It is increasingly recognized by the scientific community that the field of epigenetics is a key step for a better understanding of human biology in both normal and pathological states. Its implication in cancer, and in particular in breast cancer, is now well accepted. Breast cancer, responsible for more than 450,000 deaths worldwide yearly, is a heterogenous disease at the histological and clinical levels as well as at the molecular level. Despite considerable efforts to develop new treatments and improve patient management, patients with a same “profile” of breast cancer can respond differently to therapies and have completely different clinical outcomes. There is therefore a critical need to improve our understanding of breast cancer biology and diversity, in order to find new markers that should provide a better management of patients and the development of new therapies. An increasing number of biologists, pathologists as well as clinicians are currently working towards these goals. During my PhD, we have conducted two studies in order to gain new insights into the contribution of epigenetics in breast cancer biology.<p>In the first study, by performing large genome-scale DNA methylation profiling of numerous breast tumors as well as of normal breast tissues, we first revealed the existence of six groups of breast tumors based on their DNA methylation profiles. Three of these groups showed a strong association with the basal-like, HER2 and luminal A breast cancer subtypes, previously identified by gene expression profiling. Interestingly, the three other groups were found to be a mixture of several gene expression-based subtypes, thus revealing the capacity of DNA methylation profiling to improve breast tumor taxonomy. Second, our study suggests that the establishment of DNA methylation patterns of breast tumors might help to determine their cell type of origin. Finally, we also showed that DNA methylation profiling can reflect the cell type composition of the tumor microenvironment and that a signature of T cell tumoral infiltration is associated with a good prognosis in particular categories of breast cancer patients. <p>In the second study, we revealed the clinical relevance of the KDM5 histone demethylases in breast cancer. The expression of these histone demethylases was deregulated in the analyzed breast tumors as well as in the pre-invasive samples as compared to normal breast samples. This suggests that KDM5 enzymes might be good markers for early diagnosis of breast cancer. Moreover, we showed a prognostic value of the KDM5C histone demethylase.<p>In conclusion, the above data should provide a better understanding of breast cancer biology and diversity, and this should bring new insights to improve breast cancer patient management.<p> / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished

Les cellules souches hématopoïétiques (CSH) sont rares, mais indispensables pour soutenir la production des cellules matures du sang, un tissu en constant renouvellement. Deux caractéristiques principales les définissent; la propriété d’auto-renouvellement (AR), ou la capacité de préserver leur identité cellulaire suivant une division, et la multipotence, ce potentiel de différentiation leur permettant de générer toutes les lignée hématopoïétiques. De par leurs attributs, les CSH sont utilisée en thérapie cellulaire dans le domaine de la transplantation. Une organisation tissulaire hiérarchique est aussi préservée dans la leucémie, ou cancer du sang, une masse tumorale hétérogène devant être maintenue par une fraction de cellules au potentiel prolifératif illimité, les cellules souches leucémiques (CSL). Les travaux présentés dans ce manuscrit visent à explorer les bases moléculaires de l’AR, encore mal définies. Certains membres de la famille des facteurs de transcription à homéodomaine HOX sont impliqués dans la régulation de l’hématopoïèse normale, et leur dérégulation peut contribuer à la transformation leucémique. En particulier, la surexpression du gène Hoxb4 dans les CSH influence leur destin cellulaire, favorisant des divisions d’auto-renouvellement et leur expansion en culture et in vivo. En général, les CSH s’épuisent rapidement lorsque maintenue hors de leur niche ex vivo. Différents facteurs interagissent avec les HOX et modulent leur liaison à l’ADN, dont la famille des protéines TALE (Three Amino acid Loop Extension), comme MEIS1 et PBX1. En utilisant une stratégie de surexpression combinée de Hoxb4 et d’un anti-sens de Pbx1 dans les CSH, générant ainsi des