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The anaphase-promoting complex/cyclosome (APC/C) represents a large multisubunit E3-ubiquitin ligase complex that controls the unidirectional progression through the cell cycle by the ubiquitination of specific target proteins, marking them for proteasomal destruction. Although the APC/C's role is largely conserved among eukaryotes, its subunit composition and target spectrum appear to be species specific. In this review, we focus on the plant APC/C complex, whose activity correlates with different developmental processes, including polyploidization and gametogenesis. After an introduction into proteolytic control by ubiquitination, we discuss the composition of the plant APC/C and the essential nature of its core subunits for plant development. Subsequently, we describe the APC/C activator subunits and interactors, most being plant specific. Finally, we provide a comprehensive list of confirmed and suspected plant APC/C target proteins. Identification of growth-related targets might offer opportunities to increase crop yield and resilience of plants to climate change by manipulating APC/C activity.

The Anaphase Promoting Complex/Cyclosome (APC/C) controls unidirectional progression through the cell cycle by marking key cell cycle proteins for proteasomal turnover. Its activity is temporally regulated by the docking of different activating subunits, known in plants as CDC20 and CCS52. Despite the importance of the APC/C during cell proliferation, the number of identified targets in the plant cell cycle is limited. Here, we used the growth and meristem phenotypes of Arabidopsis CCS52A2-deficient plants in a suppressor mutagenesis screen to identify APC/CCCS52A2 substrates or regulators, resulting in the identification of a mutant cyclin CYCA3;4 allele. CYCA3;4 deficiency partially rescues the early ccs52a2-1 phenotypes, whereas increased CYCA3;4 levels enhances them. Furthermore, whereas CYCA3;4 proteins are promptly broken down after prophase in wild-type plants, they remain present in later stages of mitosis in ccs52a2-1 mutant plants, marking them as APC/CCCS52A2 substrates. Strikingly, CYCA3;4 overexpression results in aberrant root meristem and stomatal divisions, mimicking phenotypes of plants with reduced RBR1 activity. Correspondingly, RBR1 hyperphosphorylation was observed in CYCA3;4-overproducing plants. Our data thus demonstrate that an inability to timely destroy CYCA3;4 attributes to disorganized formative divisions, likely in part caused by the inactivation of RBR1.ONE-SENTENCE SUMMARYTimely post-prophase breakdown of the Arabidopsis cyclin CYCA3;4 by the Anaphase Promoting Complex/Cyclosome is essential for meristem organization and development.

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International audience; Root-knot nematodes (RKN) infect a large number of plant species triggering the formation of galls by inducing vascular cells to reenter the cell cycle. These nematodes infect the host root and usurp the plant cell cycle machinery by inducing multinucleate giant-feeding cells surrounded by asymmetrically dividing neighboring cells. Cell cycle activation is a basis for gall induction and maintenance. Giant cell development is characterized by an acytokinetic mitotic phase, followed by an endoreduplication stage leading to polyploidization of nuclei during cell expansion. The endocycle involves genome duplication without cell division, resulting in polyploidy. E2Fa and E2Fb transcription factors are positive regulators of the mitotic as well the endocycle being essential to drive S phase. Via functional and molecular analysis we unraveled the role of AtE2Fa and AtE2Fb for gall growth and development. In situ transcript localization has shown that AtE2Fa and AtE2Fb are highly expressed in these nematode feeding sites. A detailed analysis of E2F knockout and overexpressing lines suggests that despite both genes seem to have slightly different timing and spatial localization during gall expansion, both are essential for gall development.

