La plupart des conditions détectées par le dépistage néonatal sont reliées à l'une des enzymes qui dégradent les acyls-CoA mitochondriaux. Le rôle physiopathologique des acyls-CoA dans ces maladies est peu connue, en partie parce que les esters liés au CoA sont intracellulaires et les échantillons tissulaires de patients humains ne sont généralement pas disponibles. Nous avons créé une modèle animal murin de l'une de ces maladies, la déficience en 3-hydroxy-3-methylglutaryl-CoA lyase (HL), dans le foie (souris HLLKO). HL est la dernière enzyme de la cétogenèse et de la dégradation de la leucine. Une déficience chronique en HL et les crises métaboliques aigües, produisent chacune un portrait anormal et distinct d'acyls-CoA hépatiques. Ces profils ne sont pas prévisibles à partir des niveaux d'acides organiques urinaires et d'acylcarnitines plasmatiques. La cétogenèse est indétectable dans les hépatocytes HLLKO. Dans les mitochondries HLLKO isolées, le dégagement de 14CO2 à partir du [2-14C]pyruvate a diminué en présence de 2-ketoisocaproate (KIC), un métabolite de la leucine. Au test de tolérance au pyruvate, une mesure de la gluconéogenèse, les souris HLLKO ne présentent pas la réponse hyperglycémique normale. L'hyperammoniémie et l'hypoglycémie, des signes classiques de plusieurs erreurs innées du métabolisme (EIM) des acyls-CoA, surviennent de façon spontanée chez des souris HLLKO et sont inductibles par l'administration de KIC. Une charge en KIC augmente le niveau d'acyls-CoA reliés à la leucine et diminue le niveau d'acétyl-CoA. Les mitochondries des hépatocytes des souris HLLKO traitées avec KIC présentent un gonflement marqué. L'hyperammoniémie des souris HLLKO répond au traitement par l'acide N-carbamyl-L-glutamique. Ce composé permet de contourner une enzyme acétyl-CoA-dépendante essentielle pour l’uréogenèse, le N-acétylglutamate synthase. Ceci démontre un mécanisme d’hyperammoniémie lié aux acyls-CoA. Dans une deuxième EIM des acyls-CoA, la souris SCADD, déficiente en déshydrogénase des acyls-CoA à chaînes courtes. Le profil des acyls-CoA hépatiques montre un niveau élevé du butyryl-CoA particulièrement après un jeûne et après une charge en triglycérides à chaîne moyenne précurseurs du butyryl-CoA. / Most conditions detected by expanded newborn screening result from deficiency of one of the enzymes that degrade acyl-CoA esters in mitochondria. The role of acyl-CoAs in the pathophysiology of these disorders is poorly understood, in part because CoA esters are intracellular and samples are not generally available from human patients. We created a mouse model of one such condition, deficiency of 3-hydroxy-3-methylglutaryl-CoA lyase (HL), in liver (HLLKO mice). HL catalyses a reaction of ketone body synthesis and of leucine degradation. Chronic HL deficiency and acute crises each produced distinct abnormal liver acyl-CoA patterns, which would not be predictable from levels of urine organic acids and plasma acylcarnitines. In HLLKO hepatocytes, ketogenesis was undetectable. Measures of Krebs cycle flux diminished following incubation of HLLKO mitochondria with the leucine metabolite 2-ketoisocaproate (KIC). HLLKO mice also had suppression of the normal hyperglycemic response to a systemic pyruvate load, a measure of gluconeogenesis. Hyperammonemia and hypoglycemia, cardinal features of many inborn errors of acyl-CoA metabolism, occurred spontaneously in some HLLKO mice and were inducible by administering KIC. KIC loading also increased levels of several leucine-related acyl-CoAs and reduced acetyl-CoA levels. Ultrastructurally, hepatocyte mitochondria of KIC-treated HLLKO mice show marked swelling. KIC-induced hyperammonemia improved following administration of carglumate (N-carbamyl-L-glutamic acid), which bypasses an acetyl-CoA-dependent reaction essential for urea cycle function, thus demonstrating an acyl-CoA-related mechanism for this complication. In a second animal model of an inborn error of acyl-CoA metabolism, short chain acyl-CoA dehydrogenase (SCAD)-deficient mice, the main finding in liver acyl-CoAs is increased butyryl-CoA, particularly during fasting or after enteral loading with medium chain triglyceride precursor of butyryl-CoA.