cellules Hoxb4hiPbx1lo, il est possible de majorer encore d’avantage leur potentiel d’AR et leur expansion in vitro. Les CSH Hoxb4hiPbx1lo demeurent fonctionnellement intactes malgré une modulation extrême de leur destin cellulaire en culture. Les niveaux d’expressions de facteurs nucléaires, seules ou en combinaison, peuvent donc s’avérer des déterminants majeurs du destin des CSH. Afin d’identifier d’autres facteurs nucléaires potentiellement impliqués dans le processus d’AR des CSH, une stratégie permettant d’évaluer simultanément plusieurs gènes candidats a été élaborée. Les progrès réalisés en termes de purification des CSH et de leur culture en micro-puits ont facilité la mise au point d’un crible en RNAi (interférence de l’ARN), mesurant l’impact fonctionnel d’une diminution des niveaux de transcrits d’un gène cible sur l’activité des CSH. Les candidats sélectionnés pour cette étude font partie du grand groupe des modificateurs de la chromatine, plus précisément la famille des histones déméthylases (HDM) contenant un domaine catalytique Jumonji. Ce choix repose sur la fonction régulatrice de plusieurs membres de complexes méthyl-transférases sur l’AR des CSH, dont l’histone méthyl-transférases MLL (Mixed Lineage Leukemia). Cette stratégie a aussi été utilisée dans le laboratoire pour étudier le rôle de facteurs d’asymétrie sur le destin des CSH, en collaboration. Ces études ont permis d’identifier à la fois des régulateurs positifs et négatifs de l’activité des CSH. Entre autre, une diminution de l’expression du gène codant pour JARID1B, une HDM de la lysine 4 de l’histone H3 (H3K4), augmente l’activité des CSH et s’accompagne d’une activation des gènes Hox. En conclusion, divers déterminants nucléaires, dont les facteurs de transcription et les modificateurs de la chromatine peuvent influencer le destin des CSH. Les mécanismes sous-jacents et l’identification d’autres modulateurs de l’AR demeurent des voies à explorer, pouvant contribuer éventuellement aux stratégies d’expansion des CSH ex vivo, et l’identification de cibles thérapeutiques contre les CSL. Mots-clés : cellules souches hématopoïétiques, Hoxb4, Pbx1, auto-renouvellement, histone déméthylases, RNAi / Hematopoietic stem cells (HSC) are rare, but essential to sustain the constant production of all mature blood cells, a constantly renewing tissue. They are defined by two main characteristics; namely self-renewal (SR), or the capacity to preserve cell identity following division, and multipotency, the differentiation potential that allows them to generate all hematopoietic lineages. Given their attributes, HSC are used for cellular therapy in the transplantation field. A hierarchy in tissue organisation is also preserved in leukemia, or blood cancer, a heterogeneous tumor mass that is sustained by a subset of cells with unlimited SR potential, the leukemia stem cells (LSC). Studies presented in this manuscript aim to explore the molecular basis underlying SR, which are still poorly defined. Certain members of the HOX family of homeodomain transcription factors are involved in the regulation of normal hematopoiesis, and their deregulation can contribute to leukemia development. In particular, Hoxb4 overexpression in HSC influences cells fate, favouring SR divisions and their subsequent expansion in culture and in vivo. In general, HSC exhaust rapidly when maintained ex vivo, outside of their niche. Several factors interact with HOX and modulate their binding to DNA, including members of the TALE (Three Amino acid Loop Extension) protein family, such as MEIS1 and PBX1. Using a strategy of combined overexpression of Hoxb4 and an anti-sense to Pbx1in HSC, generating Hoxb4hiPbx1lo cells, it is possible to further impact on their SR potential and expansion in vitro. These Hoxb4hiPbx1lo cells remain functionally intact despite extreme modulation of their cell fate in culture. Levels of expression of nuclear factors, alone or in combination, can thus impact significantly on HSC fate. In order to identify other nuclear factors potentially involved in the process of HSC self-renewal, a strategy enabling simultaneous assessment several gene candidates was elaborated. To this end, progress made in terms of HSC purification and their culture in micro-wells facilitated the setup of an RNAi (RNA interference) screen, measuring the functional impact of lowering gene candidate transcript levels on HSC activity. Gene candidates selected for this study belong to the greater group of chromatin modifiers, more specifically the family