Hayat Hage

Marie-Noëlle Rosso

Lionel Tarrago
email: lionel.tarrago@inrae.fr

Distribution of methionine sulfoxide reductases in fungi and conservation of the freemethionine-R-sulfoxide reductase in multicellular eukaryotes

Keywords: fungi, genome, horizontal gene transfer, methionine sulfoxide, methionine sulfoxide reductase, protein oxidation, thiol oxidoreductase

à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

Life in the presence of dioxygen necessarily exposes the biological systems to oxidant molecules. Due to its high reactivity, dioxygen can be converted to reactive oxygen species (ROS), which play key roles in physiological and pathological contexts through redox modifications of macromolecules [START_REF] Sies | Reactive oxygen species (ROS) as pleiotropic physiological signalling agents[END_REF]. Met, either as a free amino acid or as a residue included in a protein, can be oxidized into Met sulfoxide (MetO) by the addition of an oxygen atom to the sulfur of the lateral chain. MetO exists as diastereomer R or S (Met-R-O and Met-S-O, respectively) [START_REF] Sharov | Diastereoselective reduction of proteinbound methionine sulfoxide by methionine sulfoxide reductase[END_REF], and can be reduced back to Met by the action of oxidoreductases called methionine sulfoxide reductases (Msr). The two main types are the (seleno)thiol-containing MsrA and MsrB, which display strict stereoselectivities toward the S-and the R-diastereomer of MetO, respectively [START_REF] Brot | Enzymatic reduction of protein-bound methionine sulfoxide[END_REF][START_REF] Grimaud | Repair of oxidized proteins. Identification of a new methionine sulfoxide reductase[END_REF][START_REF] Sharov | Diastereoselective protein methionine oxidation by reactive oxygen species and diastereoselective repair by methionine sulfoxide reductase, Free Radic[END_REF]. Whereas MsrA reduces free MetO and protein-bound MetO with similar catalytic efficiencies, MsrB generally only reduces efficiently the protein-bound MetO [START_REF] Tarrago | Methionine sulfoxide reductases preferentially reduce unfolded oxidized proteins and protect cells from oxidative protein unfolding[END_REF]. Another enzyme, fRMsr (free methionine-(R)-sulfoxide reductase) specifically reduces the free form of Met-R-O [START_REF] Gruez | Structural and biochemical characterization of free methionine-R-sulfoxide reductase from Neisseria meningitidis[END_REF][START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF][START_REF] Lin | Free methionine-(R)sulfoxide reductase from Escherichia coli reveals a new GAF domain function[END_REF]. Some bacteria also possess several molybdoenzymes that reduce exclusively the free MetO [START_REF] Dhouib | A Novel, Molybdenum-Containing Methionine Sulfoxide Reductase Supports Survival of Haemophilus influenzae in an In vivo Model of Infection[END_REF][START_REF] Ezraty | Methionine sulfoxide reduction and assimilation in Escherichia coli: new role for the biotin sulfoxide reductase BisC[END_REF] or both the free and protein-bound MetO [START_REF] Andrieu | Characterisation of the periplasmic methionine sulfoxide reductase (MsrP) from Salmonella Typhimurium, Free Radic[END_REF][START_REF] Gennaris | Repairing oxidized proteins in the bacterial envelope using respiratory chain electrons[END_REF][START_REF] Tarrago | Reduction of Protein Bound Methionine Sulfoxide by a Periplasmic Dimethyl Sulfoxide Reductase[END_REF][START_REF] Tarrago | Rhodobactersphaeroides methionine sulfoxide reductase P reduces R-and S-diastereomers of methionine sulfoxide from a broad-spectrum of protein substrates[END_REF]. Despite the lack of sequence and structure similarities, MsrA, MsrB and fRMsr generally catalyze the reduction of MetO using a similar 3-steps-mechanism [START_REF] Lin | Free methionine-(R)sulfoxide reductase from Escherichia coli reveals a new GAF domain function[END_REF][START_REF] Lowther | The mirrored methionine sulfoxide reductases of Neisseria gonorrhoeae pilB[END_REF] : i) a 'catalytic' Cys (or, less frequently, a selenocysteine, Sec) reduces the target MetO and is converted into a sulfenic (or selenic) acid [START_REF] Boschi-Muller | A sulfenic acid enzyme intermediate is involved in the catalytic mechanism of peptide methionine sulfoxide reductase from Escherichia coli[END_REF], ii) an internal 'resolving' Cys reduces it through the formation of an intramolecular disulfide bond, and finally iii) the oxidized Msr is regenerated through disulfide exchange with a thioredoxin [START_REF] Tarrago | Recharging oxidative protein repair: catalysis by methionine sulfoxide reductases towards their amino acid, protein, and model substrates[END_REF]. Variations of this mechanism exist, with some Msrs devoid of any resolving Cys, in which the sulfenic acid is directly reduced by an external reducer [START_REF] Kim | Glutaredoxin serves as a reductant for methionine sulfoxide reductases with or without resolving cysteine[END_REF][START_REF] Tarrago | Plant thioredoxin CDSP32 regenerates 1-cys methionine sulfoxide reductase B activity through the direct reduction of sulfenic acid[END_REF][START_REF] Tarrago | Regeneration mechanisms of Arabidopsis thaliana methionine sulfoxide reductases B by glutaredoxins and thioredoxins[END_REF]. Alternatively, in other Msrs, a disulfide exchange occurs with a second internal resolving Cys before regeneration by the thioredoxin [START_REF] Gruez | Structural and biochemical characterization of free methionine-R-sulfoxide reductase from Neisseria meningitidis[END_REF][START_REF] Lowther | Thiol-disulfide exchange is involved in the catalytic mechanism of peptide methionine sulfoxide reductase[END_REF][START_REF] Rouhier | Functional and structural aspects of poplar cytosolic and plastidial type a methionine sulfoxide reductases[END_REF]. Genome mining analyses indicated that msrA and msrB genes originated from early prokaryotes and are now present in all organisms across the tree of life, with few exceptions [START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF][START_REF] Tarrago | Protein-repairing methionine sulfoxide reductases in photosynthetic organisms: gene organization, reduction mechanisms, and physiological roles[END_REF][START_REF] Zhang | Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases[END_REF]. Most organisms have few msr genes, generally one of each type [START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF][START_REF] Tarrago | Protein-repairing methionine sulfoxide reductases in photosynthetic organisms: gene organization, reduction mechanisms, and physiological roles[END_REF][START_REF] Zhang | Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases[END_REF], but this number is generally higher in plants (e.g. up to 5 msrA and 9 msrB genes in Arabidopsis thaliana [START_REF] Tarrago | Protein-repairing methionine sulfoxide reductases in photosynthetic organisms: gene organization, reduction mechanisms, and physiological roles[END_REF]). Moreover, some bacteria encode a bifunctional MsrA/MsrB fusion and some others lack a MsrB, but no organism of any kind was so far described with only a MsrB [START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF][START_REF] Zhang | Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases[END_REF]. Finally, only very few organisms do not possess any msr gene at all, such as a few endosymbiotic or obligatory parasitic bacteria and some archaea [START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF][START_REF] Zhang | Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases[END_REF]. fRMsr were so far only reported in bacteria and a few unicellular eukaryotes [START_REF] Gruez | Structural and biochemical characterization of free methionine-R-sulfoxide reductase from Neisseria meningitidis[END_REF][START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF][START_REF] Lin | Free methionine-(R)sulfoxide reductase from Escherichia coli reveals a new GAF domain function[END_REF]. Most interestingly, the only eukaryote for which the absence of msr gene was observed is the fungus Encephalitozoon cuniculi, a microsporidium having a remarkably reduced genome (∼2.9 Mb) and living as intracellular parasite of mammals [START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF][START_REF] Katinka | Genome sequence and gene compaction of the eukaryote parasite Encephalitozoon cuniculi[END_REF].

The conservation of typical MsrAs and MsrBs in almost all known organisms argues for the critical role of MetO reduction in cellular metabolism, and numerous studies showed that Msrs are involved in the protection against oxidative stress and the regulation of protein functions. Schematically, Msr protective roles against oxidative injuries occur through two main functions: i) the repair of oxidized proteins, and ii) an antioxidant function through ROS scavenging by cyclic Met oxidation and reduction. Moreover, the reversible Met oxidation was shown to act as a post-translational modification responsible for the activation of enzymes and transcription factors or the regulation of protein-protein interactions [START_REF] Drazic | Methionine oxidation activates a transcription factor in response to oxidative stress[END_REF][START_REF] Erickson | A dynamic pathway for calciumindependent activation of CaMKII by methionine oxidation[END_REF][START_REF] Lee | MsrB1 and MICALs regulate actin assembly and macrophage function via reversible stereoselective methionine oxidation[END_REF].

The role of fRMsr has been far less studied, but it is suspected to have an antioxidant function by reducing free MetO and maintaining the pool of Met available for protein synthesis and for the production of sulfur-containing metabolites [START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF][START_REF] Lee | Expression of the methionine sulfoxide reductase lost during evolution extends Drosophila lifespan in a methionine-dependent manner[END_REF]. Overall, these functions were well established in animals, bacteria and plants, and have recently been reviewed [START_REF] Jiang | The Functions of the Mammalian Methionine Sulfoxide Reductase System and Related Diseases[END_REF][START_REF] Lim | Methionine in Proteins: It's Not Just for Protein Initiation Anymore[END_REF][34][START_REF] Rey | Physiological Roles of Plant Methionine Sulfoxide Reductases in Redox Homeostasis and Signaling[END_REF][START_REF] Singh | The Role of Methionine Sulfoxide Reductases in Oxidative Stress Tolerance and Virulence of Staphylococcus aureus and Other Bacteria[END_REF].

Fungi are heterotrophic eukaryotes that colonize virtually all niches on Earth and exist as unicellular or multicellular organisms. They can have numerous lifestyles, either as free living organisms playing key roles in organic and inorganic matter cycling, or as symbionts or pathogens with crucial impacts on plant and animal health [START_REF] Naranjo-Ortiz | Fungal evolution: diversity, taxonomy and phylogeny of the Fungi[END_REF]. As any other organism living in aerobic conditions and exposed to environmental constraints, fungi are exposed to oxidative constraints and protein oxidation [START_REF] Breitenbach | Oxidative Stress in Fungi: Its Function in Signal Transduction, Interaction with Plant Hosts, and Lignocellulose Degradation[END_REF]. However, the effects of Met oxidation and the roles of Msrs were largely overlooked in these eukaryotes. The Msr system was mainly characterized in the yeast Saccharomyces cerevisiae, which possesses one Msr of each type [START_REF] Tarrago | Methionine sulfoxide reductases preferentially reduce unfolded oxidized proteins and protect cells from oxidative protein unfolding[END_REF][START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF]. In S. cerevisiae, MsrA is located in the cytosol, MsrB is found in mitochondria and in the cytosol and fRMsr is located both in the cytosol and the nucleus [START_REF] Kaya | Compartmentalization and regulation of mitochondrial function by methionine sulfoxide reductases in yeast[END_REF][START_REF] Nicklow | Activity of the yeast cytoplasmic Hsp70 nucleotide-exchange factor Fes1 is regulated by reversible methionine oxidation[END_REF]. Genetic manipulations have shown that these Msrs are involved in the protection against oxidative stress and in maintaining the yeast lifespan [START_REF] Tarrago | Methionine sulfoxide reductases preferentially reduce unfolded oxidized proteins and protect cells from oxidative protein unfolding[END_REF][START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF][START_REF] Koc | Methionine sulfoxide reductase regulation of yeast lifespan reveals reactive oxygen species-dependent and -independent components of aging[END_REF][START_REF] Moskovitz | Overexpression of peptide-methionine sulfoxide reductase in Saccharomyces cerevisiae and human T cells provides them with high resistance to oxidative stress[END_REF]. Consistently, the overexpression of msrA in the basidiomycete Pleurotus ostreatus and of msrB in the yeast Schizosaccharomyces pombe increased the resistance to oxidative constraints [START_REF] Jo | Protective roles of methionine-R-sulfoxide reductase against stresses in Schizosaccharomyces pombe[END_REF][START_REF] Yin | Enhancing stress tolerance by overexpression of a methionine sulfoxide reductase A (MsrA) gene in Pleurotus ostreatus[END_REF]. Moreover, the filamentous fungus Aspergillus nidulans possesses one msrA and one msrB genes, and the deletion of one or both genes increased the sensitivity of the fungus to oxidative treatments [START_REF] Soriani | Functional characterization of the Aspergillus nidulans methionine sulfoxide reductases (msrA and msrB)[END_REF]. Few proteins with oxidized Met residues have been characterized in fungi, but remarkably interesting findings were obtained. For instance, in S. cerevisiae, the reversible oxidation of Met regulates the oligomerization state of the ataxin-2 protein and the activity of the co-chaperone Fes1 [START_REF] Nicklow | Activity of the yeast cytoplasmic Hsp70 nucleotide-exchange factor Fes1 is regulated by reversible methionine oxidation[END_REF][START_REF] Kato | Redox State Controls Phase Separation of the Yeast Ataxin-2 Protein via Reversible Oxidation of Its Methionine-Rich Low-Complexity Domain[END_REF]. Moreover, Met oxidation enhanced the activity of an α-galactosidase in Trichoderma reesei [START_REF] Kachurin | Role of methionine in the active site of alpha-galactosidase from Trichoderma reesei[END_REF]. Finally, in A. nidulans, it was demonstrated that the nuclear localization of the nitrate-responsive transcription factor NirA was actively regulated through cyclic Met oxidation [START_REF] Gallmetzer | Reversible Oxidation of a Conserved Methionine in the Nuclear Export Sequence Determines Subcellular Distribution and Activity of the Fungal Nitrate Regulator NirA[END_REF]. These data indicate that, as for other organisms, fungal Msrs certainly play key roles under many conditions of oxidative stress, such as biotic interactions or abiotic constraints.

In this study, we searched for msr genes in about 700 fungal genomes. We show that the great majority of fungi have one msrA and one msrB genes. Moreover, we identified fRmsr genes in almost all the fungi we analyzed and thereby demonstrate that the enzyme is conserved in these multicellular eukaryotes. Finally, using a phylogenetic analysis and a close inspection of the sequence features, we identified fungal Msrs with unusual sequence characteristics and uncovered horizontal gene transfers from bacteria to fungi.