Diatoms are unicellular algae with a fundamental role in global biogeochemical cycles as major primary producers at the base of aquatic food webs. In recent years, chemical communication between diatoms and associated bacteria has emerged as a key factor in diatom ecology, spurred by conceptual and technological advancements to study the mechanisms underlying these interactions. Here, we use a combination of physiological, transcriptomic, and metabolomic approaches to study the influence of naturally co-existing bacteria, Maribacter sp. and Roseovarius sp., on the sexual reproduction of the biofilm inhabiting marine pennate diatom Seminavis robusta. While Maribacter sp. severely reduces the reproductive success of S. robusta cultures, Roseovarius sp. slightly enhances it. Contrary to our expectation, we demonstrate that the effect of the bacterial exudates is not caused by altered cell-cycle regulation prior to the switch to meiosis. Instead, Maribacter sp. exudates cause a reduced production of diproline, the sexual attraction pheromone of S. robusta. Transcriptomic analyses show that this is likely an indirect consequence of altered intracellular metabolic fluxes in the diatom, especially those related to amino acid biosynthesis, oxidative stress response, and biosynthesis of defense molecules. This study provides the first insights into the influence of bacteria on diatom sexual reproduction and adds a new dimension to the complexity of a still understudied phenomenon in natural diatom populations.

Plants can acclimate by using tropisms to link the direction of growth to environmental conditions. Hydrotropism allows roots to forage for water, a process known to depend on abscisic acid (ABA) but whose molecular and cellular basis remains unclear. Here, we show that hydrotropism still occurs in roots after laser ablation removed the meristem and root cap. Additionally, targeted expression studies reveal that hydrotropism depends on the ABA signalling kinase, SnRK2.2, and the hydrotropism-specific MIZ1, both acting specifically in elongation zone cortical cells. Conversely, hydrotropism, but not gravitropism, is inhibited by preventing differential cell-length increases in the cortex, but not in other cell types. We conclude that root tropic responses to gravity and water are driven by distinct tissue-based mechanisms. In addition, unlike its role in root gravitropism, the elongation zone performs a dual function during a hydrotropic response, both sensing a water potential gradient and subsequently undergoing differential growth.

The stem cell niche of the Arabidopsis (Arabidopsis thaliana) primary root apical meristem is composed of the quiescent (or organizing) center surrounded by stem (initial) cells for the different tissues. Initial cells generate a population of transit-amplifying cells that undergo a limited number of cell divisions before elongating and differentiating. It is unclear whether these divisions occur stochastically or in an orderly manner. Using the thymidine analog 5-ethynyl-2'-deoxyuridine to monitor DNA replication of cells of Arabidopsis root meristems, we identified a pattern of two, four, and eight neighboring cells with synchronized replication along the cortical, epidermal, and endodermal cell files, suggested to be daughters, granddaughters, and great-granddaughters of the direct progeny of each stem cell. Markers of mitosis and cytokinesis were not present in the region closest to the transition zone where the cells start to elongate, suggesting that great-granddaughter cells switch synchronously from the mitotic cell cycle to endoreduplication. Mutations in the stem cell niche-expressed ASH1-RELATED3 (ASHR3) gene, encoding a SET-domain protein conferring histone H3 lysine-36 methylation, disrupted this pattern of coordinated DNA replication and cell division and increased the cell division rate in the quiescent center. E2Fa/E2Fb transcription factors controlling the G1-to-S-phase transition regulate ASHR3 expression and bind to the ASHR3 promoter, substantiating a role for ASHR3 in cell division control. The reduced length of the root apical meristem and primary root of the mutant ashr3-1 indicate that synchronization of replication and cell divisions is required for normal root growth and development.

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Background: At meiosis, two successive rounds of chromosome segregation lead to ploidy halving. This is achieved through a stepwise release of sister chromatid cohesion, along chromosome arms to allow homolog segregation at anaphase I and at centromeres to allow sister chromatid segregation at anaphase II. Cohesins, the protein complex that ensures cohesion, must then be protected at centromeres throughout meiosis, until the onset of anaphase II. Members of the Shugoshin protein family have been shown to protect centromeric cohesins at anaphase I, but much less is known about the protection of cohesion during interkinesis, the stage between meiosis I and meiosis II. [br/] Results: Here, we (1) show that both Arabidopsis SHUGOSHINs paralogs are required for complete protection of centromeric cohesins during meiosis I, without. apparent somatic function, and (2) identified PATRONUS (PANS1), a novel protein required for protection of meiotic centromeric cohesion. Although AtSGO1 and AtSGO2 protect centromeric cohesion during anaphase I, PANS1 is required at a later stage, during interkinesis. Additionally, we identified PANS2, a paralog of PANS1, whose mutation is synthetically lethal with pans1 suggesting that PANS genes are also essential for mitosis. PANS1 interacts directly with the CDC27b and the CDC20.1 subunit of the Anaphase Promoting Complex (APC/C), in a manner suggesting that PANS1 could be both a regulator and a target of the APC/C. [br/] Conclusions: This study reveals that centromeric cohesion is actively protected at two successive stages of meiosis, by SHUGOSHINs at anaphase I and by PATRONUS at interkinesis.