of histone demethylases (HDM) containing a Jumonji catalytic domain. This choice stems from the regulatory function of several members of histone methyl-transferase complexes on HSC self-renewal, including the histone methyl-transferase MLL (Mixed Lineage Leukemia). This strategy was also used in the laboratory to study the role of asymmetry factors on HSC fate, in a collaborative study. These studies enabled identification of both positive and negative regulators of HSC activity. Among these, reduced expression of the gene coding for JARID1B, a histone 3 lysine 4 (H3K4) HDM, increased HSC activity was associated with Hox genes activation. In conclusion, several nuclear determinants, including transcription factors and chromatin modifiers, can influence HSC fate. Underlying mechanisms and identification of additional modulators of SR remain areas to explore, which could eventually contribute to HSC expansion strategies ex vivo, and identification of therapeutic targets against LSC. Keywords: hematopoietic stem cells, Hoxb4, Pbx1, self-renewal, histone demethylases, RNAi

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

L’O-GlcNAcylation est une modification post-traductionnelle qui consiste en l’ajout covalent du N-acetylglucosamine au groupement hydroxyle des sérines et thréonines des protéines nucléaires et cytoplasmiques. Ce type de glycosylation atypique est régulé de manière très dynamique par l’action de l’O-GlcNAc transférase (OGT) et de l’O-GlcNAcase (OGA) qui catalysent et hydrolysent cette modification respectivement. Aujourd’hui, OGT émerge comme un régulateur transcriptionnel et senseur critique du métabolisme où les protéines ciblées par l’O-GlcNAcylation couvrent la presque totalité des voies de signalisation cellulaire. Récemment, des études ont aussi proposé qu’OGT soit impliquée dans la régulation épigénétique par l’O-GlcNAcylation des histones. Dans le but de caractériser le rôle fonctionnel d’OGT dans la régulation épigénétique, nous avons revisité le concept d’O-GlcNAcylation des histones et, de manière surprenante, n’avons pu confirmer cette observation. En fait, nos données indiquent que les outils disponibles pour détecter l’O-GlcNAcylation des histones génèrent des artéfacts. De ce fait, nos travaux supportent plutôt un modèle où la régulation épigénétique médiée par OGT se fait par l’O-GlcNAcylation de régulateurs transcriptionnels recrutés à la chromatine. Parmi ceux-ci, OGT s’associe au complexe suppresseur de tumeurs BAP1. En étudiant le rôle d’OGT dans ce complexe, nous avons identifié le facteur de transcription FOXK1 comme un nouveau substrat d’OGT et démontrons qu’il est régulé par O-GlcNAcylation durant la prolifération cellulaire. Enfin, nous démontrons que FOXK1 est aussi requis pour l’adipogenèse. Ensemble, nos travaux suggèrent un rôle important d’OGT dans la régulation du complexe BAP1. / O-GlcNAcylation is a post-translational modification which consists in the covalent addition of an N-acetylglucosamine sugar to the hydroxyl group of serine and threonine residues of nuclear and cytoplasmic substrates. This atypical glycosylation is regulated in a very dynamic manner through the action of the O-GlcNAc transferase (OGT) and the O-GlcNAcase (OGA) that catalyze and hydrolyze this modification respectively. OGT has emerged as a critical transcriptional regulator and sensor of metabolism whereby proteins targeted by O-GlcNAcylation cover several cell signaling pathways. Recently, studies have also suggested that OGT may be involved in epigenetic regulation through the O-GlcNAcylation of histones. For the purpose of characterizing the functional role of OGT in epigenetic regulation, our group revisited the concept of histone O-GlcNAcylation and surprisingly, our work could not confirm this observation. In fact, our data indicate that the available tools for histone O-GlcNAcylation detection generate artifacts. Consequently, our work rather supports a model whereby OGT-mediated epigenetic regulation is indirectly achieved through O-GlcNAcylation of chromatin-associated transcriptional regulators. Among these, OGT strongly associates with the BAP1 tumor suppressor complex. Thus, by focusing on the role of OGT in this complex, we identified the transcription factor FOXK1 as a novel substrate of OGT and demonstrate that it is regulated throught O-GlcNAcylation during cell proliferation. Finally, we demonstrate that FOXK1 is also required for adipogenesis. Taken together, these data suggest an important role of OGT in regulating the BAP1 complex.