Material and methods

Search for Msr homologs in fungi.

The protein sequence of MsrA (Uniprot accession # C8Z745), MsrB (Uniprot accession # P25566) and fRMsr (Uniprot accession # P36088) from S. cerevisiae, MsrP (Uniprot accession # P76342) and BisC (Uniprot accession # P20099) from Escherichia coli, TorZ (Uniprot accession # P44798) from Haemophilus influenzae, and DorA (Uniprot accession # Q57366) from Rhodobacter sphaeroides were used as BLASTP and TBLASTN [START_REF] Altschul | Basic local alignment search tool[END_REF] queries to identify msr genes in 683 genomes available in the MycoCosm database [START_REF] Grigoriev | MycoCosm portal: gearing up for 1000 fungal genomes[END_REF] (https://mycocosm.jgi.doe.gov/mycocosm/home).

List of all explored fungal genomes.

Aaosphaeria arxii CBS 175.79 Rhizoscyphus ericae UAMH 7357 v1.0 [START_REF] Martino | Comparative genomics and transcriptomics depict ericoid mycorrhizal fungi as versatile saprotrophs and plant mutualists[END_REF]; Anaeromyces robustus v1.0; Neocallimastix californiae G1 v1.0; Piromyces finnis v3.0; Piromyces sp. E2 v1.0 [START_REF] Haitjema | A parts list for fungal cellulosomes revealed by comparative genomics[END_REF]; Antonospora locustae HM-2013 [START_REF] Slamovits | Genome compaction and stability in microsporidian intracellular parasites[END_REF]; Armillaria cepistipes B5; Armillaria gallica 21-2 v1.0; Armillaria ostoyae C18/9; Armillaria solidipes 28-4 v1.0 [START_REF] Sipos | Genome expansion and lineage-specific genetic innovations in the forest pathogenic fungi Armillaria[END_REF]; Armillaria mellea DSM 3731 [START_REF] Collins | Genomic and proteomic dissection of the ubiquitous plant pathogen, Armillaria mellea: toward a new infection model system[END_REF]; Arthrobotrys oligospora ATCC 24927 [START_REF] Yang | Genomic and proteomic analyses of the fungus Arthrobotrys oligospora provide insights into nematode-trap formation[END_REF];

Ascobolus immersus RN42 v1.0; Choiromyces venosus 120613-1 v1.0; Morchella importuna CCBAS932 v1.0; Terfezia boudieri ATCC MYA-4762 v1.1; Tuber aestivum var. urcinatum v1.0; Tuber borchii Tbo3840 v1.0; Tuber magnatum v1.0 [START_REF] Murat | Pezizomycetes genomes reveal the molecular basis of ectomycorrhizal truffle lifestyle[END_REF]; Ascochyta rabiei ArDII [START_REF] Verma | Draft genome sequencing and secretome analysis of fungal phytopathogen Ascochyta rabiei provides insight into the necrotrophic effector repertoire[END_REF]; Ascocoryne sarcoides NRRL50072 [START_REF] Gianoulis | Genomic analysis of the hydrocarbon-producing, cellulolytic, endophytic fungus Ascocoryne sarcoides[END_REF]; Ascodesmis nigricans CBS 389.68 [START_REF] Moore | Draft Genome Sequence of an Aflatoxigenic Aspergillus Species, A. bombycis[END_REF]; Aspergillus calidoustus [START_REF] Horn | Draft Genome Sequences of Fungus Aspergillus calidoustus[END_REF]; Aspergillus campestris IBT 28561 v1.0; CBS 109695 v1.0; Laetiporus sulphureus var. sulphureus v1.0; Neolentinus lepideus v1.0; Peniophora sp. v1.0; Sistotremastrum niveocremeum HHB9708 ss-1 1.0; Sistotremastrum suecicum v1.0; Calocera cornea v1.0; Calocera viscosa v1.0 [START_REF] Nagy | Comparative Genomics of Early-Diverging Mushroom-Forming Fungi Provides Insights into the Origins of Lignocellulose Decay Capabilities[END_REF]; Daldinia eschscholtzii EC12 v1.0; Hypoxylon sp. CI-4A v1.0; Hypoxylon sp. CO27-5 v1.0; Hypoxylon sp. EC38 v3.0 [START_REF] Wu | Characterization of four endophytic fungi as potential consolidated bioprocessing hosts for conversion of lignocellulose into advanced biofuels[END_REF]; Debaryomyces hansenii [START_REF] Sacerdot | Promiscuous DNA in the nuclear genomes of hemiascomycetous yeasts[END_REF]; Dekkera bruxellensis CBS 2499 v2.0 [START_REF] Piškur | The genome of wine yeast Dekkera bruxellensis provides a tool to explore its food-related properties[END_REF]; Dentipellis sp. KUC8613 v1.0 [START_REF] Park | Comprehensive genomic and transcriptomic analysis of polycyclic aromatic hydrocarbon degradation by a mycoremediation fungus, Dentipellis sp. KUC8613[END_REF]; Dichomitus squalens CBS463.89 v1.0; Dichomitus squalens CBS464.89 v1.0; Dichomitus squalens OM18370.1 v1.0 [START_REF] Casado López | Draft Genome Sequences of Three Monokaryotic Isolates of the White-Rot Basidiomycete Fungus Dichomitus squalens[END_REF];

Encephalitozoon cuniculi GB-M1 [START_REF] Peyretaillade | Identification of transcriptional signals in Encephalitozoon cuniculi widespread among Microsporidia phylum: support for accurate structural genome annotation[END_REF]; Encephalitozoon hellem ATCC 50504; Encephalitozoon romaleae SJ-2008 [START_REF] Pombert | Gain and loss of multiple functionally related, horizontally transferred genes in the reduced genomes of two microsporidian parasites[END_REF]; Encephalitozoon intestinalis ATCC 50506 [START_REF] Corradi | The complete sequence of the smallest known nuclear genome from the microsporidian Encephalitozoon intestinalis[END_REF]; Endocarpon pusillum Z07020 [START_REF] Wang | Genome characteristics reveal the impact of lichenization on lichenforming fungus Endocarpon pusillum Hedwig (Verrucariales, Ascomycota)[END_REF]; Endogone sp FLAS 59071; Jimgerdemannia flammicorona AD002; Jimgerdemannia flammicorona GMNB39; Jimgerdemannia lactiflua OSC166217 [START_REF] Chang | Phylogenomics of Endogonaceae and evolution of mycorrhizas within Mucoromycota[END_REF]; Enterocytozoon bieneusi H348 [START_REF] Akiyoshi | Genomic survey of the non-cultivatable opportunistic human pathogen, Enterocytozoon bieneusi[END_REF]; Eremothecium gossypii ATCC 10895 [START_REF] Dietrich | The Ashbya gossypii genome as a tool for mapping the ancient Saccharomyces cerevisiae genome[END_REF]; Erysiphe necator c [START_REF] Jones | Adaptive genomic structural variation in the grape powdery mildew pathogen, Erysiphe necator[END_REF]; Eurotium rubrum v1.0 [START_REF] Kis-Papo | Genomic adaptations of the halophilic Dead Sea filamentous fungus Eurotium rubrum[END_REF]; Exophiala dermatitidis UT8656 [START_REF] Chen | Comparative genomic and transcriptomic analysis of wangiella dermatitidis, a major cause of phaeohyphomycosis and a model black yeast human pathogen[END_REF]; Fibroporia radiculosa TFFH 294 [START_REF] Tang | Short-read sequencing for genomic analysis of the brown rot fungus Fibroporia radiculosa[END_REF]; Fonsecaea monophora CBS 269.37 [START_REF] Bombassaro | Draft Genome Sequence of Fonsecaea monophora Strain CBS 269.37, an Agent of Human Chromoblastomycosis[END_REF]; Fonsecaea nubica CBS 269.64 [START_REF] Costa | Draft Genome Sequence of Fonsecaea nubica Strain CBS 269.64, Causative Agent of Human Chromoblastomycosis[END_REF]; Fusarium fujikuroi IMI 58289 [START_REF] Wiemann | Deciphering the cryptic genome: genome-wide analyses of the rice pathogen Fusarium fujikuroi reveal complex regulation of secondary metabolism and novel metabolites[END_REF]; Fusarium graminearum v1.0 [START_REF] Cuomo | The Fusarium graminearum genome reveals a link between localized polymorphism and pathogen specialization[END_REF]; Fusarium oxysporum f. sp. conglutinans race 2 54008 (PHW808); Fusarium oxysporum f. sp. cubense tropical race 4 54006 (II5); Fusarium oxysporum f. sp. lycopersici MN25 (FoMN25) NRRL 54003; Fusarium oxysporum f. sp. radicis-lycopersici 26381 (CL57); Fusarium oxysporum f. sp. raphani 54005; Fusarium oxysporum f. sp. vasinfectum 25433 (Cotton); Fusarium oxysporum Fo47; Fusarium oxysporum NRRL 32931 [START_REF] Deiulio | Kinome Expansion in the Fusarium oxysporum Species Complex Driven by Accessory Chromosomes[END_REF]; Fusarium oxysporum f. sp. lycopersici 4287 v2; Fusarium verticillioides 7600 v2 [START_REF] Ma | Comparative genomics reveals mobile pathogenicity chromosomes in Fusarium[END_REF]; Fusarium oxysporum f. sp. melonis (FoMelon) NRRL 26406 [START_REF] Ma | Genome Sequence of Fusarium oxysporum f. sp. melonis Strain NRRL 26406, a Fungus Causing Wilt Disease on Melon[END_REF]; Fusarium oxysporum f. sp. pisi HDV247 [START_REF] Williams | Comparative genomics and prediction of conditionally dispensable sequences in legume-infecting Fusarium oxysporum formae speciales facilitates identification of candidate effectors[END_REF]; Fusarium pseudograminearum CS3096 [START_REF] Gardiner | Comparative pathogenomics reveals horizontally acquired novel virulence genes in fungi infecting cereal hosts[END_REF]; Gaeumannomyces graminis var. tritici R3-111a-1; Magnaporthiopsis poae ATCC 64411 [START_REF] Okagaki | Genome Sequences of Three Phytopathogenic Species of the Magnaporthaceae Family of Fungi[END_REF]; Glarea lozoyensis ATCC 20868 [START_REF] Chen | Genomics-driven discovery of the pneumocandin biosynthetic gene cluster in the fungus Glarea lozoyensis[END_REF]; Grosmannia clavigera kw1407 [START_REF] Diguistini | Genome and transcriptome analyses of the mountain pine beetle-fungal symbiont Grosmannia clavigera, a lodgepole pine pathogen[END_REF]; Gymnopus androsaceus JB14 v1.0; Chalara longipes BDJ v1.0 [START_REF] Barbi | Fungal ecological strategies reflected in gene transcription -a case study of two litter decomposers[END_REF]; Heterobasidion annosum v2.0 [START_REF] Olson | Insight into trade-off between wood decay and parasitism from the genome of a fungal forest pathogen[END_REF]; Homolaphlyctis polyrhiza JEL142 v1.0 [START_REF] Joneson | Genomic transition to pathogenicity in chytrid fungi[END_REF]; Hortaea werneckii EXF-2000 M0 v1.0 [START_REF] Lenassi | Whole genome duplication and enrichment of metal cation transporters revealed by de novo genome sequencing of extremely halotolerant black yeast Hortaea werneckii[END_REF]; Ilyonectria sp. v1.0 [START_REF] Liao | Fungal Endophytes of Populus trichocarpa Alter Host Phenotype, Gene Expression, and Rhizobiome Composition[END_REF]; Kazachstania africana CBS 2517; Torulaspora delbrueckii CBS 1146 [START_REF] Gordon | Evolutionary erosion of yeast sex chromosomes by mating-type switching accidents[END_REF]; Kluyveromyces lactis; Yarrowia lipolytica (strain CLIB122) [START_REF] Dujon | Genome evolution in yeasts[END_REF]; Kuraishia capsulata CBS 1993 [START_REF] Morales | Complete DNA sequence of Kuraishia capsulata illustrates novel genomic features among budding yeasts (Saccharomycotina)[END_REF]; Laccaria bicolor v2.0 [START_REF] Martin | The genome of Laccaria bicolor provides insights into mycorrhizal symbiosis[END_REF]; Lentinula edodes B17 v1.1 [START_REF] Park | Long-read transcriptome data for improved gene prediction in Lentinula edodes[END_REF];