International audience; The establishment of galls and syncytia as feeding sites induced by root-knot and cyst nematodes, respectively, involves a progressive increase in nuclear and cellular size. Here we describe the functional characterization of endocycle activators CCS52A, CCS52B and a repressor of the endocycle, DEL1, during two types of nematode feeding site development in Arabidopsis thaliana. In situ hybridization analysis showed that expression of CCS52A1 and CCS52B was strongly induced in galls and syncytia and DEL1 was stably but weakly expressed throughout feeding site development. Down-regulation and over-expression of CCS52 and DEL1 in Arabidopsis drastically affected giant cell and syncytium growth, resulting in restrained nematode development, illustrating the need for mitotic activity and endo-reduplication for feeding site maturation. Exploiting the mechanism of endo-reduplication may be envisaged as a strategy to control plant-parasitic nematodes.

In multicellular organisms, morphogenesis relies on a strict coordination in time and space of cell proliferation and differentiation. In contrast to animals, plant development displays continuous organ formation and adaptive growth responses during their lifespan relying on a tight coordination of cell proliferation. How developmental signals interact with the plant cell-cycle machinery is largely unknown. Here, we characterize plant A2-type cyclins, a small gene family of mitotic cyclins, and show how they contribute to the fine-tuning of local proliferation during plant development. Moreover, the timely repression of CYCA2;3 expression in newly formed guard cells is shown to require the stomatal transcription factors FOUR LIPS/MYB124 and MYB88, providing a direct link between developmental programming and cell-cycle exit in plants. Thus, transcriptional downregulation of CYCA2s represents a critical mechanism to coordinate proliferation during plant development.

Analysis of gene expression data generated by high-throughput microarray transcript profiling experiments has demonstrated that genes with an overall similar expression pattern are often enriched for similar functions. This guilt-by-association principle can be applied to define modular gene programs, identify cis-regulatory elements, or predict gene functions for unknown genes based on their coexpression neighborhood. We evaluated the potential to use Gene Ontology (GO) enrichment of a gene's coexpression neighborhood as a tool to predict its function but found overall low sensitivity scores (13%-34%). This indicates that for many functional categories, coexpression alone performs poorly to infer known biological gene functions. However, integration of cis-regulatory elements shows that 46% of the gene coexpression neighborhoods are enriched for one or more motifs, providing a valuable complementary source to functionally annotate genes. Through the integration of coexpression data, GO annotations, and a set of known cis-regulatory elements combined with a novel set of evolutionarily conserved plant motifs, we could link many genes and motifs to specific biological functions. Application of our coexpression framework extended with cis-regulatory element analysis on transcriptome data from the cell cycle-related transcription factor OBP1 yielded several coexpressed modules associated with specific cis-regulatory elements. Moreover, our analysis strongly suggests a feed-forward regulatory interaction between OBP1 and the E2F pathway. The ATCOECIS resource (http://bioinformatics.psb.ugent.be/ATCOECIS/) makes it possible to query coexpression data and GO and cis-regulatory element annotations and to submit user-defined gene sets for motif analysis, providing an access point to unravel the regulatory code underlying transcriptional control in Arabidopsis (Arabidopsis thaliana).

The phytopathogenic actinomycete Rhodococcus fascians induces neoplastic shooty outgrowths on infected hosts. Upon R. fascians infection of Arabidopsis (Arabidopsis thaliana), leaves are formed with small narrow lamina and serrated margins. These symptomatic leaves exhibit reduced tissue differentiation, display more but smaller cells that do not endoreduplicate, and accumulate in the G1 phase of the cell cycle. Together, these features imply that leaf growth occurs primarily through mitotic cell division and not via cell expansion. Molecular analysis revealed that cell cycle gene expression is activated continuously throughout symptomatic leaf development, ensuring persistent mitotic cycling and inhibition of cell cycle exit. The transition at the two major cell cycle checkpoints is stimulated as a direct consequence of the R. fascians signals. The extremely reduced phenotypical response of a cyclind3;1-3 triple knockout mutant indicates that the D-type cyclin/retinoblastoma/E2F transcription factor pathway, as a major mediator of cell growth and cell cycle progression, plays a key role in symptom development and is instrumental for the sustained G1-to-S and G2-to-M transitions during symptomatic leaf growth.

