Ornithine-$ delta$-biaminotransferase (OAT) is a mitochondrial matrix a enzyme which catalyzes the reversible conversion of ornithine to glutamic semialdehyde. In the retinoblastoma cell line Y79, the 5$ sp prime$ UTR of the OAT mRNA is alternatively-spliced. Previous work has shown that the two alternatively-spliced 5$ sp prime$ UTR are translated with different efficiencies. / The alternatively-spliced 5$ sp prime$ UTRs were isolated and subcloned. Expression vectors were constructed containing each of the OAT 5$ sp prime$ UTRs and the human growth hormone as a reporter gene. Transient transfection expression analysis in COS-1 cells confirmed that the 5$ sp prime$ UTRs conferred differential expression on the reporter gene. The shorter 5$ sp prime$ UTR was expressed at a level 3-fold higher than that of the longer 5$ sp prime$ UTR. / RNA mobility shift assays were performed to determine whether a transacting factor was interacting with one of the alternatively-spliced 5$ sp prime$ UTRs. A binding activity was detected that bound to the longer, exon 2-containing 5$ sp prime$ UTR with a significantly higher affinity than the shorter 5$ sp prime$ UTR. Competition studies demonstrated that this binding activity was specific for the longer 5$ sp prime$ UTR.

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Ornithine Aminotransferase (OAT) is expressed at high levels in liver, kidney, and retina. Tissue-specific regulation of OAT has been described for rat kidney and liver. To characterize OAT regulation in retinal lines, we studied OAT synthesis in retinoblastomas RB355 and Y79. / OAT transcription and mRNA levels in the two lines were similar, but 3-fold greater immunoreactive OAT protein and enzyme activity were observed for Y79. Characterization of polysome-associated OAT mRNAs indicated that they were translated less efficiently, due to decreased initiation, in RB355. Initiation factor eIF-4E mRNA and protein were reduced in RB355; eIF-4E overexpression in RB355 increased OAT translation and OAT protein to the level observed in Y79. / Estrogen and thyroid hormone increased OAT expression in both strains. Estrogen increased translational initiation, with no effect on transcription, whereas thyroid hormone was primarily a transcriptional activator. / An alternatively-spliced OAT mRNA was identified in these retinoblastomas; this mRNA was poorly translated and was not affected by eIF-4E overexpression, estrogen, or thyroid hormone. / This study has demonstrated several post-transcriptional regulatory mechanisms for OAT in retinal tissue.

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Les acides aminés sont des molécules importantes pour la synthèse des protéines, la fourniture de substrats énergétiques ou la production de facteurs de régulation de l'activité cellulaire. L'ornithine aminotransférase {OAT} est une enzyme clef du métabolisme des acides aminés qui relie l'arginine, un acide aminé indispensable à la synthèse protéique au cours de la croissance, et le glutamate, la glutamine et l'a-cétoglutarate, qui peuvent être transformées en glucose. Nous nous sommes intéressés à la caractérisation biochimique de l'activité de l'OAT et à ses variations chez la souris, en utilisant deux contraintes physiologiques, le développement postnatal et l'exposition au froid. Au cours d'une première étude, nous avons caractérisé une activité OAT fonctionnelle dans tous les organes étudiés. Nous avons démontré l'existence d'un dimorphisme sexuel de l'activité, en faveur des femelles, dans la plupart des organes. Dans une seconde étude, nous avons montré des variations organe et sexe spécifiques de l'activité OAT au cours du développement postnatal en réponse aux besoins spécifiques en acides aminés : une régulation de l'expression de l'OAT par la testostérone et le changement de régime alimentaire lors du sevrage ; une contribution potentielle importante et insoupçonnée du muscle squelettique au métabolisme de l'ornithine. Enfin, nous avons mis en évidence que l'exposition au froid augmente ou diminue l'activité OAT de façon tissu-spécifique en liaison avec les besoins énergétiques de l'organisme. Nos résultats montrent que l'activité OAT est modulée dans les différents organes en fonction des besoins en acides aminés et en substrats. Cela suggère un rôle important du métabolisme des acides aminés dans les réponses adaptatives des organismes à leur environnement / Amino acids are important molecules for protein synthesis and energetic substrates supply. Ornithine aminotransferase {OAT} is a key enzyme of amino acids metabolism linking arginine, an essential amino acid for protein synthesis during growth, and glutamate, glutamine and alpha-ketoglutarate which could be converted into glucose. We principally focused on biochemical characterization of OAT activity and its variations in the mouse using two physiological constraints, postnatal development and cold exposition. During a first study, we characterized a functional OAT activity in all the males and females mice organs studied. We showed the existence of a sexual dimorphism of OAT activity in almost all the organs studied, with a higher OAT activity in females. ln a second study, we showed that the variations of OAT activity were organ and sex specific during the postnatal development in response to the specific needs in amino acids during the growth. We linked these variations to the regulation of OAT expression by testosterone and the change of the diet during weaning. We demonstrated an important and unsuspected contribution of skeletal muscles in pups' ornithine metabolism. Finally, we showed that cold exposition modulates OAT activity in a tissue specific way, in relation with energetic needs and the multiple roles of the substrates generated by the metabolic pathways of OAT. Our results show that in the different organs, OAT activity is modulated in function of amino acids needs, energetic substrates or detoxification needs of the organism during the different metabolic constraints. This is suggesting an important role for the amino acids metabolism in adaptive responses of organisms to their environment