Lentinula edodes W1-26 v1.0 [START_REF] Chen | Genome Sequence of the Edible Cultivated Mushroom Lentinula edodes (Shiitake) Reveals Insights into Lignocellulose Degradation[END_REF]; Lentinus tigrinus ALCF2SS1-6 v1.0; Lentinus tigrinus ALCF2SS1-7 v1.0 [START_REF] Wu | Genomics and Development of Lentinus tigrinus: A White-Rot Wood-Decaying Mushroom with Dimorphic Fruiting Bodies[END_REF]; Leptosphaeria maculans [START_REF] Rouxel | Effector diversification within compartments of the Leptosphaeria maculans genome affected by Repeat-Induced Point mutations[END_REF]; Leucoagaricus gongylophorus Ac12 [START_REF] Aylward | Leucoagaricus gongylophorus produces diverse enzymes for the degradation of recalcitrant plant polymers in leaf-cutter ant fungus gardens[END_REF]; v2.0; Rhizoclosmatium globosum JEL800 v1.0 [START_REF] Mondo | Widespread adenine N6-methylation of active genes in fungi[END_REF]; Lichtheimia corymbifera JMRC:FSU:9682 [START_REF] Schwartze | Gene expansion shapes genome architecture in the human pathogen Lichtheimia corymbifera: an evolutionary genomics analysis in the ancient terrestrial mucorales (Mucoromycotina)[END_REF]; Macrophomina phaseolina MS6 [START_REF] Islam | Tools to kill: genome of one of the most destructive plant pathogenic fungi Macrophomina phaseolina[END_REF]; Magnaporthe oryzae 70-15 v3.0 [START_REF] Dean | The genome sequence of the rice blast fungus Magnaporthe grisea[END_REF]; Malassezia globosa CBS 131958 [START_REF] Moreno | Draft Genome Sequence of the Ant-Associated Fungus Phialophora attae (CBS 131958)[END_REF]; Phlebia centrifuga FBCC195 [START_REF] Mäkelä | Draft Genome Sequence of the Basidiomycete White-Rot Fungus Phlebia centrifuga[END_REF]; Phlebia radiata Fr. (isolate 79, FBCC0043) [START_REF] Kuuskeri | Time-scale dynamics of proteome and transcriptome of the white-rot fungus Phlebia radiata: growth on spruce wood and decay effect on lignocellulose[END_REF]; Phlebiopsis gigantea v1.0 [START_REF] Hori | Analysis of the Phlebiopsis gigantea genome, transcriptome and secretome provides insight into its pioneer colonization strategies of wood[END_REF]; Phyllosticta capitalensis CBS 128856 v1.0; Phyllosticta citriasiana CBS 120486 v1.0; Phyllosticta citribraziliensis CBS 100098 v1.0; Phyllosticta citricarpa CBS 127454 v1.0; Phyllosticta citrichinaensis CBS 130529 v1.0; Phyllosticta paracitricarpa CBS 141357 v1.0; Phyllosticta sp. CPC 27913 v1.0 [START_REF] Guarnaccia | Phyllosticta citricarpa and sister species of global importance to Citrus[END_REF]; Pichia kudriavzevii CBS573 [START_REF] Douglass | Population genomics shows no distinction between pathogenic Candida krusei and environmental Pichia kudriavzevii: One species, four names[END_REF]; Pichia pastoris [START_REF] Schutter | Genome sequence of the recombinant protein production host Pichia pastoris[END_REF]; Piriformospora indica DSM 11827 from MPI [START_REF] Zuccaro | Endophytic life strategies decoded by genome and transcriptome analyses of the mutualistic root symbiont Piriformospora indica[END_REF]; Pleurotus ostreatus PC9 v1.0 [START_REF] Alfaro | Comparative and transcriptional analysis of the predicted secretome in the lignocellulosedegrading basidiomycete fungus Pleurotus ostreatus[END_REF];

Leucosporidiella
Pneumocystis jirovecii; [START_REF] Cissé | De novo assembly of the Pneumocystis jirovecii genome from a single bronchoalveolar lavage fluid specimen from a patient[END_REF]; Pochonia chlamydosporia 170 [START_REF] Wang | Biosynthesis of Antibiotic Leucinostatins in Bio-control Fungus Purpureocillium lilacinum and Their Inhibition on Phytophthora Revealed by Genome Mining[END_REF]; Podospora anserina S mat+ [START_REF] Espagne | The genome sequence of the model ascomycete fungus Podospora anserina[END_REF];

Polyporus brumalis BRFM 1820 v1.0 [START_REF] Miyauchi | Integrative visual omics of the white-rot fungus Polyporus brumalis exposes the biotechnological potential of its oxidative enzymes for delignifying raw plant biomass[END_REF]; Postia placenta MAD 698-R v1.0 [START_REF] Martinez | Genome, transcriptome, and secretome analysis of wood decay fungus Postia placenta supports unique mechanisms of lignocellulose conversion[END_REF]; Postia placenta MAD-698-R-SB12 v1.0 [START_REF] Gaskell | Draft genome sequence of a monokaryotic model brown-rot fungus Postia (Rhodonia) placenta SB12[END_REF]; Pseudocercospora (Mycosphaerella) fijiensis v2.0 [START_REF] Arango Isaza | Combating a Global Threat to a Clonal Crop: Banana Black Sigatoka Pathogen Pseudocercospora fijiensis (Synonym Mycosphaerella fijiensis) Genomes Reveal Clues for Disease Control[END_REF];

Pseudogymnoascus destructans 20631-21 [START_REF] Drees | Use of Multiple Sequencing Technologies To Produce a High-Quality Genome of the Fungus Pseudogymnoascus destructans, the Causative Agent of Bat White-Nose Syndrome[END_REF]; Pseudozyma antarctica T-34 [START_REF] Morita | Genome Sequence of the Basidiomycetous Yeast Pseudozyma antarctica T-34, a Producer of the Glycolipid Biosurfactants Mannosylerythritol Lipids[END_REF]; Pseudozyma hubeiensis SY62 [START_REF] Konishi | Draft Genome Sequence of the Basidiomycetous Yeast-Like Fungus Pseudozyma hubeiensis SY62, Which Produces an Abundant Amount of the Biosurfactant Mannosylerythritol Lipids[END_REF]; Psilocybe cubensis v1.0; Psilocybe serbica v1.0 [START_REF] Fricke | Enzymatic Synthesis of Psilocybin[END_REF]; Puccinia coronata avenae 12NC29; Puccinia coronata avenae 12SD80 [START_REF] Nazareno | Puccinia coronata f. sp. avenae: a threat to global oat production[END_REF]; Puccinia graminis f. sp. tritici 21-0 haplotype A; Puccinia graminis f. sp. tritici 21-0 haplotype B; Puccinia graminis f. sp. tritici Ug99 haplotype A; Puccinia graminis f. sp. tritici Ug99 haplotype C [START_REF] Li | Emergence of the Ug99 lineage of the wheat stem rust pathogen through somatic hybridisation[END_REF]; Puccinia striiformis f. sp. tritici 104 E137 A- [START_REF] Schwessinger | A Near-Complete Haplotype-Phased Genome of the Dikaryotic Wheat Stripe Rust Fungus Puccinia striiformis f. sp. tritici Reveals High Interhaplotype Diversity[END_REF]; Puccinia striiformis f. sp. tritici PST-130 [START_REF] Cantu | Next generation sequencing provides rapid access to the genome of Puccinia striiformis f. sp. tritici, the causal agent of wheat stripe rust[END_REF]; Puccinia striiformis f. sp. tritici PST-78 v1.0; Puccinia triticina 1-1 BBBD Race 1 [START_REF] Cuomo | Comparative Analysis Highlights Variable Genome Content of Wheat Rusts and Divergence of the Mating Loci[END_REF]; Pycnoporus cinnabarinus BRFM 137 [START_REF] Levasseur | The genome of the white-rot fungus Pycnoporus cinnabarinus: a basidiomycete model with a versatile arsenal for lignocellulosic biomass breakdown[END_REF];

Pycnoporus coccineus BRFM 310 v1.0; Pycnoporus puniceus CIRM-BRFM 1868 v1.0; Pycnoporus sanguineus BRFM 1264 v1.0; Ramaria rubella (R. acris) UT-36052-T v1.0 [START_REF] Miyauchi | Conserved white-rot enzymatic mechanism for wood decay in the Basidiomycota genus Pycnoporus[END_REF]; Pyrenophora teres f. teres [START_REF] Ellwood | A first genome assembly of the barley fungal pathogen Pyrenophora teres f. teres[END_REF]; Pyrenophora tritici-repentis [START_REF] Manning | Comparative genomics of a plant-pathogenic fungus, Pyrenophora tritici-repentis, reveals transduplication and the impact of repeat elements on pathogenicity and population divergence[END_REF]; Pyronema confluens CBS100304 [START_REF] Traeger | The genome and development-dependent transcriptomes of Pyronema confluens: a window into fungal evolution[END_REF]; Rhizoctonia solani AG-1 IB [START_REF] Wibberg | Establishment and interpretation of the genome sequence of the phytopathogenic fungus Rhizoctonia solani AG1-IB isolate 7/3/14[END_REF]; Rhizophagus irregularis A1 v1.0; Rhizophagus irregularis A4 v1.0; Rhizophagus irregularis A5 v1.0; Rhizophagus irregularis B3 v1.0; Rhizophagus irregularis C2 v1.0; Rhizophagus irregularis DAOM 197198 v2.0 [START_REF] Chen | High intraspecific genome diversity in the model arbuscular mycorrhizal symbiont Rhizophagus irregularis[END_REF]; Rhizophagus irregularis DAOM 181602 v1.0 [START_REF] Tisserant | Genome of an arbuscular mycorrhizal fungus provides insight into the oldest plant symbiosis[END_REF]; Rhizopogon vesiculosus Smith; Rhizopogon vinicolor AM-OR11-026 v1.0 [START_REF] Mujic | Comparative Genomics of the Ectomycorrhizal Sister Species Rhizopogon vinicolor and Rhizopogon vesiculosus (Basidiomycota: Boletales) Reveals a Divergence of the Mating Type B Locus[END_REF]; Rhizopus delemar 99-880 from Broad [START_REF] Ma | Genomic analysis of the basal lineage fungus Rhizopus oryzae reveals a wholegenome duplication[END_REF]; Rhizopus microsporus ATCC11559 v1.0; Rhizopus microsporus var. microsporus ATCC52814 v1.0 [START_REF] Lastovetsky | Lipid metabolic changes in an early divergent fungus govern the establishment of a mutualistic symbiosis with endobacteria[END_REF]; Rhizopus microsporus var. chinensis CCTCC M201021 [START_REF] Wang | Draft Genome Sequence of Rhizopus chinensis CCTCCM201021, Used for Brewing Traditional Chinese Alcoholic Beverages[END_REF]; Rhodosporidium toruloides IFO0559_1; Rhodosporidium toruloides IFO0880 v2.0; Rhodosporidium toruloides IFO1236_1 [START_REF] Zhang | Engineering Rhodosporidium toruloides for increased lipid production[END_REF]; Rhodosporidium toruloides IFO0880 v4.0 [START_REF] Coradetti | Functional genomics of lipid metabolism in the oleaginous yeast Rhodosporidium toruloides[END_REF]; Rhodosporidium toruloides NP11 [START_REF] Zhu | A multi-omic map of the lipid-producing yeast Rhodosporidium toruloides[END_REF]; Rhodotorula graminis strain WP1 v1.1 [START_REF] Firrincieli | Genome sequence of the plant growth promoting endophytic yeast Rhodotorula graminis WP1[END_REF]; Rhodotorula sp. JG-1b [START_REF] Goordial | Improved High-Quality Draft Genome Sequence of the Eurypsychrophile Rhodotorula sp. JG1b, Isolated from Permafrost in the Hyperarid Upper-Elevation McMurdo Dry Valleys, Antarctica[END_REF];

Rickenella fibula HBK330-10 v1.0 [START_REF] Korotkin | Stable isotope analyses reveal previously unknown trophic mode diversity in the Hymenochaetales[END_REF]; Rickenella mellea v1.0 (SZMC22713; [START_REF] Krizsán | Transcriptomic atlas of mushroom development reveals conserved genes behind complex multicellularity in fungi[END_REF]; Rigidoporus microporus ED310 v1.0 [START_REF] Oghenekaro | Genome sequencing of Rigidoporus microporus provides insights on genes important for wood decay, latex tolerance and interspecific fungal interactions[END_REF]; Rozella allomycis CSF55 [START_REF] James | Shared signatures of parasitism and phylogenomics unite Cryptomycota and microsporidia[END_REF]; Saccharomyces arboricola H-6 [START_REF] Liti | High quality de novo sequencing and assembly of the Saccharomyces arboricolus genome[END_REF]; Saccharomyces cerevisiae M3707 Dikaryon; Saccharomyces cerevisiae M3836 v1.0; Saccharomyces cerevisiae M3837 v1.0; Saccharomyces cerevisiae M3838 v1.0; Saccharomyces cerevisiae M3839 v1.0; Arthroderma benhamiae CBS 112371; Trichophyton verrucosum HKI 0517 [START_REF] Burmester | Comparative and functional genomics provide insights into the pathogenicity of dermatophytic fungi[END_REF]; Saccharomyces cerevisiae S288C [START_REF] Goffeau | Life with 6000 genes[END_REF]; Saksenaea vasiformis B4078 [START_REF] Chibucos | An integrated genomic and transcriptomic survey of mucormycosis-causing fungi[END_REF]; Scheffersomyces stipitis NRRL Y-11545 v2.0 [START_REF] Jeffries | Genome sequence of the lignocellulose-bioconverting and xylose-fermenting yeast Pichia stipitis[END_REF]; Schizophyllum commune H4-8 v3.0 [START_REF] Ohm | Genome sequence of the model mushroom Schizophyllum commune[END_REF]; Schizopora paradoxa KUC8140 v1.0 [START_REF] Min | Genome sequence of a white rot fungus Schizopora paradoxa KUC8140 for wood decay and mycoremediation[END_REF];

Schizosaccharomyces cryophilus OY26; Schizosaccharomyces japonicus yFS275;
Schizosaccharomyces octosporus yFS286 [START_REF] Rhind | Comparative functional genomics of the fission yeasts[END_REF]; Schizosaccharomyces pombe [292]; Sclerotinia sclerotiorum v1.0 [START_REF] Amselem | Genomic analysis of the necrotrophic fungal pathogens Sclerotinia sclerotiorum and Botrytis cinerea[END_REF]; Serpula himantioides (S.lacrymans var shastensis) MUCL38935 v1.0 [START_REF] Balasundaram | The fungus that came in from the cold: dry rot's pre-adapted ability to invade buildings[END_REF];

Serpula lacrymans S7.3 v2.0; Serpula lacrymans S7.9 v2.0 [START_REF] Eastwood | The plant cell wall-decomposing machinery underlies the functional diversity of forest fungi[END_REF]; Smittium culicis GSMNP; Smittium culicis ID-206-W2; Smittium mucronatum ALG-7-W6; Zancudomyces culisetae COL-18-3 [START_REF] Wang | Genome-Wide Survey of Gut Fungi (Harpellales) Reveals the First Horizontally Transferred Ubiquitin Gene from a Mosquito Host[END_REF];