A proteína dermicidina (DCD) é codificada por um gene localizado na porção 12q13 do cromossomo 12, presente apenas em primatas e humanos. A proteína é secretada por células de glândulas da pele, melanócitos, neurônios e células epiteliais da mama normal. Alguns estudos inciais revelaram a participação da proteína DCD em processos oncogênicos nos carcinomas de mama, próstata e melanoma pela sua capacidade de atuar como um fator de crescimento e sobrevivência celular. Nos estudos realizados no nosso laboratório mostramos que a DCD é expressa em células normais da pele, placenta, cérebro e em vários tumores, incluindo os carcinomas de mama e melanoma maligno. Ensaios biológicos e bioquímicos mostraram que o \"knockdown\" da expressão de DCD no melanoma maligno G-361 via RNA de interferência (RNAi) diminuiu significativamente o crescimento in vitro em cultura celular e a formação de tumores em camundongos Nude. Resultados similares foram obtidos quando camundongos Nude transplantados com células de melanoma G-361 foram tratados com anticorpos policlonais de coelhos contra a proteína DCD. Para compreender melhor o papel da proteína DCD na transformação de células de melanoma G-361 foram feitos ensaios de microarranjo de DNA para identificar os genes diferencialmente expressos entre as sublinhagens pLKO (controle) e IBC-I que expressa o shRNA para o mRNA da DCD. Entre os 374 genes alterados pelo silenciamento, encontramos 162 com expressão aumentada e 212 com a expressão reduzida. Os estudos de bioinformática pelo software MetaCore mostraram que o silenciamento do gene DNA modula as vias canônicas e redes de sinalização mediadas pelo receptores e ligantes da família BAFF/APRIL que contralam a ativação do fator de transcrição NF-kB, bem como para histonas envolvidas na remodelação da cromatina. Os níveis de expressão de mRNA de 9 genes de interesse foram validados por ensaios de RT-qPCR. Em uma segunda fase do estudo, foram analisadas as proteínas presentes em extratos nuclares dos clones pLKO e IBC-I de melanoma maligno G-361 por espectrometria de massas. Nos extratos proteicos da sublinhagem pLKO foram identificadas 74 proteínas nucleares, enquanto que na sublinhagem IBC-I foram identificadas 31 proteínas. Um grupo de 21 proteínas foi identificado em ambas sublinhagens. Estudos de bioinformática pelo programa STRING revelou que 14 das proteínas identificadas na sublinagem G-361-pLKO faziam interações diretas ou indiretas com a DCD. A rede formada por estas proteínas tem como centro a proteína p53, uma proteína chave na regulação do programa de morte celular e sobrevivência ao estresse oxidativo. Por outro lado, notou-se que a maioria das proteínas identificadas no extrato nuclear da sublinhagem IBC-I é da família das histonas e que poderiam atuar em complexos de remodelação da cromatina nas células G-361-IBC-I. Nossos resultados nos possibilitaram sugerir que futuros estudos sobre a expressão das histonas e suas modificações pós-traducionais poderão ajudar a desvendar o possível papel da DCD na regulação epigenética do melanoma e em outros tipos de cânceres / Dermcidin (DCD) is a human gene mapped to chromosome 12q13 region, only identified in primates and humans, and normally expressed in the eccrine glands of skin and brain. Several studies have confirmed that DCD-derived peptides contribute to innate and immune surveillance and in the oncogenic processes of breast, prostate and skin cancers, as revealed by its role as a growth factor and cell survival. We have further explored DCD function and its tumorigenic potential on skin melanocytes by specifically knocking down its expression in G-361 malignant melanoma cells via expressing constitutively short hairpin RNA against DCD mRNA. Biological and biochemical assays showed that the \"knockdown\" in the expression of DCD in G-361-pLKO control clone and a G-361-IBC-I clone expressing constitutively short hairpin RNA against DCD mRNA decreased significantly the in vitro growth