La plupart des conditions détectées par le dépistage néonatal sont reliées à l'une des enzymes qui dégradent les acyls-CoA mitochondriaux. Le rôle physiopathologique des acyls-CoA dans ces maladies est peu connue, en partie parce que les esters liés au CoA sont intracellulaires et les échantillons tissulaires de patients humains ne sont généralement pas disponibles. Nous avons créé une modèle animal murin de l'une de ces maladies, la déficience en 3-hydroxy-3-methylglutaryl-CoA lyase (HL), dans le foie (souris HLLKO). HL est la dernière enzyme de la cétogenèse et de la dégradation de la leucine. Une déficience chronique en HL et les crises métaboliques aigües, produisent chacune un portrait anormal et distinct d'acyls-CoA hépatiques. Ces profils ne sont pas prévisibles à partir des niveaux d'acides organiques urinaires et d'acylcarnitines plasmatiques. La cétogenèse est indétectable dans les hépatocytes HLLKO. Dans les mitochondries HLLKO isolées, le dégagement de 14CO2 à partir du [2-14C]pyruvate a diminué en présence de 2-ketoisocaproate (KIC), un métabolite de la leucine. Au test de tolérance au pyruvate, une mesure de la gluconéogenèse, les souris HLLKO ne présentent pas la réponse hyperglycémique normale. L'hyperammoniémie et l'hypoglycémie, des signes classiques de plusieurs erreurs innées du métabolisme (EIM) des acyls-CoA, surviennent de façon spontanée chez des souris HLLKO et sont inductibles par l'administration de KIC. Une charge en KIC augmente le niveau d'acyls-CoA reliés à la leucine et diminue le niveau d'acétyl-CoA. Les mitochondries des hépatocytes des souris HLLKO traitées avec KIC présentent un gonflement marqué. L'hyperammoniémie des souris HLLKO répond au traitement par l'acide N-carbamyl-L-glutamique. Ce composé permet de contourner une enzyme acétyl-CoA-dépendante essentielle pour l’uréogenèse, le N-acétylglutamate synthase. Ceci démontre un mécanisme d’hyperammoniémie lié aux acyls-CoA. Dans une deuxième EIM des acyls-CoA, la souris SCADD, déficiente en déshydrogénase des acyls-CoA à chaînes courtes. Le profil des acyls-CoA hépatiques montre un niveau élevé du butyryl-CoA particulièrement après un jeûne et après une charge en triglycérides à chaîne moyenne précurseurs du butyryl-CoA. / Most conditions detected by expanded newborn screening result from deficiency of one of the enzymes that degrade acyl-CoA esters in mitochondria. The role of acyl-CoAs in the pathophysiology of these disorders is poorly understood, in part because CoA esters are intracellular and samples are not generally available from human patients. We created a mouse model of one such condition, deficiency of 3-hydroxy-3-methylglutaryl-CoA lyase (HL), in liver (HLLKO mice). HL catalyses a reaction of ketone body synthesis and of leucine degradation. Chronic HL deficiency and acute crises each produced distinct abnormal liver acyl-CoA patterns, which would not be predictable from levels of urine organic acids and plasma acylcarnitines. In HLLKO hepatocytes, ketogenesis was undetectable. Measures of Krebs cycle flux diminished following incubation of HLLKO mitochondria with the leucine metabolite 2-ketoisocaproate (KIC). HLLKO mice also had suppression of the normal hyperglycemic response to a systemic pyruvate load, a measure of gluconeogenesis. Hyperammonemia and hypoglycemia, cardinal features of many inborn errors of acyl-CoA metabolism, occurred spontaneously in some HLLKO mice and were inducible by administering KIC. KIC loading also increased levels of several leucine-related acyl-CoAs and reduced acetyl-CoA levels. Ultrastructurally, hepatocyte mitochondria of KIC-treated HLLKO mice show marked swelling. KIC-induced hyperammonemia improved following administration of carglumate (N-carbamyl-L-glutamic acid), which bypasses an acetyl-CoA-dependent reaction essential for urea cycle function, thus demonstrating an acyl-CoA-related mechanism for this complication. In a second animal model of an inborn error of acyl-CoA metabolism, short chain acyl-CoA dehydrogenase (SCAD)-deficient mice, the main finding in liver acyl-CoAs is increased butyryl-CoA, particularly during fasting or after enteral loading with medium chain triglyceride precursor of butyryl-CoA.