Sodiomyces alkalinus v1.0 [START_REF] Grum-Grzhimaylo | The obligate alkalophilic soda-lake fungus Sodiomyces alkalinus has shifted to a protein diet[END_REF]; Sphaerosporella brunnea Sb_GMNB300 v2.0 [START_REF] Benucci | Draft Genome Sequence of the Ectomycorrhizal Ascomycete Sphaerosporella brunnea[END_REF]; Spizellomyces punctatus DAOM BR117 [START_REF] Russ | Genome Sequence of Spizellomyces punctatus[END_REF]; Sporisorium reilianum SRZ2 [START_REF] Schirawski | Pathogenicity determinants in smut fungi revealed by genome comparison[END_REF]; Stagonospora nodorum SN15 v2.0 [START_REF] Hane | Dothideomycete plant interactions illuminated by genome sequencing and EST analysis of the wheat pathogen Stagonospora nodorum[END_REF]; Stemphylium lycopersici CIDEFI-216 [START_REF] Franco | Draft Genome Sequence and Gene Annotation of Stemphylium lycopersici Strain CIDEFI-216[END_REF]; Suillus brevipes Sb2 v2.0 [START_REF] Branco | Genetic isolation between two recently diverged populations of a symbiotic fungus[END_REF]; Talaromyces borbonicus CBS 141340 [START_REF] Varriale | Talaromyces borbonicus, sp. nov., a novel fungus from biodegraded Arundo donax with potential abilities in lignocellulose conversion[END_REF]; Talaromyces marneffei ATCC 18224 [START_REF] Nierman | Genome Sequence of the AIDS-Associated Pathogen Penicillium marneffei (ATCC18224) and Its Near Taxonomic Relative Talaromyces stipitatus (ATCC10500)[END_REF]; Taphrina deformans [START_REF] Cissé | Genome sequencing of the plant pathogen Taphrina deformans, the causal agent of peach leaf curl[END_REF];

Thermomyces lanuginosus SSBP [START_REF] Mchunu | Xylanase Superproducer: Genome Sequence of a Compost-Loving Thermophilic Fungus, Thermomyces lanuginosus Strain SSBP[END_REF]; Tolypocladium inflatum NRRL 8044 [START_REF] Bushley | The genome of tolypocladium inflatum: evolution, organization, and expression of the cyclosporin biosynthetic gene cluster[END_REF]; Trametes pubescens FBCC735 [START_REF] Granchi | Genome Sequence of the Basidiomycete White-Rot Fungus Trametes pubescens FBCC735[END_REF]; [START_REF] Druzhinina | Massive lateral transfer of genes encoding plant cell wall-degrading enzymes to the mycoparasitic fungus Trichoderma from its plant-associated hosts[END_REF]; Trichoderma asperellum TR356 v1.0; Trichoderma harzianum TR274 v1.0 [START_REF] Kubicek | Evolution and comparative genomics of the most common Trichoderma species[END_REF]; Trichoderma atrobrunneum ITEM 908 [START_REF] Fanelli | Genomic characterization of Trichoderma atrobrunneum (T. harzianum species complex) ITEM 908: insight into the genetic endowment of a multi-target biocontrol strain[END_REF]; Trichoderma atroviride v2.0; Trichoderma virens Gv29-8 v2.0 [START_REF] Kubicek | Comparative genome sequence analysis underscores mycoparasitism as the ancestral life style of Trichoderma[END_REF]; Trichoderma hamatum GD12 [START_REF] Studholme | Investigating the beneficial traits of Trichoderma hamatum GD12 for sustainable agriculture-insights from genomics[END_REF]; Trichoderma parareesei CBS 125925 [START_REF] Yang | Genome Sequence and Annotation of Trichoderma parareesei, the Ancestor of the Cellulase Producer Trichoderma reesei[END_REF]; Trichoderma pleuroti TPhu1 [START_REF] Marik | Diversity Profile and Dynamics of Peptaibols Produced by Green Mould Trichoderma Species in Interactions with Their Hosts Agaricus bisporus and Pleurotus ostreatus[END_REF]; Trichoderma reesei QM6a [START_REF] Li | Trichoderma reesei complete genome sequence, repeat-induced point mutation, and partitioning of CAZyme gene clusters[END_REF];

Trichoderma
Trichoderma reesei RUT C-30 v1.0 [START_REF] Jourdier | Proximity ligation scaffolding and comparison of two Trichoderma reesei strains genomes[END_REF]; Trichoderma reesei v2.0 [START_REF] Martinez | Genome sequencing and analysis of the biomass-degrading fungus Trichoderma reesei (syn. Hypocrea jecorina)[END_REF]; Trichosporon asahii var.

asahii CBS 2479 [START_REF] Yang | Disseminated trichosporonosis in China[END_REF]; Trichosporon asahii var. asahii CBS 8904 [START_REF] Yang | Genome sequence of the Trichosporon asahii environmental strain CBS 8904[END_REF]; Trichosporon oleaginosus IBC0246 v1.0 [START_REF] Kourist | Genomics and Transcriptomics Analyses of the Oil-Accumulating Basidiomycete Yeast Trichosporon oleaginosus: Insights into Substrate Utilization and Alternative Evolutionary Trajectories of Fungal Mating Systems[END_REF]; Tuber melanosporum Mel28 v1.2 [START_REF] Martin | Périgord black truffle genome uncovers evolutionary origins and mechanisms of symbiosis[END_REF]; Ustilaginoidea virens [START_REF] Kumagai | Genome Sequence of Ustilaginoidea virens IPU010, a Rice Pathogenic Fungus Causing False Smut[END_REF]; Ustilago hordei Uh4857_4 [START_REF] Laurie | Genome comparison of barley and maize smut fungi reveals targeted loss of RNA silencing components and species-specific presence of transposable elements[END_REF]; Ustilago maydis 521 v2.0 [START_REF] Kämper | Insights from the genome of the biotrophic fungal plant pathogen Ustilago maydis[END_REF]; Venturia inaequalis; Venturia pirina [START_REF] Deng | Comparative analysis of the predicted secretomes of Rosaceae scab pathogens Venturia inaequalis and V. pirina reveals expanded effector families and putative determinants of host range[END_REF];

Verticillium alfalfae VaMs.102; Verticillium dahliae VdLs.17 [START_REF] Klosterman | Comparative genomics yields insights into niche adaptation of plant vascular wilt pathogens[END_REF]; Volvariella volvacea V23 [START_REF] Bao | Sequencing and comparative analysis of the straw mushroom (Volvariella volvacea) genome[END_REF];

Wallemia ichthyophaga EXF-994 [START_REF] Zajc | Genome and transcriptome sequencing of the halophilic fungus Wallemia ichthyophaga: haloadaptations present and absent[END_REF]; Wallemia mellicola v1.0 [START_REF] Padamsee | The genome of the xerotolerant mold Wallemia sebi reveals adaptations to osmotic stress and suggests cryptic sexual reproduction[END_REF]; Xylona heveae TC161 v1.0 [START_REF] Gazis | The genome of Xylona heveae provides a window into fungal endophytism[END_REF]; Yarrowia lipolytica CLIB89(W29) [START_REF] Magnan | Sequence Assembly of Yarrowia lipolytica Strain W29/CLIB89 Shows Transposable Element Diversity[END_REF]; Yarrowia lipolytica FKP355 v1.0 [START_REF] Pomraning | Regulation of Yeast-to-Hyphae Transition in Yarrowia lipolytica[END_REF]; Yarrowia lipolytica PO1f v1.0; Yarrowia lipolytica YlCW001 v1.0 [START_REF] Walker | Draft Genome Assemblies of Ionic Liquid-Resistant Yarrowia lipolytica PO1f and Its Superior Evolved Strain, YlCW001[END_REF]; Yarrowia lipolytica YB392 v1.0;

Yarrowia lipolytica YB419 v1.0; Yarrowia lipolytica YB420 v1.0; Yarrowia lipolytica YB566 v1.0;

Yarrowia lipolytica YB567 v1.0 [START_REF] Walker | Draft Genome Assemblies of Five Robust Yarrowia lipolytica Strains Exhibiting High Lipid Production, Pentose Sugar Utilization, and Sugar Alcohol Secretion from Undetoxified Lignocellulosic Biomass Hydrolysates[END_REF]; Zygosaccharomyces rouxii CBS732 [START_REF] Génolevures Consortium | Comparative genomics of protoploid Saccharomycetaceae[END_REF]; Zymoseptoria ardabiliae STIR04_1.1.1; Zymoseptoria pseudotritici STIR04_2.2.1 [START_REF] Stukenbrock | Fusion of two divergent fungal individuals led to the recent emergence of a unique widespread pathogen species[END_REF]; Zymoseptoria brevis Zb18110 [START_REF] Grandaubert | RNA-seq-Based Gene Annotation and Comparative Genomics of Four Fungal Grass Pathogens in the Genus Zymoseptoria Identify Novel Orphan Genes and Species-Specific Invasions of Transposable Elements[END_REF]; Melampsora allii-populina 12AY07 v1.0; Melampsora americana R15-033-03 v1.0 (unpublished).

Protein sequence analysis and search for potential subcellular targeting.

The protein sequences were aligned using Clustal Omega [START_REF] Sievers | Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega[END_REF] (https://www.ebi.ac.uk/Tools/msa/clustalo/) and manually checked. The predictions for subcellular localizations were done based on the assessment obtained from the TargetP [START_REF] Armenteros | Detecting sequence signals in targeting peptides using deep learning[END_REF] (http://www.cbs.dtu.dk/services/TargetP/), SignalP [START_REF] Almagro Armenteros | SignalP 5.0 improves signal peptide predictions using deep neural networks[END_REF] (http://www.cbs.dtu.dk/services/SignalP/abstract.php), WolfPsort [START_REF] Horton | WoLF PSORT: protein localization predictor[END_REF] (https://wolfpsort.hgc.jp/) and ESLpred [START_REF] Bhasin | ESLpred: SVM-based method for subcellular localization of eukaryotic proteins using dipeptide composition and PSI-BLAST[END_REF] (http://crdd.osdd.net/raghava/eslpred/) software suites.

Phylogenomic analyses and search for horizontal gene transfers.

For each type of Msr, we built a multiple sequence alignment with all the identified sequences using MAFFT version v7.429 [START_REF] Yamada | Application of the MAFFT sequence alignment program to large data-reexamination of the usefulness of chained guide trees[END_REF]. Each alignment was trimmed to remove poorly aligned regions using trimAl 1.2 [START_REF] Capella-Gutiérrez | trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses[END_REF] and manually inspected for the conservation of the catalytic residues. A phylogenetic tree was constructed for each type of Msr using RAxML Master Pthread [START_REF] Stamatakis | RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies[END_REF] version 8.2.12 (PROTGAMMAWAG model and 500 bootstraps). The phylogenetic tree was represented using iTOL (https://itol.embl.de/) [START_REF] Letunic | Interactive Tree Of Life (iTOL) v4: recent updates and new developments[END_REF]. To investigate potential horizontal gene transfers, the non-canonical Msr sequences along with the sequences whose phylogeny did not fit the species phylogeny were retrieved. Those sequences were used as BLAST queries against the non-redundant protein sequence (nr) database of NCBI (https://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastp&PAGE_TYPE=BlastSearch&LINK_LO C=blasthome) and the best 100 hits were retrieved. The fungal sequences for which protein accessions outside the fungal kingdom were identified among the highest identity scores were in further used as BLAST queries against nr excluding the taxon Fungi (taxid: 4751) and the 25 hits with the highest identity scores were retrieved. In parallel, the same fungal genes were used as queries against the fungal nr database and the 50 hits with the highest identity scores were retrieved, excluding those with 100% identity with the query. The retrieved fungal hits were manually inspected to exclude incomplete sequences. Non-canonical sequences (lacking canonical active site or of abnormal length) were also removed to avoid potential inconsistencies and long branches in the phylogenetic trees. Finally, for each type of Msr, a phylogenetic tree was constructed that included the candidate fungal gene for horizontal gene transfer, plus 25 non-fungal and 25 fungal sequences with the highest identity scores. For each candidate to HGT, the numbers of exons, the GC content of the coding sequence and the GC content at the third positions of the codons were calculated using the gene structures and coding sequences downloaded from the MycoCosm database. The GC content on the third positions of the codons were calculated only for the coding sequences starting with ATG. All calculations were made using Microsoft® Excel® version 2101.

Results

MsrA, MsrB and fRMsr are largely conserved across the fungal kingdom.

Currently, around 136,000 species of fungi are known and classified into 9 phyla (Figure 1) [START_REF] Naranjo-Ortiz | Fungal evolution: diversity, taxonomy and phylogeny of the Fungi[END_REF][START_REF] James | Toward a Fully Resolved Fungal Tree of Life[END_REF]. Together, the Ascomycota and the Basidiomycota form the subkingdom Dikarya, which regroups more than 97 % of all described fungal species (∼84,000 species and ∼48,000 species, respectively) [START_REF] James | Toward a Fully Resolved Fungal Tree of Life[END_REF]. Each phylum contains three monophyletic subphyla (Pucciniomycotina, Ustilaginomycotina and Agaricomycotina for the Basidiomycota, and Taphrinomycotina, Saccharomycotina and Pezizomycotina for the Ascomycota). The other seven phyla are described as 'early-diverging fungi' [START_REF] James | Toward a Fully Resolved Fungal Tree of Life[END_REF] (Fig. 1, Data S1). Using the S. cerevisiae MsrA, MsrB and fRMsr protein sequences as queries, we used the BLASTP and TBLASTN software suites to search for msr genes in 683 available genomes in the MycoCosm database [START_REF] Grigoriev | MycoCosm portal: gearing up for 1000 fungal genomes[END_REF]. The selected genomes were from 595 species that spanned the kingdom Fungi (Fig. 1, Data S1), including 65% Ascomycota species, 25%

Basidiomycota species and 10% early-diverging fungi. We found that the very great majority of these genomes contained one gene coding for a MsrA and one coding for a MsrB (Fig. 1, Data S1), indicating that most fungi have a simple Msr system dedicated to protein oxidation repair, similarly to most other known organisms [START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF][START_REF] Zhang | Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases[END_REF]. Most interestingly, we found that a fRmsr gene was present in almost all the analyzed genomes (Fig. 1, Data S1). The distribution of fRmsr across the fungal kingdom clearly showed that the presence of fRMsr is not limited to bacteria and unicellular eukaryotes as previously described [START_REF] Gruez | Structural and biochemical characterization of free methionine-R-sulfoxide reductase from Neisseria meningitidis[END_REF][START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF][START_REF] Lin | Free methionine-(R)sulfoxide reductase from Escherichia coli reveals a new GAF domain function[END_REF]. Of note, the search for homologs of the bacterial molybdoenzymes able to reduce MetO gave no significant hits (data not shown).

