Jason Banda

Kevin Bellande

Daniel Von Wangenheim

Tatsuaki Goh

Soazig Guyomarc'h

Laurent Laplaze
email: laurent.laplaze@ird.fr

Malcolm J Bennett +
email: malcolm.bennett@nottingham.ac.uk

Lateral root formation in arabidopsis: a well-ordered LRexit

Keywords: Auxin, biomechanics, live microscopy, cell fate, environmental patterning

New dimensions to lateral root morphogenesis in arabidopsis

Lateral roots originate primarily from pericycle tissue in angiosperm species [START_REF] Casimiro | Dissecting Arabidopsis lateral root development[END_REF][START_REF] Torres-Martínez | Lateral Root Primordium Morphogenesis in Angiosperms[END_REF]. The pericycle consists of a single cell layer surrounding the vascular tissues and which is overlain by endodermal, cortex and epidermal tissues (Fig. 1). In arabidopsis (Arabidopsis thaliana), lateral roots derive from six pericycle cell files overlaying the xylem pole (Fig. 1) [START_REF] Casero | Lateral root initiation by asymmetrical transverse divisions of pericycle cells in adventitious roots ofAllium cepa[END_REF][START_REF] Laskowski | Formation of lateral root meristems is a two-stage process[END_REF][START_REF] Wangenheim | Rules and self-organizing properties of post-embryonic plant organ cell division patterns[END_REF].

Phloem-pole pericycle (PPP) cells are reported to be mitotically dormant, while xylem-polepericycle (XPP) cells retain stem cell activity after leaving the primary root meristem and can therefore form LRs [START_REF] Parizot | Diarch symmetry of the vascular bundle in Arabidopsis root encompasses the pericycle and is reflected in distich lateral root initiation[END_REF].

In arabidopsis, lateral root development can be divided in five main steps (Figure 1A): [START_REF] Casimiro | Dissecting Arabidopsis lateral root development[END_REF] pre-branch site formation which takes place in the basal meristem/elongation zone [START_REF] De Smet | Auxin-dependent regulation of lateral root positioning in the basal meristem of Arabidopsis[END_REF][START_REF] Moreno-Risueno | Oscillating Gene Expression Determines Competence for Periodic Arabidopsis Root Branching[END_REF];

(2) LR initiation which features pericycle nuclear migration to a common cell wall between pairs of founder cells (LRFCs) followed by an asymmetric cell division in the differentiation zone [START_REF] Casimiro | Auxin transport promotes Arabidopsis lateral root initiation[END_REF][START_REF] Goh | The establishment of asymmetry in Arabidopsis lateral root founder cells is regulated by LBD16/ASL18 and related LBD/ASL proteins[END_REF]; [START_REF] Casero | Lateral root initiation by asymmetrical transverse divisions of pericycle cells in adventitious roots ofAllium cepa[END_REF] LR morphogenesis in which the LRFCs divide further to form a lateral root primordium (LRP) that eventually acquires a root meristem organization [START_REF] Malamy | Organization and cell differentiation in lateral roots of Arabidopsis thaliana[END_REF][START_REF] Goh | Quiescent center initiation in the Arabidopsis lateral root primordia is dependent on the SCARECROW transcription factor[END_REF]; [START_REF] Laskowski | Formation of lateral root meristems is a two-stage process[END_REF] concomitant LR emergence where the new organ grows through overlaying tissue layers to emerge from the parent root in the differentiation zone [START_REF] Du | PLETHORA transcription factors orchestrate de novo organ patterning during Arabidopsis lateral root outgrowth[END_REF] and finally [START_REF] Wangenheim | Rules and self-organizing properties of post-embryonic plant organ cell division patterns[END_REF] LR meristem activation corresponding to the initiation of cell divisions in the newly emerged lateral root meristem [START_REF] John | A pathway for lateral root formation in Arabidopsis thaliana[END_REF].

Until recently, arabidopsis lateral root development was studied in a 2D manner, primarily visualising primordia stages from a 'side on' viewpoint (Fig 1 ) and not considering the wider 3D morphogenic events taking place. Advanced microscopy approaches such as confocal imaging and Light Sheet Fluorescence Microscopy (LSFM, [START_REF] Ovečka | Multiscale imaging of plant development by light-sheet fluorescence microscopy[END_REF]) has helped revolutionise the field's perspective, providing static [START_REF] Mairhofer | Extracting multiple interacting root systems using X-ray microcomputed tomography[END_REF][START_REF] Morris | Shaping 3D Root System Architecture[END_REF][START_REF] Goh | Long-term live-cell imaging approaches to study lateral root formation in Arabidopsis thaliana[END_REF][START_REF] Rellán-Álvarez | GLO-Roots: an imaging platform enabling multidimensional characterization of soil-grown root systems[END_REF] or real-time [START_REF] Wangenheim | Rules and self-organizing properties of post-embryonic plant organ cell division patterns[END_REF][START_REF] Mairhofer | Extracting multiple interacting root systems using X-ray microcomputed tomography[END_REF][START_REF] Rellán-Álvarez | GLO-Roots: an imaging platform enabling multidimensional characterization of soil-grown root systems[END_REF]20] 3D visualization of lateral root development (Figure 1B). For example, LR formation features a stereotypical sequence of cell layer generation [START_REF] Wangenheim | Rules and self-organizing properties of post-embryonic plant organ cell division patterns[END_REF][START_REF] Malamy | Organization and cell differentiation in lateral roots of Arabidopsis thaliana[END_REF]. However, 4D time-lapse imaging studies of LR morphogenesis revealed that after the first anticlinal cell division the pattern of divisions does not follow a specific sequence [START_REF] Wangenheim | Rules and self-organizing properties of post-embryonic plant organ cell division patterns[END_REF][START_REF] Goh | Quiescent center initiation in the Arabidopsis lateral root primordia is dependent on the SCARECROW transcription factor[END_REF]21]. There is a high level of plasticity in the spatiotemporal regulation of cell divisions with a higher order of rules to form the shape of a new root tip.

