SHORT-CHAIN DEHYDROGENASES IN RETINOL/STEROL METABOLISM

描述(申请人提供)：维生素A衍生物全反式维甲酸调节超过530个不同基因的表达。因此，胚胎发育期间的维甲酸水平在空间和时间上都得到了精确的控制。然而，这种调控的分子机制尚不完全清楚。该项目的长期目标是确定短链脱氢酶/还原酶(SDR)在调节维甲酸生物合成中的作用。最近，我们在蛙类中发现了SDR超家族的一个新成员，rdhe 2，它是一种高度活跃的全反式视黄醇脱氢酶，对非洲爪哇的胚胎发育至关重要。重要的是，在哺乳动物中似乎有一个与青蛙rdhe2功能相当的基因，它表现出全反式视黄醇脱氢酶活性，并在早期胚胎发育中表达。我们认为，这种新的酶，命名为RDH-E2S，在哺乳动物胚胎发育过程中对维甲酸的生物合成是必不可少的，可能在成年后也是如此。为了验证这一假设，我们将利用转基因小鼠模型(特定目标1)来表征哺乳动物RDH-E2S的催化特性，并确定其在体内对维甲酸生物合成的贡献。我们的初步研究表明，沉默视网膜短链脱氢酶/还原酶1(RetSDR1)基因在人类细胞中的表达导致视黄酸和视黄醛水平显著增加，这转化为视黄酸反应基因的显著上调。这一发现表明，视黄酸的生物合成速率是由视黄醇脱氢酶和视黄醛还原酶的相对活性决定的，这两种酶共同控制着视黄酸前体视黄醛的水平。为了验证这一假说，我们建议使用转基因小鼠、人类皮肤器官培养和非洲爪哇早期胚胎发育的体外模型来表征retSDR1的催化特性，并确定其在体内调节维甲酸水平中的作用(特定目标2)。这些研究将通过提供有关维甲酸调节系统潜在重要组成部分的作用的新信息，填补我们对维甲酸维持稳态机制的理解的空白。这些研究的结果将对了解与维甲酸稳态紊乱相关的疾病的病理生理学，如胎儿酒精综合征、酒精性肝病、癌症发生和糖尿病具有重要意义。

DESCRIPTION (provided by applicant): Vitamin A derivative all-trans-retinoic acid regulates the expression of over 530 different genes. Consequently, the levels of retinoic acid during embryogenesis are controlled in a spatially and temporally precise manner. However, the molecular mechanisms underlying this regulation are not yet fully understood. The long-term objective of this project is to determine the role of short-chain dehydrogenases/reductases (SDRs) in the regulation of retinoic acid biosynthesis in health and disease. Recently, we have identified a new member of the SDR superfamily of proteins in frogs, rdhe 2, that is highly active as an all-trans-retinol dehydrogenase and is critical for embryonic development in Xenopus laevis. Importantly, there appears to be a functional equivalent of the frog rdhe2 in mammals, which exhibits an all-trans-retinol dehydrogenase activity and is expressed during early embryonic development. We propose that this novel enzyme, named RDH-E2S, is essential for retinoic acid biosynthesis in mammals during embryogenesis and, possibly, in adulthood. To test this hypothesis, we will characterize the catalytic properties of mammalian RDH-E2S and determine its contribution to retinoic acid biosynthesis in vivo using genetically modified mouse model (Specific Aim 1). Our preliminary studies indicate that silencing of retina short-chain dehydrogenase/reductase 1 (retSDR1) gene expression in human cells results in significant increase in the levels of both retinoic acid and retinaldehyde, which translates into dramatic upregulation of retinoic acid-responsive genes. This finding suggests that the rate of retinoic acid biosynthesis is determined by the relative activities of retinol dehydrogenases and retinaldehyde reductases, which together control the levels of retinoic acid precursor, retinaldehyde. To test this hypothesis, we propose to characterize the catalytic properties of retSDR1 and to determine its role in the regulation of retinoic acid levels in vivo using genetically modified mice, human skin organ culture, and Xenopus laevis in vitro model of early embryonic development (Specific Aim 2). These studies will fill the gaps in our understanding of the mechanisms responsible for the maintenance of retinoic acid homeostasis by providing new information regarding the roles of potentially important components of the retinoid regulatory system. The results of these studies will be important for understanding the pathophysiology of disorders associated with disruptions of retinoid homeostasis, such as fetal alcohol syndrome, alcoholic liver disease, carcinogenesis, and diabetes.