Figure 1. Numbers of msrA, msrB and fRmsr genes identified in fungal genomes. The simplified phylogenetic tree was built according to [START_REF] Naranjo-Ortiz | Fungal evolution: diversity, taxonomy and phylogeny of the Fungi[END_REF][START_REF] James | Toward a Fully Resolved Fungal Tree of Life[END_REF]. The precise number of msr genes per genome is available in Data S1.

We observed some variations in the numbers of msr genes in several genomes. We found 72 457 genomes, corresponding to 57 species, that had more than one copy of at least one msr gene (Table 1, 458 Data S1-4). We found 42 genomes having two copies of msrA and 17 having three or four copies (Table 459 1, Data S1-2). The highest number of msrA copies was 4 in the genome of the Agaricomycetes 460 Crucibulum laeve CBS 166.37 and Dendrothele bispora CBS 962.96 (Table 1). In the case of msrB, 461 only 14 genomes had two gene copies (Data S1, S3), and the Kickxellomycotina Smittium culicis 462 GSMNP was the only genome with three copies (Table 1). For fRmsr, only six genomes had two copies 463 (Data S1, S4), and the Agaricomycetes Ramaria rubella (R. acris) UT-36052-T and Coprinellus 464 micaceus FP101781 had 4 and 3 gene copies, respectively (Table 1). Overall, for the three types of Msr, 465 the fungal species having more than one copy were sporadically spread across the fungal kingdom 466 Also, we found only 74 genomes (58 species), in which one or more Msr types were absent (Table 2). Among them, the nine Microsporidia species, two Neocallimastigomycetes species (out of five species analyzed) and one Taphrinomycotina species, Pneumocystis jirovecii (out of eight species analyzed) lacked all three Msr types. These species are either obligate intracellular parasites (Microsporidia and Pneumocystis jirovecii) or live in the anaerobic conditions of the animal gut (Piromyces finnis and Piromyces sp. E2) [START_REF] Naranjo-Ortiz | Fungal evolution: diversity, taxonomy and phylogeny of the Fungi[END_REF][START_REF] Skalski | Pathobiology of Pneumocystis pneumonia: life cycle, cell wall and cell signal transduction[END_REF]. Interestingly, the three other Neocallimastigomycetes species (Anaeromyces robustus, Neocallimastix californiae G1 and Orpinomyces sp.), also living in anaerobic conditions [START_REF] James | Toward a Fully Resolved Fungal Tree of Life[END_REF], do not possess MsrA nor MsrB but have a gene coding for a fRMsr (Table 2). Our results show that, additionally to Encephalitozoon cuniculi, previously identified as the unique eukaryote lacking both MsrA and MsrB [START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF], 14 other fungal species are devoid of any protein-bound

(
MetO reduction enzyme (Table 2). Furthermore, we found six species lacking msrA but having a msrB and a fRmsr gene (Table 2). These are the agaricomycete Moniliophthora perniciosa, the eurotiomycetes Cladophialophora immunda, Penicillium coprophilum and Penicillium flavigenum, the sordiariomycete Magnaporthiopsis poae and the saccharomycotina Saturnispora dispora. To our knowledge, they constitute the first species, over all kingdoms, described to have only a MsrB to reduce and repair oxidized proteins, as none was found so far in genome surveys [START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF][START_REF] Zhang | Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases[END_REF]. Of note, we did not find fungal species having only a MsrB, as these six species also possessed a fRMsr (Table 2). We found that the five species Trichosporon asahii (Tremellomycetes), Clohesyomyces aquaticus (Dothideomycetes), Cladonia grayi (Lecaronomycetes) and Piptocephalis cylindrospora (Zoopagomycotina) possessed a MsrA and a fRMsr but lacked a MsrB and that the Zoopagomycotina Syncephalis pseudoplumigaleata had a MsrA only (Table 2). Finally, we found only 27 species having both a MsrA and a MsrB but lacking a fRMsr (Table 2). These species were sporadically dispersed among the fungal kingdom, but three groups stood out as remarkable: the Glomeromycotina, for which none of the four species analyzed had a fRMsr, the Taphrinomycotina for which six species out of eight analyzed (including the four Schizosaccharomyces species analyzed) were devoid of fRMsr, and the Pucciniomycotina for which all the Melampsora and Puccinia species analyzed here lacked a fRMsr (Table 2).

Altogether, these results showed that the great majority of fungi possess one gene coding for each protein-repairing Msr type (i.e. MsrA and MsrB), as well as one gene coding for the free MetO reductase fRMsr. Y, presence of one or more genes coding for the Msr; N, absence of gene coding for the Msr.

Fungal Msrs globally retained canonical sequence features for activity.

Sequence analysis and enzymatic characterizations of MsrAs from several organisms showed that the catalytic residue is located within a G[C/U]FW motif, located in the N-terminal part of the protein [START_REF] Tarrago | Methionine sulfoxide reductases preferentially reduce unfolded oxidized proteins and protect cells from oxidative protein unfolding[END_REF][START_REF] Lowther | The mirrored methionine sulfoxide reductases of Neisseria gonorrhoeae pilB[END_REF][START_REF] Boschi-Muller | A sulfenic acid enzyme intermediate is involved in the catalytic mechanism of peptide methionine sulfoxide reductase from Escherichia coli[END_REF][START_REF] Antoine | Characterization of the Amino Acids from Neisseria meningitidis MsrA Involved in the Chemical Catalysis of the Methionine Sulfoxide Reduction Step[END_REF][START_REF] Kim | Catalytic advantages provided by selenocysteine in methionine-S-sulfoxide reductases[END_REF]. The great majority of known MsrAs possess a catalytic Cys, whereas a few Sec-containing proteins are found in some insects, marine organisms and unicellular algae [START_REF] Tarrago | Protein-repairing methionine sulfoxide reductases in photosynthetic organisms: gene organization, reduction mechanisms, and physiological roles[END_REF][START_REF] Kim | Catalytic advantages provided by selenocysteine in methionine-S-sulfoxide reductases[END_REF]. In canonical MsrAs, the resolving Cys, involved in the regeneration of the catalytic Cys, is located in the C-terminal part of the protein, althought the number and positions of resolving Cys vary [START_REF] Tarrago | Methionine sulfoxide reductases preferentially reduce unfolded oxidized proteins and protect cells from oxidative protein unfolding[END_REF][START_REF] Boschi-Muller | A sulfenic acid enzyme intermediate is involved in the catalytic mechanism of peptide methionine sulfoxide reductase from Escherichia coli[END_REF][START_REF] Lowther | Thiol-disulfide exchange is involved in the catalytic mechanism of peptide methionine sulfoxide reductase[END_REF][START_REF] Rouhier | Functional and structural aspects of poplar cytosolic and plastidial type a methionine sulfoxide reductases[END_REF].

To determine whether the fungal MsrAs share these properties, we analyzed the 709 full length MsrAs identified. These MsrAs ranged from 142 to 322 amino acids, and all of them were made of a single MsrA domain (Data S2A). The length differences were mostly due to the presence of N-terminal extensions of variable sizes, indicating the possible presence of signal peptides for protein distribution in subcellular compartments. We used several targeting prediction programs (see section 2.

3) to evaluate the potential subcellular localization of the fungal MsrAs (Data S2A). Most MsrAs were predicted to be localized into the cytoplasm (45 %) or in the mitochondria (44 %) (Fig. 2A, Data S2A). The other sequences were predicted to be secreted (6%), to be localized in other compartments, or had no clearly assigned localization (6%) (Fig. 2A, Data S2A). The alignment of the primary sequences revealed that the great majority (∼ 90 %) shared common features with the previously characterized S. cerevisiae

MsrA [START_REF] Tarrago | Methionine sulfoxide reductases preferentially reduce unfolded oxidized proteins and protect cells from oxidative protein unfolding[END_REF][START_REF] Ma | Structural Plasticity of the Thioredoxin Recognition Site of Yeast Methionine S-Sulfoxide Reductase Mxr1[END_REF] (Fig. 3A, Data S2B). The catalytic Cys (position 25 in the S. cerevisiae MsrA) was located in the conserved motif 24 GCFW 27 . The Tyr 64 , Glu 76 , Asp 111 and Tyr 116 residues involved in substrate stabilization and catalysis [START_REF] Antoine | Characterization of the Amino Acids from Neisseria meningitidis MsrA Involved in the Chemical Catalysis of the Methionine Sulfoxide Reduction Step[END_REF], and the residues Gly 47 , His 100 , Gln 108 , Gly 113 , His 163 and Tyr 166 were also conserved. Finally, the Cys 176 , previously identified as resolving Cys [START_REF] Tarrago | Methionine sulfoxide reductases preferentially reduce unfolded oxidized proteins and protect cells from oxidative protein unfolding[END_REF][START_REF] Ma | Structural Plasticity of the Thioredoxin Recognition Site of Yeast Methionine S-Sulfoxide Reductase Mxr1[END_REF], was included in the 173 GYXC 176 motif (Fig. 3A, Data S2B). Because of their predominance in all the fungal kingdom, we defined the fungal MsrAs having these properties as 'canonical' sequences. The fungal MsrAs that did not match these sequence features were defined as 'non-canonical' MsrAs (Data S2A, C, D).

Particularly, we observed that in ∼ 5 % of the identified sequences, the Phe 26 residue in the 24 GCFW 27 motif containing the catalytic Cys, was substituted by a Tyr. We also observed the replacement of Asp 111 by an Asn residue in few sequences (∼ 2 %) (Data S2C). Moreover, some variations were also observed for the resolving Cys (Data S2C). Fourteen sequences (∼ 2 %) lacked the conserved 173 In the case of MsrBs, previous biochemical characterizations demonstrated that the catalytic Cys is located in a RXCXN motif in the C-terminal part of the protein [START_REF] Lowther | The mirrored methionine sulfoxide reductases of Neisseria gonorrhoeae pilB[END_REF][START_REF] Tarrago | Regeneration mechanisms of Arabidopsis thaliana methionine sulfoxide reductases B by glutaredoxins and thioredoxins[END_REF][START_REF] Kim | Different catalytic mechanisms in mammalian selenocysteine-and cysteine-containing methionine-R-sulfoxide reductases[END_REF][START_REF] Neiers | Characterization of the amino acids from Neisseria meningitidis methionine sulfoxide reductase B involved in the chemical catalysis and substrate specificity of the reductase step[END_REF][START_REF] Olry | Characterization of the methionine sulfoxide reductase activities of PILB, a probable virulence factor from Neisseria meningitidis[END_REF][START_REF] Ranaivoson | Methionine sulfoxide reductase B displays a high level of flexibility[END_REF]. Mammals express a Sec-containing form in which the Asp is replaced by a Phe [START_REF] Kim | Different catalytic mechanisms in mammalian selenocysteine-and cysteine-containing methionine-R-sulfoxide reductases[END_REF]. The resolving Cys is generally located in a CGWP motif present in the N-terminal part of the protein [START_REF] Tarrago | Regeneration mechanisms of Arabidopsis thaliana methionine sulfoxide reductases B by glutaredoxins and thioredoxins[END_REF][START_REF] Olry | Characterization of the methionine sulfoxide reductase activities of PILB, a probable virulence factor from Neisseria meningitidis[END_REF]. In the case of mammalian MsrBs, one or two resolving Cys, located in the N-terminal extremity of the protein, can be involved in the regeneration process [START_REF] Kim | Different catalytic mechanisms in mammalian selenocysteine-and cysteine-containing methionine-R-sulfoxide reductases[END_REF][START_REF] Cao | Methionine sulfoxide reductase B3 requires resolving cysteine residues for full activity and can act as a stereospecific methionine oxidase[END_REF]. Of note, most MsrBs possess two CX2C clusters coordinating a structural Zn atom [START_REF] Shumilina | Evolution of Structural and Coordination Features Within the Methionine Sulfoxide Reductase B Family[END_REF]. Here, we analyzed the 651 complete fungal MsrB protein sequences. The fungal MsrBs consisted of a single domain and ranged in length from 95 to 289 amino acids (Data S3A). Most of the variations in size were due to the presence of an N-terminal extension potentially involved in subcellular targeting. The majority of fungal MsrBs were predicted to be addressed to the cytosol (61 %) (Fig. 2B, Data S3A). The other proteins were either predicted to be localized in the mitochondria (26 %), secreted (1 %), targeted to another compartment, or were not clearly predicted to be addressed to a subcellular compartment (28 %) (Fig. 2B,Data S3A). Almost all fungal MsrBs (> 99 %) possessed the features of the S. cerevisiae enzyme (Fig. 3B, Data S3B): i) the two CX2C motifs involved in the coordination of a Zn atom, ii) the resolving Cys 97 (according to S.

cerevisiae MsrB residue numbering) included in a 97 CGW 99 motif, iii) the conserved His 134 and His 136 implicated in substrate binding, together with Arg 155 and Asn 159 [START_REF] Lowther | The mirrored methionine sulfoxide reductases of Neisseria gonorrhoeae pilB[END_REF][START_REF] Neiers | Characterization of the amino acids from Neisseria meningitidis methionine sulfoxide reductase B involved in the chemical catalysis and substrate specificity of the reductase step[END_REF], and iv) the catalytic Cys 157 located in the 155 RXCXN 159 motif. The Gly 98 , Gly 135 , Val 137 , Phe 138 and Glu 141 residues were also conserved in all these canonical fungal MsrBs (Fig. 3B, Data S3B). Only four sequences (< 1 %), from the orbiliomycetes Arthrobotrys oligospora and Monacrosporium haptotylum, the taphrinomycotina Protomyces lactucaedebilis and the chytridiomycete Blyttiomyces helices, presented remarkable differences in primary sequence features (Data S3C). These MsrBs lacked the resolving Cys at position 97, which was substituted by a Ser or a Thr, like plant and human orthologs that use an unusual regeneration process [START_REF] Tarrago | Plant thioredoxin CDSP32 regenerates 1-cys methionine sulfoxide reductase B activity through the direct reduction of sulfenic acid[END_REF][START_REF] Tarrago | Regeneration mechanisms of Arabidopsis thaliana methionine sulfoxide reductases B by glutaredoxins and thioredoxins[END_REF][START_REF] Cao | Methionine sulfoxide reductase B3 requires resolving cysteine residues for full activity and can act as a stereospecific methionine oxidase[END_REF]. Other important residues involved in substrate binding and catalysis [START_REF] Gruez | Structural and biochemical characterization of free methionine-R-sulfoxide reductase from Neisseria meningitidis[END_REF] were also conserved or substituted by residues with similar properties in positions 82, 85, 94, 123, 129 and 132. The Pro 84 , Gly 87 , His 122 and Val 134 were also strictly conserved in these canonical fungal fRMsrs (Fig. 3C, Data S4B). Only three fRMsr sequences from the Agaricomycetes Scleroderma citrinum, Dendrothele bispora and Pisolithus tinctorius presented non-canonical characteristics. The first two lacked the potential resolving Cys 101 and may be still able to reduce the free MetO, but in the latest, the catalytic Cys 125 was substituted by an Arg, likely precluding catalytic activity (Data S4C).