Patterning under pressure

LRP originate from pericycle cells located deep within the parental root and have therefore to pass through overlying endodermal, cortical and epidermal tissues before emerging [START_REF] Swarup | The auxin influx carrier LAX3 promotes lateral root emergence[END_REF] (Figure 1). This involves complex biomechanical interactions between the overlying tissues and the LRP that impact organ initiation and morphogenesis [21, [START_REF] Péret | Auxin regulates aquaporin function to facilitate lateral root emergence[END_REF][START_REF] Kumpf | Floral organ abscission peptide IDA and its HAE/HSL2 receptors control cell separation during lateral root emergence[END_REF][START_REF] Vermeer | A Spatial Accommodation by Neighboring Cells Is Required for Organ Initiation in Arabidopsis[END_REF].

During LR initiation, root cells surrounding new primordia actively adapt and remodel their properties to accommodate organ emergence [21]. For instance, during organ initiation LRFCs swell prior to asymmetric division which would require adjacent xylem and/or endodermal cells to adjust their volumes to allow the radial growth of pericycle cells (Figure 1) [START_REF] Vermeer | A Spatial Accommodation by Neighboring Cells Is Required for Organ Initiation in Arabidopsis[END_REF]. Intercellular connectivity between the LRFC and surrounding cells are likely to be involved in coordinating cell volume adjustments through control of callose deposition by plasmodesmata remodelling enzymes which regulate initiation of lateral roots [START_REF] Benitez-Alfonso | Symplastic Intercellular Connectivity Regulates Lateral Root Patterning[END_REF].

Interestingly, endodermal cell ablation is sufficient to trigger mitosis in the underlying pericycle cell [START_REF] Marhavý | Targeted cell elimination reveals an auxin-guided biphasic mode of lateral root initiation[END_REF]. Nevertheless, auxin co-treatment is required to observe an anticlinal division of the induced pericycle cells and initiate the lateral root organogenesis program [START_REF] Marhavý | Targeted cell elimination reveals an auxin-guided biphasic mode of lateral root initiation[END_REF]. In addition the cell wall remodelling enzyme EXPANSIN A1 (EXPA1), which may modulate the mechanical properties of the pericycle cell wall, is also required for radial expansion of LRFCs and to ensure the correct positioning of the first anticlinal divisions (Table 1) [START_REF] Ramakrishna | EXPANSIN A1-mediated radial swelling of pericycle cells positions anticlinal cell divisions during lateral root initiation[END_REF]. Hence, sufficient pericycle width appears necessary to trigger asymmetric pericycle cell divisions during lateral root initiation [START_REF] Ramakrishna | EXPANSIN A1-mediated radial swelling of pericycle cells positions anticlinal cell divisions during lateral root initiation[END_REF].

During LR morphogenesis, primordia have to develop under mechanical pressure from surrounding tissues. These mechanical constraints, rather than a stereotypical pattern of cell division, are responsible for determining LRP shape [START_REF] Wangenheim | Rules and self-organizing properties of post-embryonic plant organ cell division patterns[END_REF]21]. Plants, like animals, regulate morphogenesis of new organs by employing a stem cell niche regulated by a mitotically inactive organising centre [START_REF] Sarkar | Conserved factors regulate signalling in Arabidopsis thaliana shoot and root stem cell organizers[END_REF]. Interestingly, the organising centre appears once LRP form 4 cell layers (stage IV; Figure 2C) and just prior to growing through the endodermis [START_REF] Goh | Quiescent center initiation in the Arabidopsis lateral root primordia is dependent on the SCARECROW transcription factor[END_REF]. The endodermis contains an impermeable barrier termed the Casparian Strip (Figure 2) that was originally thought to regulate elemental and nutrient movement between inner and outer root tissues. However, the mechanical properties of the Casparian Strip may also provide information that impacts development of LRP. During the time course of LRP development, the mechanical properties of the endodermis cells are remodelled through an auxindependent pathway and this is necessary for LRP development to progress from stage V onwards in a wild-type fashion [21]. We suggest that the Casparian strip could behave like a "developmental traffic light" that holds back new LR (red light) until the organising centre and stem cell niche are set up (amber light) allowing the new organ to break through the endodermis and overlying tissues before emerging into the soil (green light) (Figure 2). This checkpoint system could also provide a mechanism for regulation of LR development by endogenous or environmental signals, allowing only LRP in optimal conditions to emerge [START_REF] Nacry | A role for auxin redistribution in the responses of the root system architecture to phosphate starvation in Arabidopsis[END_REF].

Breakthroughs in Lateral Root Emergence

New LRP must reprogram overlying cells to aid organ emergence. The hormone auxin functions as a local signal released by new LRP to facilitate this progression [START_REF] Swarup | The auxin influx carrier LAX3 promotes lateral root emergence[END_REF][START_REF] Vermeer | A Spatial Accommodation by Neighboring Cells Is Required for Organ Initiation in Arabidopsis[END_REF]. Key processes targeted by auxin in overlying cells include modifying their hydraulic properties, cell walls and Casparian strip [21, [START_REF] Péret | Auxin regulates aquaporin function to facilitate lateral root emergence[END_REF][START_REF] Vermeer | A Spatial Accommodation by Neighboring Cells Is Required for Organ Initiation in Arabidopsis[END_REF][START_REF] Roycewicz | Cell wall properties play an important role in the emergence of lateral root primordia from the parent root[END_REF][START_REF] Vermeer | Lateral root initiation in Arabidopsis thaliana: a force awakens[END_REF][START_REF] Reinhardt | Tonoplast aquaporins facilitate lateral root emergence[END_REF]. Regarding hydraulics, auxin represses the expression of almost every member of a family of water channels termed aquaporins [START_REF] Péret | Auxin regulates aquaporin function to facilitate lateral root emergence[END_REF].