Prenylated Rab acceptor 1 (PRA1) domain proteins are small transmembrane proteins that regulate vesicle trafficking as receptors of Rab GTPases and the vacuolar soluble N-ethylmaleimide-sensitive factor attachment receptor protein VAMP2. However, little is known about PRA1 family members in plants. Sequence analysis revealed that higher plants, compared with animals and primitive plants, possess an expanded family of PRA1 domain-containing proteins. The Arabidopsis (Arabidopsis thaliana) PRA1 (AtPRA1) proteins were found to homodimerize and heterodimerize in a manner corresponding to their phylogenetic distribution. Different AtPRA1 family members displayed distinct expression patterns, with a preference for vascular cells and expanding or developing tissues. AtPRA1 genes were significantly coexpressed with Rab GTPases and genes encoding vesicle transport proteins, suggesting an involvement in the vesicle trafficking process similar to that of their animal counterparts. Correspondingly, AtPRA1 proteins were localized in the endoplasmic reticulum, Golgi apparatus, and endosomes/prevacuolar compartments, hinting at a function in both secretory and endocytic intracellular trafficking pathways. Taken together, our data reveal a high functional diversity of AtPRA1 proteins, probably dealing with the various demands of the complex trafficking system.

Kip-related proteins (KRPs) play a major role in the regulation of the plant cell cycle. We report the identification of five putative rice (Oryza sativa) proteins that share characteristic motifs with previously described plant KRPs. To investigate the function of KRPs in rice development, we generated transgenic plants overexpressing the Orysa;KRP1 gene. Phenotypic analysis revealed that overexpressed KRP1 reduced cell production during leaf development. The reduced cell production in the leaf meristem was partly compensated by an increased cell size, demonstrating the existence of a compensatory mechanism in monocot species by which growth rate is less reduced than cell production, through cell expansion. Furthermore, Orysa;KRP1 overexpression dramatically reduced seed filling. Sectioning through the overexpressed KRP1 seeds showed that KRP overproduction disturbed the production of endosperm cells. The decrease in the number of fully formed seeds was accompanied by a drop in the endoreduplication of endosperm cells, pointing toward a role of KRP1 in connecting endocycle with endosperm development. Also, spatial and temporal transcript detection in developing seeds suggests that Orysa;KRP1 plays an important role in the exit from the mitotic cell cycle during rice grain formation.

Entry into the S phase of the cell cycle is controlled by E2F transcription factors that induce the transcription of genes required for cell cycle progression and DNA replication. Although the E2F pathway is highly conserved in higher eukaryotes, only a few E2F target genes have been experimentally validated in plants. We have combined microarray analysis and bioinformatics tools to identify plant E2F-responsive genes. Promoter regions of genes that were induced at the transcriptional level in Arabidopsis (Arabidopsis thaliana) seedlings ectopically expressing genes for the E2Fa and DPa transcription factors were searched for the presence of E2F-binding sites, resulting in the identification of 181 putative E2F target genes. In most cases, the E2F-binding element was located close to the transcription start site, but occasionally could also be localized in the 5' untranslated region. Comparison of our results with available microarray data sets from synchronized cell suspensions revealed that the E2F target genes were expressed almost exclusively during G1 and S phases and activated upon reentry of quiescent cells into the cell cycle. To test the robustness of the data for the Arabidopsis E2F target genes, we also searched for the presence of E2F-cis-acting elements in the promoters of the putative orthologous rice (Oryza sativa) genes. Using this approach, we identified 70 potential conserved plant E2F target genes. These genes encode proteins involved in cell cycle regulation, DNA replication, and chromatin dynamics. In addition, we identified several genes for potentially novel S phase regulatory proteins.