Altogether, these results uncover few proteins with non-canonical sequence features, but principally showed that for each Msr type, the residues involved in catalysis are globally conserved throughout the fungal kingdom.

The phylogenetic analysis of fungal Msrs revealed horizontal gene transfers from bacteria.

The phylogenetic relationship of fungal MsrAs globally matched the expected clustering for early-diverging fungi, Ascomycota and Basidiomycota sequences (Fig. S1-3). However, a few Ascomycota MsrA sequences clustered with Basidiomycota sequences (indicated by an asterisk on Fig. S1). We also noticed the clustering of all the MsrB sequences from Pucciniomycotina (Basidiomycetes) species with Ascomycota sequences (Fig. S2). Strikingly, the MsrB sequences from early-diverging fungi did not group in a single cluster but were interspersed in clusters containing Basidiomycota or Ascomycota sequences (Fig. S2). For fRMsrs, we observed three distinct clusters containing the protein sequences from Basidiomycota, Ascomycota and early-diverging fungi, respectively (Fig. S3).

However, two sequences from early-diverging fungi were included in the cluster containing the Ascomycota sequences (Fig. S3). Altogether, these results showed that the phylogeny of Msrs was globally congruent with the phylogeny of the species, except for a few protein sequences.

Several discrepancies suggested that some Msrs could have arisen from horizontal gene transfer.

These discrepancies were: i) the positioning in phylogenetic clusters not reflecting the phylogeny of the species from which they were isolated (Fig. S1-3), ii) the presence of non-canonical sequence features (Data S2-4), and iii) the presence of fRmsr genes in the genome of three strictly anaerobic neocallimastigomycetes, whereas other organisms from the same phylum had no msr genes (Table 2).

To evaluate the possibility of horizontal gene transfers, we selected all Msr sequences (67 MsrAs, 48

MsrBs and 8 fRMsrs) with one or more of these discrepancies and searched for their closest putative homologs by BLASTP search in all organisms recorded in the NCBI nr database. We discarded all the sequences for which the closest homologs were found in other fungal taxa. Indeed, because of the strong conservation of the protein sequences, it would be difficult to ascertain the occurrence of fungus-tofungus horizontal gene transfers. We retrieved 17 MsrA, three MsrB and three fRMsr protein sequences for further analysis (Fig. 456). For each type of Msr, we used the selected protein sequences, together with the respective 25 sequences with the highest identity score from fungi on the one hand and from non-fungal organisms on the other hand for phylogenetic analyses (Fig. 456). The phylogenetic trees highlighted two MsrA, three MsrB and three fRMsr sequences, among the selected candidates, clustering with bacterial and amoeba homologs (Fig. 456, Table 3). Moreover, we identified two additional fungal fRMsrs, not included in our primary data set, which clustered with bacterial proteins (Fig. 6, Table 3). Except for two MsrBs from Orbiliomycetes (Ascomycota), all these Msr sequences were from early-diverging fungi (Table 3). Interestingly, in the cases of MsrA and MsrB, they were all from organisms having another gene coding for a canonical enzyme in their genomes (Data S1-3).

For MsrAs, a potential horizontal gene transfer was observed for the two selenocysteinecontaining enzymes from Gonapodya prolifera (Table 3). Very interestingly, their closest homolog was also a selenocysteine-containing MsrA, from the bacteria Alkaliphilus oremlandii. Bacteria from the genus Alkaliphilus are found in sediments and ponds [START_REF] Fisher | Transformation of Inorganic and Organic Arsenic byAlkaliphilus oremlandiisp. nov. Strain OhILAs[END_REF], and Gonapodya prolifera occurs on fruits submerged into ponds [START_REF] Fuller | Observations of Gonapodya in Pure Culture: Growth, Development and Cell Wall Characterization[END_REF]. The presence of both organisms in a same ecological niche could have favored the horizontal gene transfer. The similarity between the two Gonapodya prolifera Sec-MsrA suggested they arose from one horizontal gene transfer event followed by a gene duplication (Fig. 4, Fig. S4). In the case of MsrBs, the three selected sequences clustered with bacterial MsrBs (Fig. 5). The two MsrBs from the Orbiliomycetes species Arthrobotrys oligospora and Monacrosporium haptotylum had MsrBs from Sphingomonas bacteria as closest homologs. Both fungi trap nematodes in soil [START_REF] Meerupati | Genomic mechanisms accounting for the adaptation to parasitism in nematode-trapping fungi[END_REF],

and sphingomonads have been isolated from many different land and water habitats [START_REF] Nishiyama | Identification of soil micro-habitats for growth, death and survival of a bacterium, γ-1,2,3,4,5,6-hexachlorocyclohexane-assimilating Sphingomonas paucimobilis, by fractionation of soil[END_REF], indicating that the co-occurrence in the same habitat of a sphingomonad donor and a common ancestor of A.

oligospora and M. haptotylum is plausible, which could have allowed horizontal gene transfers. Another potential horizontal gene transfer for MsrB was identified, from a donor cyanobacterium related to the marine Calothrix parasitica to the plant pathogen fission yeast Protomyces lactucaedebilis (Taphrinomycotina) (Table 3). In this case, the ecology of the extant candidate donor and acceptor do not support the co-occurrence of both organisms in a same ecological niche. The three fRMsrs we selected as candidates to horizontal gene transfer were from the neocallimastigomycetes species Anaeromyces robustus, Neocallimastix californiae and Orpinomyces sp. (Table 3). In our phylogenetic analysis, they clustered with sequences of bacteria from the phylum Firmicutes which, similarly to neocallimastigomycetes, live in the gut of ruminants (Fig. 6).

Interestingly, two other fungal fRMsr sequences, one from Piromyces sp. and one from Mortierella sp.

GBA39, were also present in the same cluster. These two sequences were omitted from our genomic search because of the absence of the corresponding genomes in the MycoCosm database. Of note, the horizontal transfer of fRmsr genes from Firmicutes to neocallimastigomycetes has been shown recently for Anaeromyces robustus and Piromyces sp. [START_REF] Murphy | Horizontal Gene Transfer as an Indispensable Driver for Evolution of Neocallimastigomycota into a Distinct Gut-Dwelling Fungal Lineage[END_REF]. Altogether, these results strongly argue for the fact that the presence of fRmsr in the genomes of these anaerobic fungi arose from horizontal gene transfers from Firmicutes (Table 3).

In addition to the phylogenetic method, we performed a parametric analysis of the numbers of exons, the overall GC contents of their coding sequence (CDS), and the GC contents of the third position of each codon to identify potential bias as supporting arguments of horizontal gene transfer (Table 3).

The comparison of these parameters with those of the other genes included in the same genomic scaffold were shown as potential indicators of horizontal gene transfers [START_REF] Ravenhall | Inferring Horizontal Gene Transfer[END_REF]. Excepted for the two Gonapodya prolifera msrA genes, which had no distinguishable values from the other genes included in their scaffolds, all other genes coding MsrBs or fRMsrs, had extreme values for at least two of the three parameters considered (Table 3). First of all, all these eight genes were made of a single exon, like bacterial genes. The global GC content of Arthrobotrys oligospora and Monacrosporium haptotylum msrB genes coding sequences were the highest of all those included in their scaffold. Similarly, the global GC content of Protomyces lactucaedebilis msrB, Anaeromyces robustus fRmsr and Orpinomyces sp. fRmsr were among the highest (Table 3). In the cases of fRmsrs from Neocallimastix californiae, Piromyces sp. and Mortierella sp., the GC contents of the third base of the codons were among the lowest by comparison with the other genes present in the scaffold. Noticeably, we also observed a lower GC content at the third position of the codons in the fRmsr gene from Piromyces sp., for which the acquisition via HGT was previously shown [START_REF] Letunic | Interactive Tree Of Life (iTOL) v4: recent updates and new developments[END_REF].

Altogether, these analyses showed that the phylogeny of fungal Msrs globally matched the phylogeny of the fungi, in accordance with vertical inheritance of the genes from bacterial ancestors [START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF][START_REF] Lin | Free methionine-(R)sulfoxide reductase from Escherichia coli reveals a new GAF domain function[END_REF][START_REF] Delaye | Molecular evolution of peptide methionine sulfoxide reductases (MsrA and MsrB): on the early development of a mechanism that protects against oxidative damage[END_REF][START_REF] Zhang | Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases[END_REF]. However, a few horizontal gene transfer events occurred in each Msr family, that contributed to the distribution of Msr genes in extant species.

* *

Discussion

This global genomic search for msr genes in nearly 700 fungal genomes covering the fungal kingdom showed that most fungi contain one gene coding for each thiol-oxidoreductase type, i.e., MsrA, MsrB and fRMsr. The phylogenetic analyses and inspection of protein sequence features revealed that Msrs from each type are globally strongly conserved across the fungal kingdom. This is consistent with the prokaryotic origin of these genes. Yet, the identification of fRmsr genes in almost all genomes across the fungal kingdom was surprising. Indeed, no fRmsr genes were identified so far from multicellular eukaryotes [START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF]. Very likely, the most obvious reason was the smaller number of eukaryotic genomes considered (i.e. 160), and the smaller number of fungal genomes available at the time of the previous study [START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF][START_REF] Grigoriev | MycoCosm portal: gearing up for 1000 fungal genomes[END_REF]. Because the number of sequenced genomes has increased dramatically in the last years, we took the opportunity of this study to reevaluate the presence of fRmsr genes in other multicellular eukaryotes. We searched for fRmsr genes in the plant and animal genomes available in the nr database of NCBI by BLAST search and found only a handful of multicellular eukaryotes apparently possessing a gene coding for a fRMsr (Table S1). These organisms, a plant and a few insects, are not phylogenetically related, indicating that the gene is not conserved in their lineage. This highlights the prevalence of fRmsr genes in the fungal kingdom, as opposite to their extremely low occurrence in other multicellular eukaryotes. What would be the advantage for the fungi to produce fRMsr whereas other multicellular organisms do not? An obvious possibility would be to protect the intracellular free Met during exposure to oxidative constraints as it was shown for S. cerevisiae [START_REF] Le | Functional analysis of free methionine-R-sulfoxide reductase from Saccharomyces cerevisiae[END_REF]. Another possibility would be to allow the reduction of the free MetO coming from the external environment or after the degradation of proteins assimilated by the fungi and its use in protein synthesis and in sulfur metabolism. Combined with MsrA, the presence of fRMsr could virtually allow to reduce the complete pool of MetO coming from the external environment. This hypothesis arose from the observation of the presence of fRmsr genes in four Neocallimastigomycetes species that lacked MsrA and MsrB. Living in the anaerobic environment of ruminant gut, the fungi very likely do not suffer oxidative stress and the fRMsr could reduce the MetO coming from the animal's food. Interestingly, the fRmsr genes in these

Neocallimastigomycetes have been probably acquired through horizontal gene transfer, reinforcing the idea that a fRMsr would provide a selective advantage in an environment unlikely to generate oxidative constraints.

We found few variations in the numbers of gene copies of each type of Msr, with only 74 genomes lacking one gene, and 72 genomes with an extra copy of at least one msr gene. In both cases, it roughly corresponded to 10 percent of the analyzed genomes. Our study revealed the absence of both MsrA and MsrB in 15 species of fungi, twelve of these being also devoid of fRMsr. The lack of all Msr appears to be related to the fungal lifestyles, since it concerned the nine Microsporidia, and the Pneumocystis jirovecii species, which live as intracellular parasites of metazoans. These 10 species have small genomes with reduced numbers of genes, from 1,831 for Encephalitozoon romaleae SJ-2008 to 3,632 for Enterocytozoon bieneusi H348, as compared to the average ~12,200 genes for the fungal genomes considered in this study. It has been proposed that the intracellular lifestyle allowed for genome compaction and gene loss, making the fungi highly dependent on the infected cell for numerous biochemical pathways [START_REF] Katinka | Genome sequence and gene compaction of the eukaryote parasite Encephalitozoon cuniculi[END_REF]. Our results suggest the fungi could also rely on the host detoxication system for the protection against oxidative constraints and limitation of protein oxidation. The absence of Msr was also observed in Neocallimastigomycetes, which live in the anaerobic environment of ruminant gut, where the protection of proteins from oxidative damage is likely not crucial. This hypothesis is reinforced by the fact that the numbers of genes in Neocallimastigomycetes genomes is above the average of the analyzed fungi (~15,500) and thus, the loss of msr genes was not due to a global genome compaction, but potentially due to the lack of selection pressure. Besides these species, for which the lack of Msr is consistent with the lifestyle, we also identified 12 other species for which msrA or msrB genes were absent from the genomes. As they are very likely living in aerobic conditions and are probably exposed to protein oxidation, the lack of MsrA or MsrB is surprising and we cannot exclude that the missing genes are due to incomplete genome sequencing. On the other hand, most of the 37 analyzed genomes from Glomeromycotina, Pucciniomycotina (Melampsora and Puccinia) and Taphrinomycotina (Schizosaccharomyces) species lacked the fRMsr. These fungi may have lost the capacity to reduce the free Met-R-O, as it has been shown for mammals [START_REF] Lee | Mammals reduce methionine-S-sulfoxide with MsrA and are unable to reduce methionine-R-sulfoxide, and this function can be restored with a yeast reductase[END_REF], or the MsrB might have significant activity on the free Met-R-O, as suggested for plants [START_REF] Le | Diversity of plant methionine sulfoxide reductases B and evolution of a form specific for free methionine sulfoxide[END_REF].