Aquaporin genes encode plasma membrane (PIP class) or tonoplast (TIP class) localised water channels that regulate hydraulic properties of plant cells. Auxin appears to fine-tune the hydraulic properties of cells in the LRP and overlying tissues through its regulation of PIP and TIP genes spatial expression. Over-expressing and/or mutating PIP and TIP class aquaporins significantly delays LR emergence (Table 1) [START_REF] Péret | Auxin regulates aquaporin function to facilitate lateral root emergence[END_REF][START_REF] Reinhardt | Tonoplast aquaporins facilitate lateral root emergence[END_REF]. Hence, auxin-dependent spatio-temporal regulation of aquaporin expression appears critical for emergence of LRP.

The importance of these water channels results from new LRP becoming symplastically isolated from surrounding tissues soon after initiation through closure of plasmodesmata [START_REF] Benitez-Alfonso | Symplastic Intercellular Connectivity Regulates Lateral Root Patterning[END_REF].

Auxin originating from new LRP also causes overlying cells to modify their cell walls and undergo cell separation to enable new organs to emerge [START_REF] Swarup | The auxin influx carrier LAX3 promotes lateral root emergence[END_REF]. Auxin does this by triggering the sequential induction of the PIN3 auxin efflux carrier and then the LAX3 influx carrier in cells directly overlying new LRP [START_REF] Peret | Sequential induction of auxin efflux and influx carriers regulates lateral root emergence[END_REF]. A 3D root mathematical model indicated that, collectively, the distinct temporal patterns of induction of different classes of auxin carrier was necessary for preferential hormone accumulation in cells overlying LRP. This functions to focus the auxin-dependent induction of cell wall remodelling enzymes and, consequentially cell separation, in advance of the emerging organ [START_REF] Swarup | The auxin influx carrier LAX3 promotes lateral root emergence[END_REF][START_REF] Peret | Sequential induction of auxin efflux and influx carriers regulates lateral root emergence[END_REF].

Reactive oxygen species (ROS) have also been proposed to function as important signals during auxin-regulated LR formation. ROS treatment increases LR density and can restore LR formation in lines in which auxin-mediated cell wall accommodation and remodelling in LRPoverlying cells is disrupted. ROS are deposited in the apoplast of overlying cells during LR emergence following induction of RESPIRATORY BURST OXIDASE HOMOLOGS (RBOH) (Table 1) [START_REF] Orman-Ligeza | RBOH-mediated ROS production facilitates lateral root emergence in Arabidopsis[END_REF]. Disrupting (or enhancing) expression of RBOH in LRP or overlying root tissues decelerates (or accelerates) LR development and emergence. Hence, RBOH-mediated ROS production appears to facilitate LR outgrowth by promoting remodelling of overlying root cell walls. ROS levels generated within LRP are also important for organ emergence [START_REF] Orman-Ligeza | RBOH-mediated ROS production facilitates lateral root emergence in Arabidopsis[END_REF][START_REF] Manzano | The Emerging Role of Reactive Oxygen Species Signaling during Lateral Root Development[END_REF].

The MYB36 TF controls the expression of a subset of peroxidase genes (e.g. PER9) in boundary cells at the base of developing LRP. Mutating MYB36 causes LRP to adopt a flattened shape compared to the dome-like appearance of WT LRP, resulting in slower organ emergence. Reducing the levels of hydrogen peroxide (H2O2) in myb36-5 rescues the LR mutant phenotype (Table 1). This suggests that MYB36-dependent induction of peroxidases reduces H 2 O 2 levels thus defining the outer boundary of the growing LRP [START_REF] Fernández-Marcos | Control of Arabidopsis lateral root primordium boundaries by MYB36[END_REF].

Lateral Root Gene Regulatory Networks: learning lessons from primary root development

A large number of genes that regulate meristem patterning and maintenance in primary roots are also expressed during equivalent processes in lateral root development. For example, AP2-/ERF PLETHORA (PLT) transcription factors are major regulators of the gene regulatory network controlling primary root meristem patterning and maintenance [START_REF] Scheres | Coordination of growth in root and shoot apices by AIL/PLT transcription factors[END_REF]. PLT genes are also expressed during LRP development [START_REF] Hofhuis | Phyllotaxis and rhizotaxis in Arabidopsis are modified by three plethora transcription factors[END_REF]. Interestingly, PLT3, PLT5, and PLT7 were shown to control the onset of PLT1, PLT 2, and PLT4 gene expression in developing LRP (Figure 3). Accordingly, the triple plt3plt5plt7 mutant exhibits a PLT-null phenotype in which the first round of periclinal cell divisions, which creates a two layered LRP, is disrupted, resulting into a highly disorganized LRP with no properly defined meristem (Table 1). Surprisingly, transgenic expression of any PLT member from stage I onwards is sufficient to restore the formative periclinal divisions and subsequent organization of root meristem-like identities. This suggests that rather than target specificity of PLTs, precise timing in PLT expression controls critical events during LRP formation [START_REF] Du | PLETHORA transcription factors orchestrate de novo organ patterning during Arabidopsis lateral root outgrowth[END_REF].

Gene regulatory networks dependent on the GRAS-family SCARECROW (SCR)/SHORT-ROOT (SHR) transcription factors control the transition from stage II to stage III in LRP development (Figure 1) and thereby impact patterning of the quiescent centre (QC) at stage IV/V [START_REF] Goh | Quiescent center initiation in the Arabidopsis lateral root primordia is dependent on the SCARECROW transcription factor[END_REF][START_REF] Drapek | Uncovering Gene Regulatory Networks Controlling Plant Cell Differentiation[END_REF]. The observed onset of QC marker gene expression is concomitant of a major transition from an early morphogenesis phase to a late, meristem organization phase (Figure 1). Interestingly, the scr mutant is still able to establish a QC and a functional root meristem later in development, thus illustrating the robustness of the LR regulatory network [START_REF] Goh | Quiescent center initiation in the Arabidopsis lateral root primordia is dependent on the SCARECROW transcription factor[END_REF]. In the primary root tip SCR and PLT proteins were shown to directly induce expression of QC marker WOX5 by cooperative interaction with TCP to coordinate LR outgrowth in a time-and space-dependant manner (Figure 3) [START_REF] Shimotohno | Root stem cell niche organizer specification by molecular convergence of PLETHORA and SCARECROW transcription factor modules[END_REF].

