Short-Chain Dehydrogenases in Retinol/Sterol Metabolism

描述（由申请人提供）：维甲酸在多种胎儿和成人组织的分化和发育中起重要作用。视黄醇生产维甲酸分为两个步骤：首先，视黄醇通过可逆氧化转化为视黄醛，然后视黄醛被不可逆氧化为视黄酸。视黄醇的氧化是视黄酸生产的限速步骤，决定了视黄醇生物合成视黄酸的总速率。在之前的资助期内，我们已经确定了人类短链脱氢酶/还原酶（sdr）超家族蛋白的两个亚家族，它们对类维甲酸有活性。当在完整细胞中表达时，rodh样sdr赋予将视黄醇氧化为视黄醛的能力，而ralr1样酶赋予将视黄醛还原为视黄醇的能力。基于这些观察结果，我们提出两组人类sdr通过调节视黄醇和视黄醛之间的平衡，从而调节视黄酸的生物合成速率，从而促进人体组织中类视黄酸的稳态。为了验证我们的假设，我们建议在存在和不存在RoDH-和ralr1样sdr的情况下表征人类和动物组织中的类视黄醇代谢。第一个特定目的下的实验将验证一个假设，即在人器官型皮肤筏培养中，沉默rodh样SDR基因表达会导致视黄醇氧化为视黄醇的速率降低，而沉默RalR1样SDR基因表达会导致视黄醇转化为视黄醇的速率降低。第二个特定目标下的实验将用RalR1敲除小鼠模型的体内研究来补充人类皮肤支架的离体研究，并将验证RalR1对于小鼠组织中视黄醛还原为视黄醇至关重要的假设。由于RalR1的小鼠同源物在体外对视黄醛以外的中链醛具有高度活性，因此我们还将测试RalR1是否有助于中链醛的还原。据报道，类维生素a稳态在癌细胞、酒精性肝病和胎儿酒精综合征中被破坏。所提出的研究将更好地了解在各种病理状态下类视黄醇代谢失调的分子基础。

DESCRIPTION (provided by applicant): Retinoic acid plays an important role in differentiation and development of a wide variety of fetal and adult tissues. Retinoic acid is produced from retinol in two steps: first, retinol is converted to retinaldehyde by a reversible oxidation, and then retinaldehyde is irreversibly oxidized to retinoic acid. The oxidation of retinol is the rate-limiting step in retinoic acid production that determines the overall rate of retinoic acid biosynthesis from retinol. During the previous funding period, we have identified two subfamilies of the human short-chain dehydrogenase/reductase (SDRs) superfamily of proteins that are active toward retinoids. When expressed in intact cells, RoDH-like SDRs confer the ability to oxidize retinol to retinaldehyde, whereas RalR1-like enzymes confer the ability to reduce retinaldehyde back to retinol. Based on these observations, we propose that both groups of human SDRs contribute to retinoid homeostasis in human tissues by regulating the equilibrium between retinol and retinaldehyde, and thereby, regulating the rate of retinoic acid biosynthesis. To test our hypothesis, we propose to characterize retinoid metabolism in human and animal tissues in the presence and in the absence of RoDH- and RalR1-like SDRs. Experiments under the first specific aim will test a hypothesis that silencing of RoDH-like SDR gene expression in human organotypic skin raft culture results in a decreased rate of retinol oxidation to retinaldehyde, whereas silencing of RalR1- like SDR gene expression results in a decreased rate of retinaldehyde conversion to retinol. Experiments under the second specific aim will complement the ex vivo studies in human skin rafts with in vivo studies in RalR1 knockout mouse model and will test a hypothesis that RalR1 is essential for the reduction of retinaldehyde to retinol in mouse tissues. Because in vitro the mouse ortholog of RalR1 is highly active toward medium-chain aldehydes in addition to retinaldehydes, we will also test whether RalR1 contributes to the reduction of medium-chain aldehydes. It has been reported that retinoid homeostasis is disrupted in cancer cells, in alcoholic liver disease and in fetal alcohol syndrome. The proposed studies will provide a better understanding of the molecular bases underlying disregulation of retinoid metabolism in various pathological states.