The dual-specificity CDC25 phosphatases are critical positive regulators of cyclin-dependent kinases (CDKs). Even though an antagonistic Arabidopsis thaliana WEE1 kinase has been cloned and tyrosine phosphorylation of its CDKs has been demonstrated, no valid candidate for a CDC25 protein has been reported in higher plants. We identify a CDC25-related protein (Arath;CDC25) of A. thaliana , constituted by a sole catalytic domain. The protein has a tyrosine-phosphatase activity and stimulates the kinase activity of Arabidopsis CDKs. Its tertiary structure was obtained by NMR spectroscopy and confirms that Arath;CDC25 belongs structurally to the classical CDC25 superfamily with a central five-stranded β-sheet surrounded by helices. A particular feature of the protein, however, is the presence of an additional zinc-binding loop in the C-terminal part. NMR mapping studies revealed the interaction with phosphorylated peptidic models derived from the conserved CDK loop containing the phosphothreonine-14 and phosphotyrosine-15. We conclude that despite sequence divergence, Arath;CDC25 is structurally and functionally an isoform of the CDC25 superfamily, which is conserved in yeast and in plants, including Arabidopsis and rice.

Three decades have passed since the first recognition of restriction checkpoints in the plant cell cycle. Although many core cell cycle genes have been cloned, the mechanisms that control the G(1)-->S and G(2)-->M transitions in plants have only recently started to be understood. The cyclin-dependent kinases (CDKs) play a central role in the regulation of the cell cycle, and the activity of these kinases is steered by regulatory subunits, the cyclins. The activities of CDK-cyclin complexes are further controlled by an intricate panoply of monitoring mechanisms, which result in oscillating CDK activity during the division cycle. These fluctuations trigger transitions between the different stages of the cell cycle.

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E2F/DP complexes play a pivotal role in the regulation of the G1/S transition in animals. Recently, plant E2F homologs have been cloned, but DP-related sequences have not been identified so far. Here we report that Arabidopsis thaliana contains at least two different DP-related genes, AtDPa and AtDPb. They exhibit an overall domain organization similar to that of their animal counterparts, although phylogenetic analysis demonstrated that they form a separate subgroup. AtDPs efficiently heterodimerize in vitro with the Arabidopsis E2F-related proteins, AtE2Fa and AtE2Fb through their dimerization domains. AtDPa and AtE2Fa are predominantly produced in actively dividing cells with highest transcript levels in early S phase cells.

In this thesis, we studied a mathematical modelling approach of systems biology in plants. We have concentrated on two different issues related to the cell cycle and cell division (especially in the plant Arabidopsis). The first issue is that of the epidermal cell population in the Arabidopsis leaf and the second issue deals with gene networks which play an important role during the cell cycle. The chapters are grouped into four parts. In Part I, we described the cell cycle as the series of events that takes place in a cell leading to its division and duplication. We also stated the general objective of the study. We addressed the various aspects of the problems and the key factors that are assumed to influence or cause the problems. We provided a comprehensive mathematical framework in Part II to be used in the other chapters for the modelling, simulation and analysing purposes. Here we introduced and studied Michaelis-Menten kinetics (a model of enzyme kinetics) and the quasi-steady state assumption to reduce the complexity of the model. We also introduced two basic mathematical models for the growth of cell size in plants. In Part III, we considered two case studies related to the cell cycle and cell division in Arabidopsis. The first case study is the temporal control of epidermal cell divisions in the Arabidopsis leaf. The growth of plant organs is the result of two processes acting on the cellular level, namely cell division and cell expansion. The precise nature of the interaction between these two processes is still largely unknown as it is experimentally challenging to disentangle them. The lower epidermal tissue layer of the Arabidopsis leaf is composed of two cell types, puzzle shaped pavement cells and guard cells, which build the stomata. We determined the cell number and the individual cell areas separately for both cell types during development. To dissect the rules whereby different cell types divide and expand, the experimental data were fit into a computational model that describes all possible changes a cell can undergo from a given day to the next day. The model allows to calculate the probabilities for a precursor cell to become a guard or pavement cell, the maximum size at which it can divide into two pavement cells or two guard cells, the cell cycle duration and two different growth rates for two kinds of cells (pavement and guard cells) in one population. The second case study deals with the fact that atypical E2F activity restrains APC/CCCS52A2 function obligatory for endocycle onset. We have demonstrated that the atypical E2F transcription factor E2Fe/DEL1 controls the onset of the endocycle through a direct transcriptional control of APC/C activity. Because E2Fe/DEL1 represses the CCS52A2 promoter, we hypothesize that its level must drop below a critical threshold to allow sufficient accumulation of CCS52A2 during late S and G2 phase for cells to proceed from division to endoreduplication. We built a mathematical model to analyse the above hypothesis. The model is based on an ODE model used for the binding of ligands to proteins with the help of Hill functions. This mathematical model helps to understand mechanistically how decreasing E2Fe/DEL1 levels can account for the division-to-endoreduplication transition. Finally, in Part IV, we discussed some future work to extend the above research. We suggested several ideas to design new experiments and increase the value of the models. The term ”we” is used throughout the text to underline the fact that every single result of the author’s work as represented in the thesis was only possible because of the provision of experiments, equipment, materials and scientific input from others.