Lateral Root Gene Regulatory Networks: emerging properties

Lateral root research has recently moved beyond studying the role of individual or a few genes to characterising many genes that compose regulatory networks controlling lateral root development employing systems and -omics based approaches. LR-related transcriptomic datasets have recently been produced in different conditions, monitoring gene expression dynamics during root branching in an unbiased fashion [START_REF] Birnbaum | A gene expression map of the Arabidopsis root[END_REF][START_REF] Himanen | Transcript profiling of early lateral root initiation[END_REF][START_REF] Brady | A high-resolution root spatiotemporal map reveals dominant expression patterns[END_REF][START_REF] De Smet | Receptor-like kinase ACR4 restricts formative cell divisions in the Arabidopsis root[END_REF][START_REF] Voß | The circadian clock rephases during lateral root organ initiation in Arabidopsis thaliana[END_REF]. These offer unprecedented resources to explore single gene expression dynamics and identify new candidate regulators of LR formation. More importantly understanding how these components interact to form regulatory networks is crucial if we want to understand the emerging properties of LR development [START_REF] Lucas | Plant systems biology: network matters[END_REF][START_REF] Haque | Computational prediction of gene regulatory networks in plant growth and development[END_REF].

LRP can be induced in a highly synchronised manner by gravistimulation [START_REF] Péret | Auxin regulates aquaporin function to facilitate lateral root emergence[END_REF][START_REF] Voß | The circadian clock rephases during lateral root organ initiation in Arabidopsis thaliana[END_REF]49].

Researchers have therefore exploited gravistimulation-based LR induction to sequentially sample root bending zones every 3 hours and to generate a high resolution time course transcriptomic dataset spanning LR induction to organ emergence and meristem activation [START_REF] Voß | The circadian clock rephases during lateral root organ initiation in Arabidopsis thaliana[END_REF]. This unique transcriptome resource offers unprecedented insights to explore expression dynamics during LR organogenesis, ranging from a single gene up to the network scale. More than 8,000 (out of 22,000) genes were differentially expressed during LRP formation. Clustering identified 77 distinct classes of gene expression profiles, highlighting the complex regulation of LR formation [START_REF] Voß | The circadian clock rephases during lateral root organ initiation in Arabidopsis thaliana[END_REF]. Interestingly a number of circadian-clock regulated genes were identified, revealing a surprising clock re-phasing mechanism during LR initiation, which were proposed to 'insulate' the LRP from diurnal fluctuations in water fluxes which may interfere with organ emergence [START_REF] Voß | The circadian clock rephases during lateral root organ initiation in Arabidopsis thaliana[END_REF]. This LR transcriptomic dataset has been used to infer the topology of the gene regulatory network coordinating lateral root morphogenesis [START_REF] Lavenus | Inference of the Arabidopsis lateral root gene regulatory network suggests a bifurcation mechanism that defines primordia flanking and central zones[END_REF]. The predicted GRN controlling primordia patterning was organised in two main auxin-regulated modules, an early network dependent on ARF7 and ARF19 (Figure 3), and a later network, involving ARF5. In general, genes in the early ARF7-associated sub-network are expressed in all primordia cells during early stages of LRP development and then their expression decreases and is confined to the base of LRP. Concomitant with this transition, expression of genes associated with the ARF5 module rises. This module includes many transcription factors involved in primary root meristem organization whose expression is observed in central and tip LRP zones [START_REF] Du | PLETHORA transcription factors orchestrate de novo organ patterning during Arabidopsis lateral root outgrowth[END_REF][START_REF] Lavenus | Inference of the Arabidopsis lateral root gene regulatory network suggests a bifurcation mechanism that defines primordia flanking and central zones[END_REF].

Interestingly, the inference approach indicated that these two sub-networks were linked by multiple mutual inhibition relationships [START_REF] Lavenus | Inference of the Arabidopsis lateral root gene regulatory network suggests a bifurcation mechanism that defines primordia flanking and central zones[END_REF]. This topology is predicted to confer a toggle switch behaviour to the GRN [START_REF] Gardner | Construction of a genetic toggle switch in Escherichia coli[END_REF][START_REF] Shoval | SnapShot: Network Motifs[END_REF][START_REF] Tyson | Sniffers, buzzers, toggles and blinkers: dynamics of regulatory and signaling pathways in the cell[END_REF]. Altogether, this suggests that a switch between a module controlling early stages of LR development and a second module regulating meristem organisation occurs between 20h and 30h after gravistimulation i.e. at the transition from stage IV to V. Interestingly, this corresponds to the appearance of the organising centre identity in LRPs that marks a major transition in LRP cell divisions and expansion [START_REF] Goh | Quiescent center initiation in the Arabidopsis lateral root primordia is dependent on the SCARECROW transcription factor[END_REF]. Thus, the predicted topology of the network suggests systems-scale mechanisms that contribute to the patterning of LRP. Network analysis further revealed an unexpected role for very long chain fatty acids (VLCFA) acting downstream of the transcription factor PUCHI to control cell proliferation during lateral root formation (Trinh et al, PNAS, in press).