Após a realização de uma sessão de treinamento (ST) é comum a ocorrência do fenômeno denominado dano muscular induzido pelo exercício (DMIE), que se caracteriza por prejuizos a estrutura da fibra muscular, com ruptura de alguns sarcômeros, desordem miofibrilar e alargamento das linhas Z. Ainda em consequência ao DMIE, surgem alguns sintomas que são utilizados como marcadores indiretos: dor muscular de início tardio (DMIT), redução na produção de força, aumento de enzimas e proteínas na corrente sanguínea e inchaço. O presente estudo examinou os efeitos da suplementação nutricional a fim de minimizar os efeitos deletérios do DMIE em 3 experimentos. No 1° estudo, 36 indivíduos inexperientes em treinamento de força (TF) foram suplementados com: placebo (PLA, n=12, 50mg·kg-1 de carboidrato); leucina (LEU) baixa dose (LBD, n=12, 50mg·kg-1 de LEU + 50mg·kg-1 de carboidrato) e LEU alta dose (LAD, n=12, 250mg·kg-1 de LEU + 50mg·kg-1 de carboidrato) por 6 dias antecedentes a sessão de treinamento (ST), e nos 3 dias seguintes. Foi observada redução significante, p<0.05, na dor muscular de início tardio (DMIT) do peitoral por palpação, e alongamento nos momentos 48h, e 72h após a ST no grupo LBD comparado ao PLA. A redução no teste de 1 repetição máxima (1RM) apresentou significância no grupo PLA em todos momentos após ST. O aumento na atividade da creatina quinase (CK) foi significante no grupo PLA comparado ao LAD em 24h, 48h e 72h após a ST, enquanto o aumento da concentração de mioglobina (Mb) foi significante no grupo PLA comparado ao grupo LBD e LAD em 24h, 48h e 72h após a ST. O 2° estudo contou com a participação de 28 indivíduos com até 6 meses de experiência em TF. Os sujeitos foram suplementados com 3g de &beta;-hidroxi-&beta;-metilbutirato (HM) por 14 dias (H14, n=07); 7 dias (H07, n=07) e placebo por 14 dias (P14) ou 7 dias (P07, n=07) antecedentes a ST, e nos 3 dias seguintes. O aumento da DMIT por palpação e alongamento foi significante no grupo P14 comparado ao H14 em 24h (apenas alongamento), 48h e 72h após ST, ainda no momento 72h o grupo P07 era superior ao H07. A redução no teste de 1RM ocorreu nos 4 grupos imediatamente após, foi mantida em 24h após a ST nos grupos H14, H07 e P07, sem diferenças entre os grupos. O aumento na concentração de Mb foi significante no grupo P14 comparado ao grupo H14. No 3° estudo, 24 indivíduos experientes em TF foram suplementados com 7g de arginina (ARG, n=12) ou placebo (PLA, n=12, 7g carboidrato) 30 minutos pré-ST. O grupo PLA apresentou aumento significante na DMIT por palpação em 24h comparado ao grupo ARG. A redução no teste de 1RM alcançou significância apenas em 24h após a ST no grupo PLA, mas sem diferença entre os grupos. Os resultados do presente estudo permitem concluir que a suplementação nutricional implementada atenuou o comportamento de alguns marcadores indiretos DMIE, com maior efeito para a DMIT e parametros bioquímicos / After performing a training session (TS) is common the occurrence of the phenomenon called muscle damage induced by exercise (DMIE), which is characterized by damage to muscle fiber structure, breaking some sarcomeres, myofibrillar disorder and extension lines Z. As a consequence of DMIE, there are some symptoms that are measured as indirect markers: delayed onset muscle soreness (DOMS), reduction in strength production, increase of enzymes and proteins in the bloodstream, and swelling. The effect of nutritional interventions to minimize deleterious responses associated with exercise-induced muscle damage (EIMD) were investigated in 3 experiments. In study 1, 36 inexperienced