The WEE1 kinase is a conserved cell cycle inhibitor that, together with CDC25, controls CDK activity and consequently the G2/M progression in yeast and animals. The same control mechanism is utilized by the DNA damage checkpoint to induce a cell cycle arrest upon DNA stress. In plants, a homolog of the WEE1 gene is present and is also responsible for phosphorylation of CDKs, although only during DNA damage. Plants that not contain a functional WEE1 gene develop normally during normal growth circumstances, but show a hypersensitive growth response to replication stress. To understand this, a transcriptomics analysis was performed, indicating prolonged S-phase duration. A role for WEE1 during S phase was substantiated by its specific accumulation in replicating nuclei that suffered from DNA stress. Besides an extended replication phase, WEE1 knockout plants accumulated dead cells that were associated with the onset of premature vascular differentiation. Furthermore, a new synchronization method for Arabidopsis roots was presented and the initial research on the posttranslational control of plant WEE1 was executed, both providing intriguing results.

Accounting for almost one fifth of the primary production on Earth, diatoms play a key ecological and biogeochemical role in our contemporary oceans. Furthermore, as producers of various lipids and pigments, and characterized by their finely ornamented silica cell wall, diatoms gained an emerging interest of different industrial fields, including biofuel production, nanotechnology and pharmaceutics. However, despite their major ecological importance and their high commercial value, little is known about the mechanisms that control their life cycle. Their ability to live and dominate in highly unstable and sometimes harsh environments, suggests that diatoms have evolved specific strategies to adapt to and survive in such conditions. Unraveling the regulatory mechanisms that underlie their unique life cycle strategies will therefore be of crucial importance to understand diatom ecology and evolution and to further exploit their industrial potential. In this thesis I aimed to gain insights in the genetic mechanisms and environmental factors that control the diatom cell cycle. Using a variety of molecular analyses, it is demonstrated that diatoms use a common eukaryotic base of cell cycle regulatory components to control their cell division, complemented with some novel diatom-specific features, including an expanded set of diatom-specific cyclins, which most probably are part of a complex integrative network allowing them to pace the cell cycle with the surrounding conditions.

In the recent years many studies contributed to our understanding of the mechanisms that drives the cell cycle machinery. Studies performed in yeast, animals, worms, flies and plants revealed that despite the evolutionary distance between these species, an universal picture can be drawn on how the basic cell cycle machinery is regulated. However, in spite of their highly conserved cell cycle machinery, it is remarkable how plants and animals have integrated the control of cell cycle differently into their specific developmental programs. In contrast to animals, plants develop mostly post-embryonic, which is characterized by continuous growth and organ formation during their entire life-span. This developmental style relies on the existence of stem cell niches within the root and shoot apical meristems, continuously supplying new cells. Next to this, plants are able to form organs de novo, like lateral roots, requiring cell cycle reactivation within already differentiated cells. Additionally, besides cell proliferation, endoreduplication plays as well an important role during different developmental processes. The plant body shows an amazing flexibility rendering them the ability to cope with different environmental and developmental signals. However, although different reports describe the influence of different environmental and developmental cues on cell cycle progression and endoreduplication, we have currently only limited knowledge on how these signals connect to the core cell cycle machinery. The aim of this project was to gain insight in how these intrinsic and extrinsic signals are integrated with the regulation of the cell cycle machinery. Although different core cell cycle genes display developmental and cell cycle-phase dependent transcriptional regulation, it is intriguing how little is known on their transcriptional regulators. Therefore the work was focused on determining new transcriptional regulators of core cell cycle genes and to try to connect these with specific environmental or developmental processes.