LR transcriptome analysis and meta-analysis approaches also contributed to identify numerous small signalling-peptide encoding genes involved in intercellular communications during root branching [START_REF] Ghorbani | Expanding the repertoire of secretory peptides controlling root development with comparative genome analysis and functional assays[END_REF][START_REF] Oh | Signaling Peptides and Receptors Coordinating Plant Root Development[END_REF]. Pairs of peptides and peptide-receptor complexes were shown to contribute to the spatial patterning of the lateral root initiation by mediating cellto-cell signalling [START_REF] Kumpf | Floral organ abscission peptide IDA and its HAE/HSL2 receptors control cell separation during lateral root emergence[END_REF][START_REF] Fernandez | The GLV6/RGF8/CLEL2 peptide regulates early pericycle divisions during lateral root initiation[END_REF][START_REF] Murphy | RALFL34 regulates formative cell divisions in Arabidopsis pericycle during lateral root initiation[END_REF][START_REF] Roberts | CEP5 and XIP1/CEPR1 regulate lateral root initiation in Arabidopsis[END_REF][START_REF] Gonneau | Receptor Kinase THESEUS1 Is a Rapid Alkalinization Factor 34 Receptor in Arabidopsis[END_REF]. Recently, the secreted peptide TARGET OF LBD SIXTEEN 2/PAMP-INDUCED PEPTIDE-LIKE 3 (TOLS2/PIPL3) and its receptor RLK7 were shown to cooperate in inhibiting LR initiation near pre-existing LRFC in a non-cell autonomous manner, thus controlling minimal spacing between initiated LRs (Figure 3) [START_REF] Toyokura | Lateral Inhibition by a Peptide Hormone-Receptor Cascade during Arabidopsis Lateral Root Founder Cell Formation[END_REF]. Together with the intricate networks controlling hormones distribution and signalling across root tissues, peptide signalling pathways provide a new level of complexity in cell-to-cell communication that can contribute to the overall organisation, especially spatial patterning, in the root branching process [START_REF] Laskowski | Periodic Lateral Root Priming: What Makes It Tick?[END_REF][START_REF] Du | Lateral root formation and the multiple roles of auxin[END_REF].

In summary, network scale analyses revealed interconnectivity, redundancy and multiple feedback loops provide plasticity at a cellular level, confer robustness to LRP development at a multicellular scale, and allow for the integration of plant systemic cues. These network properties are likely to be crucial for the root developmental machinery to adapt to the myriad environment signals that they are exposed to in highly heterogeneous soil environments [START_REF] Morris | Shaping 3D Root System Architecture[END_REF].

Divining Roots in search of moisture

Recent studies have revealed that plant roots employ sophisticated regulatory mechanisms to sense and respond by branching to the availability of water and nutrients. Soil consists of air pockets, stones, nutrient rich and poor patches, whilst the spatial and temporal distribution of moisture also varies. Regulating where a lateral root will form is therefore a critical decision to maximise efficiency of foraging in soil. One of the most important soil resources for plant roots to obtain is water. It has long been known that osmotic stress can slow down lateral root emergence in arabidopsis and that this response is ABA dependent [START_REF] Deak | Osmotic regulation of root system architecture[END_REF]. In roots of barley (Hordeum vulgare cv.) and maize (Zea mays), water deficit caused lateral roots to no longer initiate [64]. A recent article revealed this is controlled by an abscisic acid (ABA)-dependent regulatory mechanism termed xerobranching [START_REF] Orman-Ligeza | The Xerobranching Response Represses Lateral Root Formation When Roots Are Not in Contact with Water[END_REF]. The effect of water deficit could be mimicked by ABA treatment, but mutants defective for key PYR/PYL ABA receptors were resistant to both signals. Additional experiments revealed that ABA accumulated in root tip tissues when exposed to a water deficit, blocking LR initiation possibly by attenuating the oscillatory auxin response network in the basal meristem.

A related adaptive response termed lateral root hydropatterning describes the ability of roots to differentiate between contact with moist soil or air, then trigger preferential branching towards water. Xerobranching (the inhibition of LR initiation by drought stress) and hydropatterning (the preferential LR emergence towards the wettest side of the root) appear to be mechanistically distinct since the latter response is unaffected in ABA mutant backgrounds [START_REF] Orman-Ligeza | The Xerobranching Response Represses Lateral Root Formation When Roots Are Not in Contact with Water[END_REF][START_REF] Bao | Plant roots use a patterning mechanism to position lateral root branches toward available water[END_REF]. Instead, modelling studies led authors to propose a "sense-by-growth" mechanism for hydropatterning in which uptake of external water into the root elongation zone conveyed a means to perceive water availability [START_REF] Bao | Plant roots use a patterning mechanism to position lateral root branches toward available water[END_REF]. Recent research has revealed that water availability controls LR initiation in the elongation zone via post-translational modification of AUXIN RESPONSE FACTOR 7 (ARF7). This key LR regulator encodes a transcription factor that contains four SUMO modification sites. Mutating all 4 ARF7 SUMOylation sites disrupts hydropatterning [START_REF] Orosa-Puente | Root branching toward water involves posttranslational modification of transcription factor ARF7[END_REF]. When roots are exposed to air, ARF7 SUMOylation rapidly increases. SUMOylation of ARF7 enables this transcription factor to recruit the Aux/IAA repressor protein, IAA3. This led to a model where the post-translational modification of ARF7 by SUMOylation on the dry side of a root causes IAA3 recruitment, repressing expression of target genes like LBD16 that are required for LR initiation and hydropatterning (Figure 4) [START_REF] Orosa-Puente | Root branching toward water involves posttranslational modification of transcription factor ARF7[END_REF].