in cell culture and tumor formation in nude mice. Similar results were obtained treating nude mice bearing G-361 melanoma xenografts with rabbit polyclonal antibodies against DCD protein. Here, we present a DNA microarray-based study that identified the genes that are up- and down-regulated in a G-361-pLKO control clone and a G-361-IBC-I clone expressing constitutively short hairpin RNA against DCD mRNA. A total of 372 genes differentially expressed were identified; being 162 genes up-regulated and 212 genes down-regulated. Bioinformatic studies showed that DCD gene silencing modulates canonical pathways and signaling networks mediated APRIL/BAFF receptors and ligands and NF-kB signaling pathway as well as chromatin remodeling mediated by histone family. The mRNA expression levels of 9 genes of interest were validated by RT-qPCR assays. Next we analyzed the proteins present in nuclear extracts from G-361- pLKO and G-361-IBC-I clones by mass spectrometry. We identified 74 proteins in the G-361-pLKO clone and 31 proteins in the G-361-IBC-I. A group of 21 proteins was identified in both sublineages. Bioinformatics analyses by STRING (Search Tool for the Retrieval of Interacting Genes/Proteins) platform showed that a small portion of the proteins identified only in G-361- pLKO cells was predicted to interact directly with DCD. The network formed by these proteins is centered in the p53 protein, a key regulator of survival and cell death program in response to DNA damage and oxidative stress. On the other hand, this network was completed abrogated using the nuclear protein from G-361-IBC-I because of absence of DCD protein. Since most of the proteins identified in nuclear extracts are of the histone family, it is likely that they are acting in the chromatin-remodeling complexes which are important to remodel nucleosomes of the G-361-IBC-I cells. Our results allowed us to suggest that future studies on the expression of histones and their posttranslational modifications may help to unravel the possible role of DCD in the epigenetic regulation of melanoma and other cancers

O nucléolo é uma organela nuclear responsável pela produção dos ribossomos, através das Regiões Organizadoras do Nucléolo (NORs). Espécies que possuem mais de um par de cromossomos contendo NORs terão, obrigatoriamente, pelo menos um par ativo, sendo as demais NORs funcionais de acordo com a demanda celular. O mecanismo de compensação de dose é visualizado e bem estabelecido em híbridos interespecíficos, conhecido como dominância nucleolar, com a inativação de NORs de um dos parentais por outras homeólogas ativas que as dominam. A arquitetura da cromatina nas NORs e o controle da sua expressão foram estudados com o objetivo de se entender os mecanismos envolvidos no fenômeno da dominância nucleolar em espécies diplóides que possuem múltiplas NORs. A espécie modelo utilizada neste estudo foi Crotalaria juncea (Leguminosae-Papilionoideae), caracterizada por conter 2n=2x=16, e NORs no braço curto do cromossomo 1, sendo este o principal organizador do nucléolo, e no braço longo do cromossomo 4 adjacente à heterocromatina centromérica, sendo este um sítio adicional (sítio menor) e de expressão facultativa, previamente determinada. Nas raízes de C. juncea sincronizadas, observou-se que a nucleologênese tem seu início durante o final da telófase, em que os 4 sítios de genes ribossomais podem ter atividade e formar até 4 nucléolos, os quais tendem a se fundir durante a interfase. A Hibridação in situ fluorescente (FISH) permitiu estudos da arquitetura da cromatina, com a visualização dos territórios cromossômicos, onde a cromatina não está organizada de forma aleatória dentro do núcleo, e consequentemente o rDNA 45S dentro do nucléolo. Observou-se também que todos os sítios de rDNA 45S possuem diferença no tamanho do arranjo repetitivo. Assim sendo, a hierarquia de dominância está de acordo com o tamanho de cada arranjo (sítio), e estes são ativados de acordo