Regarding the presence of supernumerary copies of msr genes in few genomes, it seems not to be related to the phylogeny, nor to the numbers of genes per genome, as the genomes of fungi having two copies or more of one msr gene contain an average number of ~12,700 genes, similar to the average number of genes in the considered genomes (~12,200). In most cases, the presence of several copies was due to gene duplications, as indicated by the close phylogenetic relationships of paralogous proteins and the conservation of canonical features. However, in few fungi, the presence of extra copies was due to horizontal gene transfers, like for msrsA genes in Gonapodya prolifera, and msrB genes in Arthrobotrys oligospora, Monacrosporium haptotylum and Protomyces lactucaedebilis. Together with the horizontal transfers of fRmsr genes in Neocallimastigomycetes, most of these events were strongly supported by the phylogeny and parametric values as well as by shared ecological niches for donor bacteria and recipient fungi. The case of Gonapodya prolifera msrA was remarkably interesting as it illustrated a prokaryote-to-eukaryote transfer of a selenoprotein gene. If such a transfer was observed from bacteria to archaea [START_REF] Zhang | Dynamic evolution of selenocysteine utilization in bacteria: a balance between selenoprotein loss and evolution of selenocysteine from redox active cysteine residues[END_REF], to our knowledge, the bacteria-to-eukaryote transfer of a selenoprotein gene has never been described. As this fungus possesses an eukaryotic selenocysteine insertion machinery [START_REF] Mariotti | Utilization of selenocysteine in earlybranching fungal phyla[END_REF], we hypothesize that the transferred prokaryotic msrA gene was compatible with the eukaryotic machinery, or that it was modified after the transfer to allow proper Sec insertion. In all cases, the presence of several Msr gene copies could allow a beneficial increase in gene dosage or, alternatively, different spatial and temporal expression profiles during the life cycle of the fungi through distinct transcriptional activities. It could also allow different subcellular targeting as shown for plant

Msrs [START_REF] Tarrago | Protein-repairing methionine sulfoxide reductases in photosynthetic organisms: gene organization, reduction mechanisms, and physiological roles[END_REF]. Indeed, in 23 fungal genomes, out of the 59 having more than one msrA gene, the MsrAs were predicted to be addressed to different subcellular compartments. Similarly, MsrBs from 7 genomes out of the 15 with more than one gene were predicted to be addressed to different cellular compartments.

Conversely, all the multicopy fRMsrs were predicted to be cytoplasmic. Our results indicate that, as experimentally demonstrated for yeast [START_REF] Kaya | Compartmentalization and regulation of mitochondrial function by methionine sulfoxide reductases in yeast[END_REF], most fungi might not have the ability to reduce the two diastereomers of protein-bound MetO in each subcellular compartment, suggesting variations in protein oxidation in the different cellular compartments. Another interesting aspect is the presence of potentially secreted Msrs in few fungal species, mostly from the Ascomycota genera Aspergillus and Penicillium.

Although it remains to be demonstrated experimentally, the presence of Msrs in the extracellular environment of the fungal cells could help to protect from oxidation the numerous secreted carbohydrate-active enzymes ('CAZymes') and other enzymes used by saprotrophic and pathogen fungi to degrade plant cell walls or insect chitins [START_REF] Peng | The draft genome sequence of the ascomycete fungus Penicillium subrubescens reveals a highly enriched content of plant biomass related CAZymes compared to related fungi[END_REF].

Finally, we observed that most MsrAs, MsrBs and fRMsrs had conserved canonical features, but we also highlighted potentially interesting discrepancies. For the three types of Msrs, a few sequences lacked the resolving Cys at conserved positions and might use alternative Cys residues for the regeneration of their activity, whereas others possessed only the catalytic Cys and might be regenerated by the direct reduction of the sulfenic acid formed after MetO reduction, as shown for Msrs from other organisms [START_REF] Tarrago | Plant thioredoxin CDSP32 regenerates 1-cys methionine sulfoxide reductase B activity through the direct reduction of sulfenic acid[END_REF][START_REF] Cao | Methionine sulfoxide reductase B3 requires resolving cysteine residues for full activity and can act as a stereospecific methionine oxidase[END_REF]. The presence of Sec in two Gonapodya prolifera MsrAs might confer them a catalytic advantage, similarly to other Sec-MsrAs [START_REF] Kim | Catalytic advantages provided by selenocysteine in methionine-S-sulfoxide reductases[END_REF]. We also observed that ~5% of fungal MsrAs harbor a GCYW motif containing the catalytic Cys, where the Tyr residue replaces the Phe residue of the canonical GCFW motif. To our knowledge, no MsrA with such motif has been described to date. Because of the similarity between Phe and Tyr physicochemical properties, this substitution should not prevent the catalysis, but it could induce a change in substrate specificity, as demonstrated for the E. coli MsrA. Indeed, direct evolution assays showed that the substitution of the Phe by a Leu conferred to the enzyme the capacity to efficiently reduce alkyl-aryl sulfoxides [START_REF] Tarallo | Development of a simple high-throughput assay for directed evolution of enantioselective sulfoxide reductases[END_REF]. Similarly, the atypical HYCIN motif of the Protomyces lactucaedebilis MsrB could modify the enzyme substrate specificity. The substitution of residues around the catalytic Cys of Msrs could affect the specificity for oxidized protein substrates or free MetO, but also potentially confer to the enzymes the ability to reduce other molecules such as the oxidized thioether-containing metabolites involved in sulfur metabolism (e.g S-adenosyl homocysteine, methylthio-ribose…) [START_REF] Pirkov | A complete inventory of all enzymes in the eukaryotic methionine salvage pathway[END_REF]. It would be interesting to evaluate the catalytic properties of these atypical Msrs on various sulfoxide-containing substrates.

  v1.0; Amniculicola lignicola CBS 123094 v1.0; Ampelomyces quisqualis HMLAC05119 v1.0; Aplosporella prunicola CBS 121.167 v1.0; Aulographum hederae v2.0; Bimuria novae-zelandiae CBS 107.79 v1.0; Byssothecium circinans CBS 675.92 v1.0; Cercospora zeae-maydis v1.0; Clathrospora elynae CBS 161.51 v1.0; Cucurbitaria berberidis CBS 394.84 v1.0; Decorospora gaudefroyi v1.0; Delitschia confertaspora ATCC 74209 v1.0; Delphinella strobiligena CBS 735.71 v1.0; Didymella exigua CBS 183.55 v1.0; Dissoconium aciculare v1.0; Dothidotthia symphoricarpi v1.0; Elsinoe ampelina CECT 20119 v1.0; Eremomyces bilateralis CBS 781.70 v1.0; Hortaea acidophila CBS 113389 v1.0; Karstenula rhodostoma CBS 690.94 v1.0; Lentithecium fluviatile v1.0; Lindgomyces ingoldianus ATCC 200398 v1.0; Lineolata rhizophorae ATCC 16933 v1.0; Lizonia empirigonia CBS 542.76 v1.0; Lophiostoma macrostomum v1.0; Lophiotrema nucula CBS 627.86 v1.0; Lophium mytilinum CBS 269.34 v1.0; Macroventuria anomochaeta CBS 525.71 v1.0; Massarina eburnea CBS 473.64 v1.0; Massariosphaeria phaeospora CBS 611.86 v1.0; Melanomma pulvis-pyrius v1.0; Microthyrium microscopicum CBS 115976 v1.0; Myriangium duriaei CBS 260.36 v1.0; Mytilinidion resinicola CBS 304.34 v1.0; Ophiobolus disseminans CBS 113818 v1.0; Patellaria atrata v1.0; Phoma tracheiphila IPT5 v1.0; Piedraia hortae CBS 480.64 v1.1; Pleomassaria siparia v1.0; Polychaeton citri v1.0; Polyplosphaeria fusca CBS 125425 v1.0; Pseudovirgaria hyperparasitica CBS 121739 v1.0; Rhizodiscina lignyota CBS 133067 v1.0; Saccharata proteae CBS 121410 v1.0; Setomelanomma holmii CBS 110217 v1.0; Sporormia fimetaria v1.0; Teratosphaeria nubilosa CBS 116005 v1.0; Tothia fuscella CBS 130266 v1.0; Trematosphaeria pertusa CBS 122368 v1.0; Trichodelitschia bisporula CBS 262.69 v1.0; Verruculina enalia CBS 304.66 v1.0; Viridothelium virens v1.0; Westerdykella ornata CBS 379.55 v1.0; Zasmidium cellare ATCC 36951 v1.0; Zopfia rhizophila v1.0 [51]; Acaromyces ingoldii MCA 4198 v1.0; Ceraceosorus guamensis MCA 4658 v1.0; Jaminaea sp. MCA 5214 v1.0; Meira miltonrushii MCA 3882 v1.0; Pseudomicrostroma glucosiphilum gen et sp. nov. MCA 4718 v1.0; Testicularia cyperi MCA 3645 v1.0; Tilletiopsis washingtonensis MCA 4186 v1.0; Violaceomyces palustris SA 807 v1.0 [52]; Acidomyces richmondensis BFW; Acidomyces richmondensis BFW [53]; Acremonium chrysogenum ATCC 11550 [54]; Agaricus bisporus var bisporus (H97) v2.0; Agaricus bisporus var. burnettii JB137-S8; Gigaspora rosea v1.0; Rhizophagus cerebriforme DAOM 227022 v1.0; Rhizophagus diaphanus v1.0 [55]; Amanita muscaria Koide v1.0; Gymnopus luxurians v1.0; Hebeloma cylindrosporum h7 v2.0; Hydnomerulius pinastri v2.0; Hypholoma sublateritium v1.0; Laccaria amethystina LaAM-08-1 v2.0; Paxillus adelphus Ve08.2h10 v2.0; Paxillus involutus ATCC 200175 v1.0; Piloderma croceum F 1598 v1.0; Pisolithus microcarpus 441 v1.0; Pisolithus tinctorius Marx 270 v1.0; Plicaturopsis crispa v1.0; Scleroderma citrinum Foug A v1.0; Sebacina vermifera MAFF 305830 v1.0; Sphaerobolus stellatus v1.0; Suillus luteus UH-Slu-Lm8-n1 v3.0; Tulasnella calospora AL13/4D v1.0; Oidiodendron maius Zn v1.0 [56]; Amanita thiersii Skay4041 v1.0 [57]; Ambrosiozyma philentoma NRRL Y-7523; Candida boidinii NRRL Y-2332; Citeromyces matritensis NRRL Y-2407; Nakazawaea wickerhamii NRRL Y-2563; Peterozyma xylosa NRRL Y-12939; Saccharomycopsis capsularis NRRL Y-17639; Saturnispora dispora NRRL Y-1447 [58]; Amorphotheca resinae v1.0; Meliniomyces bicolor E v2.0; Meliniomyces variabilis F v1.0;

  creatinivora 62-1032 v1.0; Kockovaella imperatae NRRL Y-17943 v1.0; Naematella encephela UCDFST 68-887.2 v1.0; Clohesyomyces aquaticus v1.0; Pseudomassariella vexata CBS 129021 v1.0; Protomyces lactucaedebilis 12-1054 v1.0; Lobosporangium transversale NRRL 3116 v1.0; Absidia repens NRRL 1336 v1.0; Hesseltinella vesiculosa NRRL3301 v2.0; Rhizopus microsporus var. microsporus ATCC52813 v1.0; Syncephalastrum racemosum NRRL 2496 v1.0; Basidiobolus meristosporus CBS 931.73 v1.0; Linderina pennispora ATCC 12442 v1.0; Catenaria anguillulae PL171

  GYXC176 motif but possessed two to four Cys in a Q[C/S/K]X2KX[C/N][C/X]XI[R/L]CYG motif, similar to poplar MsrAs[START_REF] Rouhier | Functional and structural aspects of poplar cytosolic and plastidial type a methionine sulfoxide reductases[END_REF]. Some other sequences possessed a Cys residue in the C-terminal region, but not in a GYXC motif, and others lacked any potential resolving Cys (Data S2C). Finally, a special case could be made for MsrAs from the early-diverging fungus Gonapodya prolifera (Monoblepharidomycetes). This fungus has three non-canonical MsrAs, two of which had the catalytic Cys replaced by a Sec[START_REF] Mariotti | Utilization of selenocysteine in earlybranching fungal phyla[END_REF]. Each had another Cys outside the conserved position of the resolving Cys in canonical fungal MsrAs. These two MsrA sequences had high similarity with the Sec-MsrAs from the bacterium Alkaliphilus oremlandii and the single-cell green alga Chlamydomonas reinhardtii, previously shown to use the Sec residue for the regeneration of their activity[START_REF] Kim | Catalytic advantages provided by selenocysteine in methionine-S-sulfoxide reductases[END_REF][START_REF] Lee | Evidence for the dimerization-mediated catalysis of methionine sulfoxide reductase A from Clostridium oremlandii[END_REF] (Data S2D).

Figure 2 .

 2 Figure 2. Potential subcellular targeting of fungal Msrs. The circle charts show the subcellular predictions for fungal MsrA (A), MsrB (B) and fRMsr (C), the proportion of proteins predicted to be localized in the cytosol ('Cyto.'), to be addressed to the mitochondria ('Mito.') or secreted ('Secr.'). The label 'Other' indicates the proportion of proteins predicted to be addressed to other compartments or for which no consensus prediction was obtained.

  Finally, another unusual feature was found in the MsrB from the taphrinomycotina Protomyces lactucaedebilis, with the location of the catalytic Cys in an HYCIN motif, instead of the typical RXCXN motif (Data S3C). Interestingly, searching in the NCBI nr database, we found 50 sequences, mainly from poorly characterized bacteria and archaea, that possessed this unusual motif (Data S3D). Considering His and Arg have similar physicochemical properties, we anticipate such non-canonical MsrBs might have conserved a catalytic activity.

Figure 3 .