subjects in resistance training (RT) were supplemented for 6 days prior to the training session (TS), and in the following 3 days with: placebo (PLA, n=12, 50mg·kg-1 of carbohydrate); leucine (LEU) low dose (LLD, n=12, 250mg·kg-1 LEU + 50mg·kg-1 + carbohydrate) and LEU high dose (LHD, n=12, 250mg·kg-1 LEU + 50mg·kg-1 + carbohydrate). There was a significant reduction (p <0.05) in delayed onset muscle soreness (DOMS), of the chest by palpation and stretching at 48h, after TS in the LLD group compared to PLA. A significant reduction in the one repetition maximum (1RM) test was observed in the PLA group at all times after TS. The increase in creatine kinase (CK) activity was significant in the PLA group compared to the LHD in 24h, 48h and 72h after TS, while the increase in myoglobin concentration (Mb) was significant in the PLA group compared to the LLD and LHD group in 24h, 48h, and 72h after TS. In study 2, 28 subjects with up to 6 months of RT experience were supplemented with 3g of &beta;-hydroxy-&beta;-methylbutyrate (HM&beta;) for 14 days (H14, n=7); for 7 days (H07, n=7), and placebo for 14 days (P14, n=7) or 7 days (P07, n=7) antecedent to ST, and in the next 3 days. The increase in DOMS by palpation and stretching was significant in the P14 group compared to H14 in 24h (stretching only), 48h and 72h after TS, yet at 72h the P07 group was higher than H07. The reduction in the 1RM test occurred in the 4 groups immediately after and maintained within 24h after TS in groups H14, H07 and P07, and there was no difference between groups. The increase in Mb concentration was significant in the P14 group compared to the H14 group. In study 3, 24 resistance-trained subjects were supplemented with 7g of arginine (ARG, n=12) or placebo (PLA, n=12, 7g of carbohydrate) 30 minutes pre- TS. The PLA group presented a significant increase in DOMS by palpation in 24h compared to the ARG group, and a significant reduction in the 1RM test only in 24h after ST in the PLA group, but without a significant difference between groups. The results of the present study suggest that the responses of indirect markers associated with EIMD were attenuated by nutritional interventions, with greater effect for DOMS and biochemical parameters

O músculo esquelético é um tecido muito importante para a saúde, e seu desequilíbrio está relacionado com diversas doenças. Existe um grande interesse na identificação e caracterização dos agentes/mecanismos responsáveis pelo controle das vias anabólicas/catabólicas da massa muscular que contribuem para a manutenção da homeostase do músculo esquelético. A atrofia muscular é caracterizada pela perda de massa muscular, levando a redução de capacidade funcional. Dada à importância deste tecido na locomoção e muitas outras funções fisiológicas do organismo, o processo de atrofia pode inferir ao indivíduo o aumento da morbidade e perda de sua qualidade de vida, tornando assim, de grande importância a compreensão dos processos moleculares envolvidos. Neste trabalho exploramos a capacidade da suplementação de leucina em proteger a massa muscular durante a atrofia induzida por imobilização, através da inibição de vias catabólicas. A leucina é um alvo atraente, uma vez que é considerado um aminoácido anti-atrófico capaz de regular as vias intracelulares envolvidas na síntese e degradação das proteínas, desta forma, abordamos se o efeito anti-atrófico da leucina envolve modulação do eixo GSK3-&#946;/&#946;-Catenina. Ratos Wistar, suplementados com leucina, tiveram sua pata posterior esquerda imobilizada com o músculo sóleo em posição encurtada. Em seguida, o