com a demanda celular. As análises das modificações nas histonas mostraram que a H3K9Met1 apresentou marcas fracas no nucléolo, enquanto no restante da cromatina nuclear sua marcação foi intensa. Já a H3K9Met2 apresentou marcação fortemente associada à cromatina presente no nucléolo, com alguns pequenos pontos heterocromáticos dispersos no núcleo. Pela observação entende-se que ambas metilações controlam diferentes tipos de heterocromatinas, ou seja, a H3K9Met2 controla principalmente heterocromatinas associadas aos genes ribossomais, e a H3K9Met controla heterocromatinas não associadas ao rDNA. O rDNA é hiperacetilado dentro do nucléolo para a H3K14. Não foi observada marcação nucleolar para H4K8ac, mas pôde ser observadas regiões hiperacetiladas em outras regiões da cromatina. A metilação do DNA esteve diretamente associada à diferentes níveis de organização da cromatina das NORs. As heterocromatinas adjacentes ao nucléolo apareceram fortemente metiladas, enquanto a cromatina distendida dentro do nucléolo apresentou marcação dispersa, com algumas regiões mais fortemente marcadas, onde a cromatina apresentava-se mais condensada e provavelmente não associados com a cromatina ativa. As fibras estendidas permitiram uma análise de alta resolução, onde foi possível observar que regiões não metiladas apareciam intercaladas entre grandes regiões fortemente metiladas, sugerindo que estas regiões hipometiladas estão, possivelmente, associadas com as alças de transcrição dentro do nucléolo. 12 Esses resultados contribuem para o entendimento sobre o controle genético e epigenético na arquitetura da cromatina ribossomal, bem como seu controle na expressão dos genes ribossomais no genoma das plantas. / The nucleolus is a nuclear organelle responsible for the ribosomes production, by Nucleolus Organizer Regions (NORs). Species presenting more than one chromosome pair with NORs should present, one pair expressing the genes, at least; while the other pairs expressing their genes accordingly to cellular demand. Dosage compensation mechanism is visualized and well established of interspecific hybrids as a well-described phenomena named nucleolar dominance, where a NOR from one parental could lead to inactivation of a NOR from the other parental which is dominated. The chromatin architecture and expression of the NORs were studied to address the mechanism involved in the nucleolar dominance of diploid species containing multiple sites of 45S rDNA. The model species used in the present study was the crop Crotalaria juncea (Leguminosae-Papilionoideae) characterized by 2n=2x=16 chromosomes, being the main NOR mapped into chromosome 1 short arm and presenting an additional site (minor site) in the chromosome 4 long arm adjacent to a centromeric heterochromatin and facultatively expressed. Synchronized meristematic root tip cells determined to nucleologenesis starts during the late-telophase, often expressing every ribosomal gene sites, when up to four nucleoli could be observed and these become merged during interphases. FISH allowed nucleolar chromatin architecture be accessed revealing distinct chromosomal domains (territories), suggesting a non-random distribution of the 45S rDNA, even between homologous chromosomes, into the nucleolus. The 45S rDNA sites from both chromosome pairs 1 and 4 of C. juncea showed differences in their array sizes. The differences in the 45S rDNA array sizes and the order of loci expression suggest a hierarchy of dominance, a feature of nucleolar dominance; being the small RONs activated only on demand. Immunodetection of histone modifications showed different patterns to methylation distribution across the chromatin as a whole; where H3K9Met1 was found mainly distributed along the nuclear chromatin without an evident signal into nucleolus, while H3K9Met2 was detected as conspicuous dots in the nuclear chromatin and highly accumulated into the nucleolus. The results indicate different control on heterochromatin