Branching out in search of nitrogen

Absorption of nutrients represents another crucial function of a root system and lateral roots in particular [START_REF] Morris | Shaping 3D Root System Architecture[END_REF]. Nutrients are often dispersed in soil and therefore the ability by roots to sense nutrients is vitally important for efficient resource acquisition. Nitrate is a key nutrients that has a profound effect on lateral root development, whose formation and elongation is induced under mild N stress, but this is impaired during exposure to high N or prolonged N stress (recently reviewed in [START_REF] Sun | Nitrate: A Crucial Signal during Lateral Roots Development[END_REF][69][70]). A recent study reported the importance of C-TERMINALLY ENCODED PEPTIDES (CEPs) in roots experiencing low N, where root steleexpressed CEPs travel to the shoot and accumulate in the leaf phloem, where they bind to two leucine-rich receptor kinases (LRR-RK), CEPR1 and CEPR2 [START_REF] Tabata | Perception of root-derived peptides by shoot LRR-RKs mediates systemic N-demand signaling[END_REF]. This triggers the expression of CEP DOWNSTREAM 1 (CEPD1) and CEPD2, which induce the expression of NITRATE TRANSPORTER2.1 (NRT2.1) in roots exposed to N-rich conditions [START_REF] Ohkubo | Shoot-to-root mobile polypeptides involved in systemic regulation of nitrogen acquisition[END_REF]. This boosts N uptake, but inhibits lateral root growth, as NRT2.1 KO mutants have increased lateral root initiation [START_REF] Little | The putative high-affinity nitrate transporter NRT2.1 represses lateral root initiation in response to nutritional cues[END_REF]. A close family member, NRT1.1, coordinates a multitude of genes involved in nitrogen branching response [START_REF] Alvarez | Systems approach identifies TGA1 and TGA4 transcription factors as important regulatory components of the nitrate response of Arabidopsis thaliana roots[END_REF]. This dual-affinity transporter and transceptor is phosphorylated under low nitrate conditions by CBL-Interacting Protein Kinase 23 (CIPK 23), turning the protein into a high-affinity nitrate carrier [75]. Interestingly, the nonphosphorylable form of NRT1.1 no longer represses lateral root initiation [START_REF] Bouguyon | Multiple mechanisms of nitrate sensing by Arabidopsis nitrate transceptor NRT1.1[END_REF]. Phosphorylation of NRT1.1 negatively effects local auxin build-up in LRP through its ability to co-transport auxin, which is inhibited in high nitrate patches, accounting for the local buildup of auxin, triggering localised emergence of lateral roots (Figure 4) [START_REF] Bouguyon | Multiple mechanisms of nitrate sensing by Arabidopsis nitrate transceptor NRT1.1[END_REF][START_REF] Bouguyon | Nitrate Controls Root Development through Post-Transcriptional Regulation of the NRT1.1/NPF6.3 transporter/sensor[END_REF].

Shining light on lateral root development

Recent studies have also revealed a key role for light on LR development. When roots were illuminated but the shoot kept in the dark, root growth and lateral root emergence was reduced [START_REF] Chen | Shoot-to-Root Mobile Transcription Factor HY5 Coordinates Plant Carbon and Nitrogen Acquisition[END_REF]. Interestingly the bZIP transcription factor ELONGATED HYPOCOTYL5 (HY5) KO allele is incapable of directing root growth in response to exposing the shoot to light. HY5 stability is regulated by the COP1 ubiquitin ligase, which degrades HY5 in the dark [START_REF] Ang | Molecular Interaction between COP1 and HY5 Defines a Regulatory Switch for Light Control of Arabidopsis Development[END_REF].

Accordingly, the cop4-1 mutant showed no difference in root growth in different shoot and root light treatments. Stabilized HY5 in the shoot is transported down to the root via the phloem to induce LR formation [START_REF] Chen | Shoot-to-Root Mobile Transcription Factor HY5 Coordinates Plant Carbon and Nitrogen Acquisition[END_REF]. However, adding Far-red (FR) light could increase HY5 levels to a state in which lateral root primordia development was arrested [START_REF] Van Gelderen | Far-Red Light Detection in the Shoot Regulates Lateral Root Development through the HY5 Transcription Factor[END_REF]. The authors hypothesize this is through the binding of HY5 to the ARF19 promoter, reducing its expression which negatively effects PIN3 and LAX3 auxin transport proteins in the overlaying cortex (Figure 4) [START_REF] Van Gelderen | Far-Red Light Detection in the Shoot Regulates Lateral Root Development through the HY5 Transcription Factor[END_REF]. Hence, light perception in the shoot can very finely regulate the auxin machinery locally in cells overlying LR primordia, to alter organ emergence. Interestingly, HY5 in shoots promotes carbon assimilation and in roots can induce expression of NRT2.1 [START_REF] Chen | Shoot-to-Root Mobile Transcription Factor HY5 Coordinates Plant Carbon and Nitrogen Acquisition[END_REF] whilst negatively regulating AMT1;2 [START_REF] Huang | HY5 regulates nitrite reductase 1 (NIR1) and ammonium transporter1;2 (AMT1;2) in Arabidopsis seedlings[END_REF], which increases nitrate and reduces ammonium uptake, respectively. Collectively, this reveals novel root functions for HY5 and flags it as a key node in an array of plant environmental responses.

Concluding Remarks and Future Directions

Major advances have been made elucidating the molecular and cellular basis of lateral root development using the model plant arabidopsis. Nevertheless, lateral root morphogenesis in other plant species like maize and rice exhibits far greater anatomical variation than observed in arabidopsis, reflecting the diversity of root classes [START_REF] Passot | A New Phenotyping Pipeline Reveals Three Types of Lateral Roots and a Random Branching Pattern in Two Cereals[END_REF]. However, very little is currently known about the underlying gene regulatory networks and mechanisms driving the morphological diversity of these different LR classes in crops. Despite this, genetic studies in crop models like maize and rice have identified common hormone signals and genes with arabidopsis that regulate lateral root development [START_REF] Hochholdinger | Genetic Control of Root System Development in Maize[END_REF]84]. The rich 'omic' resources available for arabidopsis has helped initiate efforts to elucidate the underlying gene regulatory networks that control morphogenesis of the new root organs.

GRN approaches have revealed system properties that confer important features in LRP development: ability to stabilize or, conversely, trigger changes in cell identity, capacity to organize those changes in time and space (patterning), capacity to reach a similar output from various transcriptomics scenarios (robustness) and at the same time, capacity to integrate external cues that may influence the GRN final state. This latter plasticity property plays a prominent role for root system adaptation to environmental signals. This will also provide the foundation on which to probe the sophisticated adaptive mechanisms that regulate how plants pattern root branching to match spatial availability of resources like water and nutrients in their external environment.