Endoreduplication is a cellular process during which nuclear DNA content (ploidy) is increased through successive genome duplication without cell division. Endoreduplication plays pivotal functions throughout the plant life cycle such as morphogenesis or cell specification, and also in response to environmental stresses. Another potential role of endoreduplication is that, by increasing gene copy number, transcription could be increased. In Tomato, the fruit pericarp tissue (fleshy part) is composed of a heterogeneous population of cells displaying a large variation of ploidy levels reaching up to 256c (c = haploid genome quantity). In this tissue, these high ploidy levels are generally correlated with large cells. However, little is known about the onset and progression of endoreduplication during tomato fruit growth and its consequences on the regulation of cell size and gene expression. We therefore aim to determine the in situ distribution of gene expression based on the ploidy levels in the pericarp during fruit development.For that, ploidy distribution in the pericarp is first quantified in situ by Fluorescent in situ Hybridization (FisH). In parallel, cell size is measured to study the potential link between ploidy and cell growth. Second, RNA extracted from nuclei sorted based on their ploidy level are used for sequencing. From this transcriptome data, a search for potential markers of ploidy and/or genes having a ploidy specific expression will be done. These ploidy distribution and transcriptomics experiments are done by harvesting fruits at five stages from 6 to 12 days post anthesis (dpa) during fruit growth. Using this data a virtual map of ploidy distribution and gene expression will be done for early development of Tomato fruit pericarp.; L’endoréduplication, qui est la source majeure d’endopolyploïdie chez les Plantes, est un processus cellulaire durant lequel la quantité d’ADN nucléaire (ploïdie) est augmentée par une succession de réplications du génome sans division cellulaire. L’endoréduplication est le résultat de la capacité des cellules à transformer le classique cycle cellulaire en un cycle cellulaire partiel, l’endocycle, où la synthèse de l’ADN intervient indépendamment de la mitose. Chez les plants, l’endoréduplication est présente dans de nombreux organes et types cellulaires et notamment ceux ayant un fort intérêt agronomique, comme la partie charnue (péricarpe) du fruit de Tomate dans lequel les niveaux de ploïdie sont important. A la fin du développement du fruit de tomate, le péricarpe est composé d’une population hétérogène de cellules dont les niveaux de ploïdie et la tailles varient. De plus, à ce stade, des gènes présentent des profils d’expression spécifiquement lié aux niveaux de ploïdie. Cependant, peu de données sont disponible à propos de l'engagement et la progression de l’endoreduplication dans les cellules durant le développement du fruit de tomate ou son effet sur l’expression des gènes. Afin d’expliquer pourquoi une population hétérogène de cellule co-existent dans le péricarpe et comme base de description des régulations transcriptionnelles par l’endoréduplication dans le péricarpe, la thèse a eu d’étudier cette population hétérogène de cellule afin d’établir les bases de détermination des programmes génétiques mis en place par l’endoreduplication. Donc, j’ai cartographié le dynamisme des niveaux de ploïdie et de l’expression des gènes en fonction des niveaux de ploïdie dans le péricarpe au cours du développement précoce du fruit, à 6-, 9- et 12 jours après anthèse. Les données ont montré qu’un doublement de l’expression était présent en relation avec le doublement de ploïdie et l’endoreduplication est déjà bien en place, organisée en gradient dans le péricarpe à 6DPA. En utilisant les données produites, une carte virtuelle des niveaux de ploïdie et de l’expression des gènes sera établie au cours de du développement du péricarpe du fruit de Tomate.