músculo sóleo foi removido, pesado e processado para avaliar a expressão de genes e proteínas por qPCR e Western blot, respectivamente. Além disso, secções transversais musculares foram utilizadas em ensaios de imunofluorescência para análise de localização celular. Após 1 dia de imobilização, foi observado a ativação de GSK3-&#946; no músculo sóleo, e a suplementação de leucina foi capaz de bloquear esse efeito. Embora os níveis proteicos de &#946;-Catenina tenham sido inalterados pela imobilização ou pela suplementação de leucina, a análise de localização celular, mostrou uma diminuição nuclear da &#946;-Catenina causada pela imobilização, porém, a suplementação de leucina foi capaz de aumentar os níveis de &#946;-Catenina no núcleo. A análise Confocal confirmou a translocação nuclear de &#946;-Catenina, durante a suplementação de leucina. Também analisamos os níveis de expressão de NF-&#954;B por ser potencialmente regulado por &#946;-Catenina, no entanto, não foram observadas alterações nos níveis desta proteína entre os grupos. Assim, este estudo revela &#946;-Catenina como uma nova molécula chave envolvida nos efeitos anti-atróficos da leucina. / Skeletal muscle is a very important tissue for health, and its depletion predicts the prognosis of several diseases. There is great interest in the identification and characterization of the agents/mechanisms responsible for the control of muscle mass through anabolic/catabolic pathways that contribute to maintenance of skeletal muscle homeostasis. Muscle atrophy is characterized by loss of muscle mass, leading to a reduction in functional capacity. Due the importance of this tissue in locomotion and many other physiological organism functions, the process of atrophy can infer to the individual the increase of the morbidity and loss of his quality of life, thus making of great importance to the understanding the molecular processes involved. In this work, leucine supplementation is sought to protect muscle mass during atrophy induced by immobilization through the inhibition of catabolic pathways. Leucine is an attractive target, since it is considered an anti-atrophic amino acid capable of regulating intracellular pathways involved in the synthesis and degradation of proteins. Therefore, we approached whether the anti-atrophic effect of leucine involves modulation of the GSK3-&#946;/&#946;-Catenin. Male Wistar rats, supplemented with leucine, had their left hind limb immobilized with the soleus muscle in a shortened position. Thereafter, the soleus muscle was removed, weighed and processed to evaluate an expression of genes and proteins by qPCR and Western blot, respectively. In addition, muscle cross-sections were performed in immunofluorescence assays for cell localization analysis. After 1 day of immobilization, activated GSK3-&#946; in the soleus muscle was observed, and leucine supplementation was able to block this effect. Although protein levels of &#946;-Catenin were unchanged by immobilization or by leucine supplementation, cell localization analysis showed a nuclear decrease of &#946;-Catenin caused by immobilization, however, leucine supplementation was able to increase levels of &#946;-Catenin in the nucleus. Confocal analysis confirmed nuclear translocation of &#946;-Catenin during leucine supplementation. We also analyzed the expression levels of NF-B by being potentially regulated by &#946;-Catenin, however, no changes were observed in NF-&#954;B protein levels between the groups. Thus, this study reveals &#946;-Catenin as a new key molecule involved in the anti-atrophic effects of leucine.