In parallel with -omics data collection and studies, advances in cell tracking, sampling and transcriptome analysis will generate a more detailed cartography of gene expression in developing LRP at various stages. When combined with time-course transcriptomic datasets, this will allow exploration of GRN at high temporal and spatial resolution. Moreover, crossreferencing with other omics datasets, such as epigenomics, or phosphoproteomics, will help decipher the complex regulation levels resulting in the observed transcriptomic profiles and properties in LRP development [START_REF] Zhang | Quantitative Phosphoproteomics after Auxin-stimulated Lateral Root Induction Identifies an SNX1 Protein Phosphorylation Site Required for Growth[END_REF]. Further studies will be needed in arabidopsis to explore and characterise the emerging properties of the lateral root GRN and how it integrates environmental signals, and how it compares to mechanisms in other plant species (see Outstanding Questions). by the nuclear migration of two pericycle cells and their first anticlinal division in the differentiation zone. During morphogenesis the LRFCs divide periclinally as well as radially to form a three/dimensional lateral root primordia (LRP). The LRP grows through the overlaying tissue layers to emerge from the primary root. After emergence the lateral root will acquire an active meristem. Colour coding was used according to the model described by Malamy and Benfey [START_REF] Malamy | Organization and cell differentiation in lateral roots of Arabidopsis thaliana[END_REF]. Six files of pericycle cell files contribute to the LRP seen in a radial perspective (B) [START_REF] Wangenheim | Rules and self-organizing properties of post-embryonic plant organ cell division patterns[END_REF]. Common colours (yellow or dark yellow) highlight daughter cells derived from individual pericycle founder cell files. ARF7, ARF19 module control PLT-dependant responses through SHR/SCR turnover to control LR patterning [START_REF] Du | PLETHORA transcription factors orchestrate de novo organ patterning during Arabidopsis lateral root outgrowth[END_REF]. TOLS2 peptide interact with RLK7 to control PUCHI and LR spacing [START_REF] Toyokura | Lateral Inhibition by a Peptide Hormone-Receptor Cascade during Arabidopsis Lateral Root Founder Cell Formation[END_REF]. A plant-specific B3 transcription factors FUS3 and LEC2 interact together to induce expression of the auxin biosynthetic gene YUC4 through binding to its promoter elements in LRFCs [START_REF] Tang | FUSCA3 interacting with LEAFY COTYLEDON2 controls lateral root formation through regulating YUCCA4 gene expression in Arabidopsis thaliana[END_REF]. The phloem pole-expressed CEP5 and its proposed leucine-rich repeat (LRR) receptor XIP1 as well as another peptide GOLVEN 6 (GLV6), might also be involved in the LRI [START_REF] Fernandez | The GLV6/RGF8/CLEL2 peptide regulates early pericycle divisions during lateral root initiation[END_REF] [START_REF] Ang | Molecular Interaction between COP1 and HY5 Defines a Regulatory Switch for Light Control of Arabidopsis Development[END_REF]. However in WT+FR light, COP1 is broken down through stabilized PhyA mediated degradation and subsequently HY5 accumulates [START_REF] Van Gelderen | Far-Red Light Detection in the Shoot Regulates Lateral Root Development through the HY5 Transcription Factor[END_REF]. HY5 is transported through phloem to inactivate LRP primordia development through inducing its own local expression and indirectly reducing LAX3 and PIN3 in the overlaying tissue layers, possibly through ARF19 interaction. In patched of low nitrogen (N) soil, NTR1.1 becomes phosphorylated by CIPK23. Phosphorylation causes NRT1.1 to perform a dual function as nitrogen and auxin transporter, which blocks auxin accumulation in the lateral root primordia (LRP). In high N soil, NRT1.1 is unphosphorylated, blocking its auxin transport function and allowing auxin to build up in the LRP to promote emergence [START_REF] Alvarez | Systems approach identifies TGA1 and TGA4 transcription factors as important regulatory components of the nitrate response of Arabidopsis thaliana roots[END_REF][75][START_REF] Bouguyon | Multiple mechanisms of nitrate sensing by Arabidopsis nitrate transceptor NRT1.1[END_REF][START_REF] Bouguyon | Nitrate Controls Root Development through Post-Transcriptional Regulation of the NRT1.1/NPF6.3 transporter/sensor[END_REF]. During periods of root growth through air gaps, xerobranching, abscisic acid (ABA) accumulates and binds to its receptor family PYR/PYL to indirectly reduce IAA levels and therefore lateral root founder cell (LRFC) initiation. However, when one side is in contact with water in the soil, ARF7 induces downstream targets such as LBD16 to initiate LRFC division on the side in contact with water, termed lateral root hydropatterning [START_REF] Orosa-Puente | Root branching toward water involves posttranslational modification of transcription factor ARF7[END_REF]. Contrary, on the air side ARF7 is SUMOylated causing recruitment of IAA3 repressor proteins resulting in ARF7 protein degradation and inhibition of LR formation.

  Lateral roots are crucial for increasing surface area of root systems to explore heterogeneous soil environments. Major advances have recently been made in the model plant Arabidopsis thaliana to elucidate the cellular basis of lateral root development and the underlying gene regulatory networks that control morphogenesis of the new root organ. This has provided the foundation on which to understand the sophisticated adaptive mechanisms that regulate how plants pattern their root branching to match the spatial availability of resources like water and nutrients in their external environment. We review new insights into the molecular, cellular and environmental regulation of LR development in arabidopsis.

