of the histone variant H3.3[END_REF].

The dual nature of histone variant H3.3 makes it necessary for a variety of nuclear processes that specifically occur in germ cells and in early embryos development of metazoans [START_REF] Banaszynski | Histone variants in metazoan development[END_REF][START_REF] Ooi | Germline histone dynamics and epigenetics[END_REF][START_REF] Orsi | Epigenetic and replacement roles of histone variant H3.3 in reproduction and development[END_REF][START_REF] Szenker | The double face of the histone variant H3.3[END_REF]. In Xenopus laevis embryos gastrulation depends on the critical involvement of H3.3 [START_REF] Szenker | A developmental requirement for HIRA-dependent H3.3 deposition revealed at gastrulation in Xenopus[END_REF]. During mouse spermatogenesis, the majority of histone H3 is replaced by the variant H3.3 by the first meiotic prophase, where most of the mRNA required for the development of sperm is synthesized. It disappears just before complete inactivation of the genome, when the transition proteins and protamines replace almost all of histones [START_REF] Akhmanova | Structure and expression of histone H3.3 genes in Drosophila melanogaster and Drosophila hydei[END_REF]. Thus, the replacement of histone H3 by histone H3.3 could be important for the initiation and maintenance of an open chromatin during meiotic phase.

Many studies revealed that H3.3 is notably required for the proper segregation of meiotic chromosomes in spermatocytes and for the global organization of early spermatid chromatin [START_REF] Akhmanova | The localization of histone H3.3 in germ line chromatin of Drosophila males as established with a histone H3.3-specific antiserum[END_REF][START_REF] Sakai | Transcriptional and developmental functions of the H3.3 histone variant in Drosophila[END_REF]. The critical requirement of H3.3 after fertilization has also been extensively studied.

In Drosophila, H3.3 deficient flies are viable but both male and female are sterile [START_REF] Hodl | Transcription in the absence of histone H3.3[END_REF][START_REF] Sakai | Transcriptional and developmental functions of the H3.3 histone variant in Drosophila[END_REF].

Interestingly the substitution of the four amino acids which distinguish Drosophila H3.3 from its canonical one (S31A, A87S, I89Vand G90M), is not sufficient to rescue the fertility of H3.3 homozygous mutation; the fertility is rescued only after providing K4-methylated versions of H3.3 [START_REF] Hodl | Transcription in the absence of histone H3.3[END_REF]. This suggests that not only the specific mere requirement of variant histone H3.3 but also K4 methylation of its residue are critical. In a more recent study, the same authors found that the null mutant flies for both H3.3 genes (His3.3A and His3.3B) rescued their fertility when they were provided with H3.2 expressed under the H3.3B promoter which suggests that the major difference between canonical and variant H3 histones resides in their mode of transcriptional regulation [START_REF] Hodl | Transcription in the Absence of Histone H3.2 and H3K4 Methylation[END_REF]. These results illustrate the complexity of the picture and the necessity of further investigations to fully understand the role of H3.3 and its epigenetic modifications. H3.3 variant deposition is mediated by a distinct group of histone chaperone complexes.

B. Histone variant H3.3 chaperones

The double incorporation of H3.3 in both transcriptionally silent and active chromatin is supported by the presence of two main histone chaperone complexes that mediate the H3.3 deposition. These two complexes are DAXX-ATRX and HIRA complexes.

DAXX-ATRX complex

The death domain-associated protein (DAXX) was shown to achieve H3.3 enrichment in heterochromatin [START_REF] Wong | ATRX interacts with H3.3 in maintaining telomere structural integrity in pluripotent embryonic stem cells[END_REF]. DAXX was first shown by Yang in 1997 to induce cell apoptosis through the cJun N-terminal kinase (JNK) pathway activation [START_REF] Yang | Daxx, a novel Fas-binding protein that activates JNK and apoptosis[END_REF]. It was initially identified as a cytoplasmic protein that interacts with FAS (fibroblast-associated surface antigen) [START_REF] Salomoni | Daxx: death or survival protein?[END_REF], while some studies reported that DAXX resides primarily in the nucleus and participates in transcriptional regulation [START_REF] Lanotte | NB4, a maturation inducible cell line with t(15;17) marker isolated from a human acute promyelocytic leukemia (M3)[END_REF]. In fact DAXX subcellular localization is submitted to regulation [START_REF] Chen | Progress on the subcellular localization of Daxx[END_REF][START_REF] Yeung | Daxx contains two nuclear localization signals and interacts with importin alpha3[END_REF]. DAXX protein cooperates with the SWI/SNF like ATP-dependent chromatin remodeling factor, α-thalassemia/mental retardation X-linked syndrome protein (ATRX) [START_REF] Gibbons | Mutations in a putative global transcriptional regulator cause X-linked mental retardation with alpha-thalassemia (ATR-X syndrome)[END_REF]. ATRX protein contains an N-terminal ADD (ATRX-DNMT3-DNMT3L domain [START_REF] Argentaro | Structural consequences of disease-causing mutations in the ATRX-DNMT3-DNMT3L (ADD) domain of the chromatin-associated protein ATRX[END_REF] and a C-terminal ATPase domain, which function in the regulation of its binding and enzymatic activities, respectively [START_REF] Dhayalan | The ATRX-ADD domain binds to H3 tail peptides and reads the combined methylation state of K4 and K9[END_REF]. The ATRX ADD

Results and Discussion

Conclusion and Future Directions