SHORT CHAIN DEHYDROGENASES IN RETINOL/STEROL METABOLISM

维甲酸和类固醇激素调节从形态发生到生殖的各种生理过程。 这两种激素的生物合成都涉及它们相应的前体氧化和还原成生物活性形式。最近的数据从这个和其他实验室表明，哺乳动物组织中含有一组相关的酶，能够利用在体外类维生素A和类固醇醇和醛作为底物。 这些酶具有超过40%的序列同一性，属于短链淀粉酶/还原酶超家族。 主要假设是：（1）这组结构相关的酶参与体内类维生素A和甾体激素的生物合成;（2）单个视黄醇/甾醇脱氢酶在人体组织中表现出不同的组织特异性表达模式，这决定了它们对每个代谢途径的贡献;和（3）这些微粒体脱氢酶的活性位点面向胞质溶胶，其中NAD+依赖性酶在氧化方向上起作用，而NADP+依赖性酶在还原方向上起作用。 共同酶的存在应提供联合调节类维生素A和类固醇信号通路的手段。 这一假设与维甲酸显著降低血清双氢睾酮水平和前列腺癌组织中维甲酸水平降低的观察结果一致。 本建议的总体目标是确定本实验室最近鉴定的人微粒体视黄醇/甾醇脱氢酶在细胞中类维生素A和类固醇代谢中的作用。 本提案的具体目的是：（1）开发具有催化活性的纯化重组视黄醇/固醇脱氢酶的制备方法;（2）表征人视黄醇/固醇脱氢酶的底物特异性和催化性质;（3）确定人体组织中视黄醇/固醇脱氢酶的表达模式;和4）确定人端粒酶在微粒体膜中跨膜插入的拓扑结构。这些目标的实现将使我们能够了解这组新的短链脱氢酶在健康和疾病中类维生素A和类固醇代谢中的作用。

Retinoid and steroid hormones regulate a variety of physiological processes from morphogenesis to reproduction. Biosynthesis of both types of hormones involves oxidation and reduction of their corresponding precursors to the biologically active forms. Recent data from this and other laboratories suggest that mammalian tissues contain a group of elated enzymes capable of utilizing in vitro both retinoid and steroid alcohols and aldehydes as substrates. These enzymes share more than 40 percent sequence identity and belong to the superfamily of short-chain dehydrogenases/reductases. The main hypotheses of this proposal are that: (1) this group of structurally related enzymes is involved in biosynthesis of both retinoid and steroid hormones in vivo; (2) individual retinol/sterol dehydrogenases exhibit different tissue-specific expression patterns in human tissues, which determines their contribution to each metabolic pathway; and (3) the active site of these microsomal dehydrogenases faces the cytosol, where the NAD+-dependent enzymes function in the oxidative direction, and the NADP+-dependent enzymes function in the reductive direction. The existence of common enzymes should provide the means for joint regulation of retinoid and steroid signaling pathways. This hypothesis is consistent with the observations that retinoic acid significantly decreases serum levels of dihydrotestosterone and that the levels of retinoic acid are decreased in prostate carcinoma tissue. The overall objective of this proposal is to determine the role of the human microsomal retinol/sterol dehydrogenases, recently identified by this laboratory, in retinoid and steroid metabolism in the cells. The specific aims of this proposal are to: (1) develop a procedure for preparation of catalytically active purified recombinant retinol/sterol dehydrogenases; (2) characterize the substrate specificity and catalytic properties of the human retinol/sterol dehydrogenases; (3) determine the expression patterns of retinol/sterol dehydrogenases in human tissues; and 4) determine the topology of transmembrane insertion of human dehydrogenases in the microsomal membrane. Accomplishment of these objectives will allow us to gain understanding of the role of this new group of short-chain dehydrogenases in retinoid and steroid metabolism in health and disease.