  Surprisingly, limited attention has been paid to the role of biomechanics in the regulation of lateral root development, to date. However, increasing evidence points to the central role for mechanical signals during development in both plants and animals [21,85]. Lateral root development is a fascinating system to address these questions given that, unlike leaf primordia formation that takes place on the flanks of the shoot apical meristem, new root organs originate deep within parental root tissues. Live cell imaging, e.g. light sheet microscopy, will further advance the field in observing the dynamics of lateral root morphogenesis in real-time. Such research could generate fundamentally new knowledge about mechanical patterning and signalling in plants. For example, uncovering how LRP QC specification is synchronised with the new organ disrupting the Casparian strip could reveal novel mechanical signalling components and mechanisms linking cell walls and cell cycle regulation.

Figure 1 .

 1 Figure 1. The birth of a new lateral root. The five main steps of lateral root formation in the

Figure 2 .

 2 Figure 2. The lateral root primordia (LRP) must break through the overlaying endodermis

Figure 3 .

 3 Figure 3. Feeding GRN with spatial and temporal information result in robust spacing and

  . Abbreviations: A-type CYCLIN-Dependent KINASE A1.1 (CDKA1); AUXIN-RESPONSE FACTOR 7, 19 (ARF7, ARF19); C-TERMINALLY ENCODED PEPTIDE5 (CEP5); CYCLINB1.1 (CYCB1); FOURLIPS (FLP); FUSCA3 (FUS3); LATERAL ORGAN BOUNDARIES 16, 18 (LBD16, LBD18; LEAFY COTYLEDON2 (LEC2); RECEPTOR-Like KINASE 7 (RLK7); SOLITARY-ROOT (SLR); TEOSINTE-branched CYCLOIDEA PCNA (TCP); TARGET OF LBD SIXTEEN 2 (TOLS2); WUSCHEL-Related HOMEOBOX 5 (WOX5); XYLEM INTERMIXED WITH PHLOEM 1 (XIP1); YUCCA4 (YUC4).

Figure 4 .

 4 Figure 4. Soil and light environment play a key role in lateral root (LR) positioning. White

Table 1 .

 1 Overview of mutant and transgenic lines affected in lateral root development.

	Category Mutants	Paper	Lateral root phenotype	Stage affected
	Auxin Signalling 35S:TOLS2	[60]	Reduced number of LRs	LR initiation
		puchi	[89]	Reduced number of LRs	LRP development/ emergence
		CASP::shy2-2	[25]	No LRs	LR initiation
	Auxin transport aux1	[90]	Reduced number of LRs	LR initiation
		lax3	[22]	Reduced number of LRs	Stage I development
	Auxin conjugation/	dao1	[91]	Increased number of LRs	?
	degradation	gh3.1,2,3,4,5,6	[91]	Increased number of LRs	?
	Auxin Biosynthesis yuc4	[88]	Reduced number of LRs	?
	Meristem	plt3plt5plt7	[39]	Reduced emergence	Promotes initiation, reduces
	establishment				emergence
	Transcription	flp	[92]	Reduced number of LRs	Delay in stage I
	factors
		fus3	[88]	Reduced number of LRs	Delayed emergence
		lec2	[88]	Reduced number of LRs	?
	Peptides ralf34	[57]	Increased number of LRs	initiation
		irAtRALF1	[93]	Increased number of LRs	?
		ida	[24]	Decreased LR density	Delayed emergence
		CEP5	[58]	Decreased LR density	stage I and II
	Receptors xip1	[58]	Decreased LR density	initiation and development
		the1	[59]	Increase number of stage I LRP	patterning and development of LRP
		GLV 1-11 OE	[94]	Reduced LR density	initiation and development
	Kinases aur1-2 aur2-2	[95]	Decreased LR density	Decreased initiation and emergence
		hae hsl2	[24]	Decreased LR density	Delayed emergence
		mkk4/mkk5	[96]	Reduced LR density	Delayed emergence
		MPK3SR and MPK6SR	[96]	Reduced LR density	Delayed emergence
	Cell wall expa1	[28]	Reduced number of LRs	Initiation
		gpat4 gpat8	[97]	Reduced number of LRs	Delayed emergence and deformation
		dcr	[97]	Reduced number of LRs	Delayed emergence and deformation
		bdg	[97]	Reduced number of LRs	Delayed emergence and deformation
		lrd5/xeg113	[31]	Increase rate of emergence	Emergence
	Oxygen and ROS rap2.12,rap2.2,rap2.3,h	[98]	Increased number of LRs	Delayed emergence
		re1,hre2
		erf VII	[98]	Increased number of LRs	Initiation
		robhc rbohd rbohe	[35]	Decreased LR density	Delayed emergence
		myb36	[37]	Decreased LR density	Delay in stage IV
		per7	[36]	Decreased LR density	Initiation
		per57	[36]	Decreased LR density	Initiation
	Water transport pdbg1,2	[26]	Increased LR density	Patterning and spacing
		pip2;1-1 and pip2;1-2	[23]	Reduced number of LRs	Delayed development
		tip1;1 tip2;1 tip2;1	[33]	Reduced number of LRs	Delayed emergence
	Light sensing hyh/hy5	[80]	Increased number of LRs	Initiation and emergence
	SUMOylation ots1 ots2	[67]	Reduced number of LRs	?
		siz1	[99]	Reduced number of LRs	?
	Circadian clock toc1-1	[46]	Decreased LR density	Delayed initiation and emergence

Acknowledgement

The authors would like to acknowledge the following funding agencies for financial support: J.B, D.v.W., T.G. and M.J.B. acknowledge the Biological and Biotechnological Science Research Council (BBSRC) for supporting Doctoral Training, responsive mode and Professorial Fellowship awards (grant numbers BB/G023972/1, BB/R013748/1, BB/L026848/1, BB/M018431/1, BB/PO16855/1, BB/M001806/1); K.B., S.G. and L.L. acknowledge support from the French National Research Agency (ANR) through the NewRoot project (ANR-17-CE13-0004-01); T.G. was supported by the Ministry of Education, Culture, Sports, Science and Technology (MEXT) through a JSPS Research Fellowship for Young Scientists (No. 12J02079)