名古屋大学博士学位論文 学位の種類 : 博士(医学)(論文) 学位授与年月日:平成4年7月20日 淺田義正氏の博士論文として提出された

The respective kinetics of bacterial leucine aminopeptidase and [beta]-glucosidase activities were investigated to improve understanding of factors controlling activity and hydrolytic capacity in estuarine organic-poor sands. Depth distributions of enzyme activity and bulk organic matter content were determined in sediments of Aransas Bay and Copano Bay Texas, to investigate enzyme dynamics as related to the geochemical properties of the sediment. Vertical profiles of activity in sediment showed that the enzymes were more active at the surface and that the potential hydrolysis rate of leucine aminopeptidase was higher than that of [beta]-glucosidase. Vertical patterns of enzyme activity correlated (weakly) with variations in sediment organic matter (TOC, TN, and carbohydrates) content. Enrichments of sediment samples with monomeric organic compounds and inorganic nutrients did not affect leucine aminopeptidase and [beta]-glucosidase activities in short- and long-term incubations. Enzyme activity was independent of nutrient availability and suggested that microbial communities were not nutrient-limited. Time-course assays of bacterial hydrolysis of TOC, TN, and carbohydrates provided information about how substrate limitation may affect enzyme activity. Positive correlations between bulk TOC and TN content and enzyme activity indicated enzyme dependence on polymeric substrate content. Induction of enzyme activity after sediment enrichments with specific labile compounds confirmed the importance of available organic substrate to enzyme hydrolysis efficiency. A kinetic approach established the occurrence of enzyme inhibition and its effects on enzyme hydrolytic capacity. The addition of a specific-enzyme substrate to sediment samples modified enzyme parameters and indicated that a substrate-reversible type of inhibitor could reduce enzyme hydrolytic capacity. The addition of polyphenol, as a natural inhibitor of enzyme activity, to the sediment resulted in a concomitant reduction of leucine aminopeptidase activity and ammonium regeneration rate, and thus demonstrated a close coupling between enzyme activity and sediment ammonium regeneration. These research results demonstrate the dynamic nature of the hydrolytic enzymes, provide information about the mechanisms of induction and inhibition of activity, and demonstrate some implications of reducing the hydrolytic capacity to organic matter decomposition and nutrient regeneration rates. / text

Une cellule dendritique (CD) qui exprime le facteur de transcription GILZ durant l'apprêtement de l'antigène et sa présentation aux cellules effectrices, génère des lymphocytes T régulateurs (LTregs) CD25high Foxp3+ sécréteurs d'IL-10. La production de GILZ est dépendante de l'action des glucocorticoïdes, de l'IL-10 et du TGF-.Nous avons mis en évidence chez l'homme qu'une corticothérapie orale de 48h induit l'expression de GILZ dans les cellules présentatrices de l'antigène circulantes (CPAs) de sujets allergiques. Les CPAs isolées après la corticothérapie génèrent des LTregs CD25high Foxp3+ IL-10+ spécifiques de l'allergène.Nous également constaté in vitro que les mastocytes participent à l'activation des CDs au cours des réactions allergiques en régulant l'expression de GILZ. Les médiateurs d'origine mastocytaire, dont l'histamine, diminuent l'expression de GILZ dans les CDs et altèrent ainsi leur capacité à activer des LTregs. Nous avons identifié le mécanisme par lequel l'histamine diminue l'expression de GILZ dans les CDs humaines. L'histamine inhibe l'activité transcriptionnelle de Foxo3, un facteur de transcription régulant l'expression de GILZ. Enfin, nous avons démontré que des souris transgéniques qui surexpriment GILZ constitutivement dans les CDs sont protégées contre le développement de l'asthme allergique. L'ensemble de ces résultats permet d'envisager de nouvelles stratégies d'immunomodulation dans l'allergie, centrée sur la régulation de l'expression de GILZ dans les CDs.

Les protéines GILZ (Glucocorticoid-Induced Leucine Zipper) et TSC-22 (Transforming growth factor-beta Stimulated Clone-22) appartiennent à la famille de protéines TSC-22D (TSC-22 Domain). GILZ a été décrit précédemment comme étant induit au cours de la déprivation en interleukine-2 (IL-2) des lymphocytes de la lignée cellulaire CTLL-2, permettant ainsi de retarder leur apoptose. Le but de notre travail était de déterminer les rôles respectifs de GILZ et TSC-22 au cours de l'apoptose des cellules CTLL-2.Nos résultats ont permis de montrer que TSC-22 augmentait l'apoptose induite par la déprivation en IL-2 des cellules CTLL-2. Nous avons mis en évidence une augmentation de l'activation des caspases ainsi qu'une régulation positive de l'expression de BIM. Nous avons en outre montré que l'expression de GILZ, protéine anti-apoptotique, induite lors de la déprivation en IL-2, était régulée négativement en présence de TSC-22. Enfin, nous avons montré que l'expression de l'ARNm de gilz était régulée négativement par TSC-22, mais que la stabilité de son ARNm n'était pas modifiée.Notre travail a donc permis de montrer que TSC-22 accélère l'entrée en apoptose des lymphocytes T en régulant négativement l'expression de la protéine anti-apoptotique GILZ.