establishment and maintenance, being the modifications specific to certain chromosomal regions. Indeed, H3K9Met is a key component in the nucleolus chromatin architecture and expression. The chromatin inside the nucleolus showed a high accumulation of H3K14ac, with a weak fluorescent signal along the nucleus; on the other hand H4K8ac showed a strong signal homogenously distributed across the nuclear chromatin, but without evident signals inside the nucleolus. DNA methylation was directly associated with different levels of chromatin organization of the NORs. The heterochromatic regions associated to RON are highly methylated, while the chromatin inside the nucleolus showed weaker signals, with some bright spots probably in condensed regions and related to chromatin inactivity. Extended DNA fiber allowed a higher resolution mapping that revealed long methylated regions intermingled by nomethylated ones, being the last probably associated to transcriptional loops of rRNA genes into the nucleolus. The results presented herein contributes to a better understand about the nucleolar chromatin architecture and the genetic and epigenetic control of the ribosomal genes expression on plant genomes.

No description available.

Within the lymphocyte lineage, GATA-3 is a major transcription factor implicated in the regulation of Th1/Th2 differentiation by promoting the expression of the Th2 cytokines, such as IL-4, IL-5, IL-10 and IL-13. Although the role of GATA-3 in the development of the Th2 lineage has been extensively described in the literature, the molecular mechanisms underlying its activity remain to be clarified. Here, we investigated whether GATA-3 might be regulated by reversible acetylation. In vivo, GATA-3 associates with class I and III HDACs. Biochemical studies unraveled the specific association of GATA-3 with the class III member SIRT1. Association with SIRT1 leads to the inhibition of GATA-3-induced IL-5 transcription. Using siRNA, we further show that SIRT1 promotes destabilization of GATA-3. Interestingly, nicotinamide, a specific inhibitor of SIRTs had no effect on the ability of SIRT1 to destabilize GATA-3 and to repress its transcriptional activity. In addition, a catalytic-defective mutant of SIRT1 (H363Y) shows similar effects to wild-type SIRT1, demonstrating that the deacetylase activity of SIRT1 is not required for its regulation of GATA-3. For the first time, our study indicates that SIRT1 is functionally linked to GATA-3. Moreover, our results also suggest that some important SIRT1 functions may not require its deacetylase activity.

La méthylation de l'ADN et les modifications post-traductionnelles des histones sont desmodifications épigénétiques qui interviennent dans la régulation des éléments transposables(ET) chez de nombreuses espèces. La proportion des ET dans les génomes varie selon lesespèces considérées et pose la question des mécanismes de régulation de ces ET. Au sein del'espèce Drosophila simulans, les populations naturelles présentent un polymorphisme uniquedans le nombre de copies des ET, ce qui en fait un excellent modèle pour étudier cettequestion. L'étude de la méthylation d'ADN et des modifications post-traductionnelles deshistones associées au rétrotransposon à LTR tirant dans la lignée germinale des populationsnaturelles a permis de montrer l'influence d'une copie d'ET sur la structure de la chromatineau site d'insertion. Dans un second volet, nous avons cherché à caractériser la méthylation del'ADN chez la drosophile, chez laquelle la fonction est encore mal connue. Nous avons, pardes approches spécifiques et globales, mesuré l'abondance de cette marque épigénétique chezla drosophile. Nous concluons que les taux de méthylation de l'ADN sont très faibles maisvariables entre espèces. Notre travail n'a pas permis de mettre en évidence un rôle de laméthylation de l'ADN dans le contrôle des ET, toutefois, nous ne pouvons pas exclure cesystème de régulation.

No description available.