Molecular mechanisms of oocyte and sperm maturation-inducing hormone

本研究旨在探讨鱼类性腺中卵母细胞和精子成熟诱导激素（17 α，20 β-二羟基-4-异戊烯-3-酮）的分子机制。17 α，20 β-二羟基-4-异戊烯-3-酮是通过类固醇生成酶20 β-羟基类固醇脱氢酶（20 β-HSD）的作用从17 α-羟孕酮生物合成的。本研究首先克隆了香鱼（Plecoglossus altivelis）和青鳉（Oryzias latipes）20 β-HSD基因的cDNA克隆，以香鱼20 β-HSD cDNA克隆为探针，从虹鳟（Oncorhynchus mykiss）卵巢滤泡中分离到两个不同的20 β-HSD样cDNA。cDNA和基因组DNA分析表明，这两个20 β-HSDs由两个不同的基因编码。北方杂交和RT-PCR分析表明，这两个基因在不同的组织中表达。两种cDNA在大肠杆菌中的体外表达使我们能够确定两种cDNA之一（A型）的产物具有将17 α-羟孕酮转化为17 α，20 β-二羟基-4-异戊烯-3-酮的能力。20 β-HSD基因的表达显着增强时，颗粒细胞分离的卵黄发生后卵泡与促性腺激素孵育。此外，重要的是要注意，A型基因，而不是B型基因，是可诱导的颗粒细胞促性腺激素刺激。由于20 β-HSD A型表现出20 β-HSD活性，20 β-HSD A型基因的增强表达增加了17 α-羟孕酮向17 α，20 β-二羟基-4-异戊烯-3-酮的转化。这些数据表明20 β-HSD的表达受促性腺激素控制。促性腺激素对20 β-HSD基因启动子区的影响将为进一步了解类固醇和蛋白激素对类固醇生成酶的调节提供帮助。

This study was designed to investigate the molecular mechanism of oocyte and sperm-maturation inducing hormone (l7alpha, 20beta-dihydroxy-4-pregnen-3-one) in fish gonads. l7alpha, 20beta-Dihydroxy-4-pregnen-3-one is biosynthesized by the action of a steroidogenic enzyme, 20beta-hydroxysteroid dehydrogenase (20beta-HSD) from 17alpha-hydroxyprogesterone. We first isolated cDNA clones encoding ayu (Plecoglossus altivelis) and medaka (Oryzias latipes) 20beta-HSD.Using the ayu 20beta-HSD cDNA clone as a probe, we isolated two different 2Obeta-HSD like cDNAs from rainbow trout (Oncorhynchus mykiss) ovarian follicles. cDNA and genomic DNA analysis show that the two 20beta-HSDs are encoded in two different genes. Northern blot analysis and RT-PCR show that these two are expressed in different tissues. In vitro expression of the two cDNAs in E.colienabled us to establish that the product of one (type A) of the two cDNAs has the ability to convert 17alpha-hydroxyprogesterone to 17alpha, 2Obeta-dihydroxy-4-pregnen-3-one. 20beta-HSD gene expression was markedly enhanced when granulosa cells isolated from postvitellogenic follicles were incubated with gonadotropin. Furthermore, it is of importance to note that type A gene, but not type B gene, is inducible in granulosa cells by gonadotropin stimulation. Since 20beta-HSD type A exhibits 20beta-HSD activity, the enhanced expression of 20beta-HSD type A gene increases the conversion of 17alpha-hydroxyprogesterone to 17alpha, 2Obeta-dihydroxy-4-pregnen-3-one. These data demonstrate that the expression of 20beta-HSD is controlled by gonadotropin. The effects of gonadotropin on the promoter regions of 20beta-HSD genes will provide further understanding of steroid and protein hormone modulation of steroidogenic enzymes.