La leucine zipper de la rétine neurale (Nrl) joue un rôle central dans le développement et l'homéostasie des bâtonnets en activant I'expression de gènes tels que le photopigment Rhodopsine. Nrl est aussi associé à la Rétinite Pigmentaire, faisant ainsi de ce gène un modèle intéressant pour la compréhension des programmes contrôlant le développement et I'homéostasie des photorécepteurs.Ce travail de thèse vise à caractériser les mécanismes régulateurs de I'expression de Nr/ au cours du développement rétinien. L'électroporation in vivo de vecteurs rapporteurs dans des rétines de souris en développement, a révélé des séquences minimales de promoteur Nr/ nécessaires à une expression spécifique dans les photorécepteurs. Nous avons identifié RORI3 comme facteur requis pour cette expression, et montré que les facteurs OTX2, CRX et CREB s'accrochent aussi directement à des régions régulatrices particulières du promoteur. Nous avons construit un virus adéno-associé (AAV) contenant un promoteur minimal Nrl de 0.3 kb, et montré qu'il est adapté à la délivrance de gène spécifiquement dans les photorécepteurs.Nous avons montré que NRL, CRX et NR2E3, les régulateurs principaux de la Rhodopsine, ont une expression rythmique au cours de 24 h, et que l'expression cyclique de Nr/ peut être due à l'activation par RORp, un composant l'horloge circadienne. Enfin, nous avons identifié un nouveau facteur de transcription, NonO, au niveau de la région du promoteur proximal de la Rhodopsine, qui en combinaison avec NRL et CRX, active le promoteur de la Rhodopsine. L'invalidation de NonO au cours du développement rétinien a prouvé son implication pour le développement et I'homéostasie des bâtonnets.

DNA damage response pathways are a complicated network of proteins that function to remove and/or reverse DNA damage. Following genetic insult, a signal cascade is generated, which alerts the cell to the presence of damaged DNA. Once recognized, the damage is either removed or the damaged region is excised, and the original genetic sequence is restored. However, when these pathways are defective the cell is unable to effectively mediate the DNA damage response and the damage persists unrepaired. Thus, the proteins that maintain the DNA damage response pathway are critical in preserving genomic stability. One essential DNA repair protein is the Breast Cancer Associated gene, BRCA1. BRCA1 is essential for mediating the DNA damage response, facilitating DNA damage repair, and activating key cell cycle checkpoints. Moreover, mutations in BRCA1 lead to a higher incidence of breast and ovarian cancer, highlighting the importance of BRCA1 as a tumor suppressor. In an effort to better understand how BRCA1 carried out these functions, researchers sought to identify additional BRCA1 interacting proteins. This led to the identification of several proteins including the BRCA1 Associated C-terminal Helicase, BACH1. Due to the direct interaction of BACH1 with a region of BRCA1 essential for DNA repair and tumor suppression, it was speculated that BACH1 may help support these BRCA1 function(s). In fact, initial genetic screenings confirmed that mutations in BACH1 correlated not only with hereditary breast cancer, but also with defects in DNA damage repair processes. The initial correlation between BACH1 and cancer predisposition was further confirmed when mutations in BACH1 were identified in the cancer syndrome Fanconi anemia (FA) (complementation group FA-J), thus giving BACH1 its new name FANCJ. These findings supported a previously established link between the FA and BRCA pathways and between FA and DNA repair. In particular, we demonstrated that similar to other FA/BRCA proteins, suppression of FANCJ lead to a substantial decrease in homologous recombination and enhanced both the cellular sensitivity to DNA interstrand cross-linking agents and chromosomal instability. What remained unknown was specifically how FANCJ functioned and whether these functions were dependent on its interaction with BRCA1 or other associated partners. In fact, we identified that FANCJ interacted directly with the MMR protein MLH1. Moreover, we found that the FANCJ/BRCA1 interaction was not required to correct the cellular defects in FA-J cells, but rather that the FANCJ/MLH1 interaction was required. Although both the FA/BRCA and MMR pathways undoubtedly mediate the DNA damage response, there was no evidence to suggest that these pathways were linked, until recently. Our findings not only indicate a physical link between these pathways by protein-protein interaction, but also demonstrated a functional link.