 3 Figure 3. Protein sequence characteristics of canonical fungal Msrs. In this representation of canonical MsrAs (A), MsrBs, (B) and fRMsrs (C), the catalytic Cys (in red), the resolving Cys (in green) and the residues previously shown to be involved in catalysis and/or substrate binding (in blue) are shown. The residues in purple are conserved in all canonical fungal Msrs. In B, the Cys residues labeled in orange correspond to Zn binding residues. The numberings are based on S. cerevisiae MsrA (A), MsrB (B) and fRMsr (C).

Figure 4 .

 4 Figure 4. Phylogenetic analysis of MsrA candidates to horizontal gene transfer. The fungal MsrAs tested for horizontal gene transfer (in black) are from the following fungal genomes (protein accessions in brackets): Piptocephalis cylindrospora RSA 2659 single-cell v3.0 (20412); Syncephalis pseudoplumigaleata Benny S71-1 single-cell v1.0 (33365; 23947); Thamnocephalis sphaerospora RSA 1356 single-cell v1.0 (27979); Coemansia reversa NRRL 1564 v1.0 (43282); Dimargaris cristalligena RSA 468 single-cell v1.0 (33956); Linderina pennispora ATCC 12442 v1.0 (280520); Smittium culicis GSMNP (5372; 7035); Smittium culicis ID-206-W2 (5101; 8129); Smittium mucronatum ALG-7-W6 (4510; 6899; 7265); Zancudomyces culisetae COL-18-3 (924) and Gonapodya prolifera v1.0 (135492; 159800/159692; 57902). The MsrA sequences from fungi, plants, amoeba and bacteria are in blue, green, yellow and purple, respectively. The phylogenetic tree was built with RAxML v. 8.2 [348] and represented using iTOL (https://itol.embl.de/) [412]. The branches with bootstrap values over 70 are in red.

Figure 5 .

 5 Figure 5. Phylogenetic analysis of MsrB candidates to horizontal gene transfer. The fungal MsrBs tested for horizontal gene transfer (in black) are from the following genomes (protein accessions in brackets): Arthrobotrys oligospora ATCC 24927 (9001); Monacrosporium haptotylum CBS 200.50 (10089) and Protomyces lactucaedebilis 12-1054 v1.0 (391028).The MsrBs sequences from fungi and bacteria are in blue and purple, respectively. The phylogenetic tree was built with RAxML v. 8.2[START_REF] Stamatakis | RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies[END_REF] and represented using iTOL (https://itol.embl.de/)[START_REF] Letunic | Interactive Tree Of Life (iTOL) v4: recent updates and new developments[END_REF]. The branches with bootstrap values over 70 are in red.

Figure 6 .

 6 Figure 6. Phylogenetic analysis of fRMsr candidates to horizontal gene transfer. The fungal fRMsrs tested for horizontal gene transfer (in black) are from the following genomes (protein accessions are in brackets): Anaeromyces robustus v1.0 (328892); Neocallimastix californiae G1 v1.0 (697339) andOrpinomyces sp (1191427). The fRMsrs sequences from fungi and bacteria are in blue and purple, respectively. Red stars indicate fRmsr genes possibly acquired by horizontal gene transfer, which were not identified in our genomics search but found as homologs of the selected fungal fRMsr candidates. The phylogenetic tree was built with RAxML v 8.2[START_REF] Stamatakis | RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies[END_REF] and represented using iTOL (https://itol.embl.de/)[START_REF] Letunic | Interactive Tree Of Life (iTOL) v4: recent updates and new developments[END_REF]. The branches with bootstrap values over 70 are in red.

Table 1 ,

 1 Data S1-4), indicating that gene enrichment was not a characteristic of a specific fungal group. 467

Table 1 .

 1 Fungi with five or six Msr genes. 468

						Nb. Genes
	Genome	MsrA	MsrB	fRMsr	Lifestyle	(Genome size in	Ref.
						Mbp)
	Agaricomycetes
	Crucibulum laeve CBS 166.37 v1.0	4	1	1		14,218 (44)	[351]
	Dendrothele bispora CBS 962.96 v1.0	4	1	1		33,645 (131)	[351]
	Tulasnella calospora AL13/4D v1.0	3	1	1	Aerobic (saprotroph)	19,659 (62)	[352]
	Ramaria rubella (R. acris) UT-36052-T v1.0	1	1	4		14,207 (62)	[353]
	Coprinellus micaceus FP101781 v2.0	1	1	3		23,559 (77)	[351]
	Dothideomycetes
	Hortaea werneckii EXF-2000 M0 v1.0	2	2	2	Aerobic	15,748 (50)	[167]
	Sordariomycetes
	Coniochaeta sp. 2T2.1 v1.0	2	2	2	Aerobic (phytopathogen)	24,735 (74)	[121]
	Saccharomycotina
	Pichia kudriavzevii CBS573	3	1	1		5,140 (11)	[354]
	Yarrowia lipolytica (strain CLIB122)	3	1	1		6,447 (21)	[355]
	Yarrowia lipolytica CLIB89(W29)	3	1	1		7,919 (21)	[356]
	Yarrowia lipolytica FKP355 v1.0	3	1	1		6,858 (20)	[357]
	Yarrowia lipolytica PO1f v1.0	3	1	1		6,798 (20)	[358]
	Yarrowia lipolytica YB392 v1.0	3	1	1	Aerobic	6,750 (20)	[359]
	Yarrowia lipolytica YB419 v1.0	3	1	1		6,751 (20)	[359]
	Yarrowia lipolytica YB420 v1.0	3	1	1		6,772 (20)	[359]
	Yarrowia lipolytica YB566 v1.0	3	1	1		6,764 (20)	[359]
	Yarrowia lipolytica YB567 v1.0	3	1	1		6,776 (20)	[359]
	Yarrowia lipolytica YlCW001 v1.0	3	1	1		6,800 (20)	[358]
	Mucoromycotina
	Absidia repens NRRL 1336 v1.0	3	1	1	Aerobic (saprotroph)	14 919 (47)	[360]
	Rhizopus microsporus ATCC11559 v1.0	2	1	2		11,135 (24)	[273]
	Rhizopus microsporus var. chinensis CCTCC M201021	2	1	2	Aerobic (phytopathogen)	17,676 (46)	[274]
	Kickxellomycotina
	Smittium mucronatum ALG-7-W6 Smittium culicis GSMNP	3 2	1 3	1 1	Anaerobic (insect gut)	8 247 (102) 12 166 (77)	[361] [361]
	Monoblepharidomycetes
	Gonapodya prolifera v1.0	3	1	1	Aerobic (aquatic)	13 902 (49)	[362]

Table 2 .

 2 Fungal genomes lacking one or more Msr types. 500

						Nb. Genes
	Genome	MsrA	MsrB fRMsr	Lifestyle	(Genome	Ref.
						size in Mbp)
	Microsporidia
	Antonospora locustae HM-2013	N	N	N		2,606 (6)	[364]
	Encephalitozoon cuniculi GB-M1	N	N	N		1,996 (2)	[365]
	Encephalitozoon hellem ATCC 50504	N	N	N		1,847 (2)	[366]
	Encephalitozoon intestinalis ATCC 50506 Encephalitozoon romaleae SJ-2008 Enterocytozoon bieneusi H348	N N N	N N N	N N N	Intracellular parasite of metazoans	1,833 (2) 1,831 (2) 3,632 (4)	[367] [366] [368]
	Mitosporidium daphniae UGP3	N	N	N		3,330 (6)	[369]
	Nematocida parisii ERTm1	N	N	N		2,661 (4)	[370]
	Nosema ceranae BRL01	N	N	N		2,060 (8)	[371]
	Neocallimastigomycetes
	Piromyces finnis v3.0	N	N	N		10,992 (56)	[372]
	Piromyces sp. E2 v1.0 Anaeromyces robustus v1.0 Neocallimastix californiae G1 v1.0	N N N	N N N	N Y Y	Anaerobic (ruminant gut)	14,648 (71) 12,832 (72) 20,219 (193) [372] [372] [372]
	Orpinomyces sp.	N	N	Y		18,936 (101) [373]
	Taphrinomycotina
	Pneumocystis jirovecii	N	N	N	Intracellular parasite of metazoans	3,520 (8)	[374]
	Neolecta irregularis DAH-1 v1.0	Y	Y	N		6,674 (15)	[375]
	Schizosaccharomyces cryophilus OY26	Y	Y	N		5,180 (12)	[376]
	Schizosaccharomyces japonicus yFS275	Y	Y	N	Aerobic	4,878 (12)	[376]
	Schizosaccharomyces octosporus yFS286	Y	Y	N		4,986 (12)	[376]
	Schizosaccharomyces pombe	Y	Y	N		5,134 (13)	[377]
	Agaricomycetes
	Moniliophthora perniciosa FA553	N	Y	Y	Aerobic (phytopathogen)	13,560 (18)	[378]
	Eurotiomycetes
	Penicillium coprophilum IBT 31321	N	Y	Y		8,999 (28)	[379]
	Penicillium flavigenum IBT 14082	N	Y	Y		10,994 (33)	[379]
	Penicillium polonicum IBT 4502	N	Y	Y	Aerobic	10,694 (32)	[379]
	Aspergillus bombycis NRRL 26010	Y	Y	N		12,265 (37)	[380]
	Aspergillus zonatus v1.0	Y	Y	N		9,886 (29)	[70]
	Sordiariomycetes
	Magnaporthiopsis poae ATCC 64411	N	Y	Y	Aerobic (phytopathogen)	12,335 (40)	[381]
	Pochonia chlamydosporia 170	Y	Y	N	Aerobic (animal pathogen)	14,204 (44)	[382]
	Saccharomycotina
	Saturnispora dispora NRRL Y-1447	N	Y	Y	Aerobic	4,881 (10)	[383]
	Tremellomycetes
	Trichosporon asahii var. asahii CBS 2479	Y	N	Y	Aerobic (human	8,300 (24)	[384]
	Trichosporon asahii var. asahii CBS 8904	Y	N	Y	pathogen)	8,507 (25)	[385]
	Dothideomycetes
	Clohesyomyces aquaticus v1.0	Y	N	Y	Aerobic (saprotrophic)	15,810 (50)	[360]
	Lecaronomycetes
	Cladonia grayi Cgr/DA2myc/ss v2.0	Y	N	Y	Aerobic (lichen)	11,389 (35)	[386]
	Zoopagomycotina
	Piptocephalis cylindrospora RSA 2659 single-cell v3.0	Y	N	Y	Aerobic (mycoparasite)	4,301 (11)	[387]

Table 3 .

 3 Potential horizontal msr gene transfers from bacteria to fungi For both Gonapodya prolifera selenocysteine-containing MsrAs, the MycoCosm accession numbers presented here refer to only a part of the proteins. Their reconstructed complete gene structures are presented in Fig.S4. The horizontal gene transfers of Anaeromyces robustus and Piromyces sp. fRmsr genes have been shown recently[START_REF] Murphy | Horizontal Gene Transfer as an Indispensable Driver for Evolution of Neocallimastigomycota into a Distinct Gut-Dwelling Fungal Lineage[END_REF]. The protein accessions are from NCBI (https://www.ncbi.nlm.nih.gov/protein). Orpinomyces sp. genes on the scaffolds are not publically available. The calculations were made on the

				Candidate gene	Mean of all genes from the scaffold
	Genome	Protein accession (Scaffold number)	Nb. of exons	GC content (%)	GC third base (%)	Nb. of exons	GC content (%)	GC third base (%)	Organisms with the closest Msr homolog
	MsrA
	Monoblepharidomycetes
	Gonapodya prolifera v1.0 Gonapodya prolifera v1.0	135492 a (36) 159800/ (105) 159692 a	3 4	56.4 58.9	61.3 69.6	5.7 ± 4.8 5.8 ± 3.8	56.5 ± 3.4 57.3 ± 4.2	65.2 ± 8.5 67.4 ± 8.2	Alkaliphilus oremlandii OhILAs
	MsrB
	Orbiliomycetes
	Arthrobotrys oligospora ATCC 24927	9001 (00170)	1	55.6	56.5	3.0 ± 2.3	48.0 ± 2.4	48.0 ± 4.6	Sphingomonas bacterium
	Monacrosporium haptotylum CBS 200.50	10089 (00497)	1	55.8	58.0	4.3 ± 4.3	49.0 ± 2.7	52.9 ± 5.3	Sphingomonas bacterium
	Taphrinomycotina
	Protomyces lactucaedebilis 12-1054 v1.0	391028 (3)	1	54.9	52.1	1.6 ± 1.0	52.6 ± 1.9	54.4 ± 5.9	Calothrix parasitica
	fRMsr
	Neocallimastigomycetes
	Anaeromyces robustus v1.0	328892 b (243)	1	30.1	14.8	4.2 ± 4.5	26.3 ± 5.0	11.8 ± 3.3	Pseudobutyrivibrio sp. LB2011
	Neocallimastix californiae G1 v1.0	697339 (7)	1	27.5	9.8	4.6 ± 4.4	28.8 ± 4.4	16.7 ± 5.4	Erysipelotrichaceae bacterium
	Orpinomyces sp.	1191427 (d_7180000085738)	1	34.8	22.5	3.0 ± 2.9 d	27.2 ± 5.4 d	16.0 ± 7.8 d	Pseudobutyrivibrio ruminis
	Piromyces sp.	AWI66787.1 b,c	1	31.3	9.9	n.d.	33.2 ± 5.7	23.3 ± 11.1	Pseudobutyrivibrio ruminis
	Mucoromycotina
	Mortierella sp. GBA39	KAF9144191.1 c (77)	1	55.2	62.4	1.4 ± 0.7	57.1 ± 3.5	71.3 ± 6.0	Paenibacillus rhizosphaerae
	complete genome.
	n.d, not determined.

a b c d The positions of

Acknowledgements.

The authors thank the U.S. Department of Energy Joint Genome Institute (JGI), Dr. Sebastien Duplessis (Université de Lorraine, INRAE, UMR1136, Interactions Arbres/Microorganismes, France) of and Dr. Christine Smart (Cornell University, NY, USA) for sharing Melampsora allii-populina 12AY07 v1.0 and Melampsora americana R15-033-03 v1.0 unpublished genome data, respectively. Financial support to Hayat Hage from Région SUD Provence-Alpes-Côte d'Azur and The French National Research Institute for Agriculture, Food and Environment (INRAE) is acknowledged.

Declaration of competing interest.

None.