Molecular mechanisms of activin B-induced spermatogenesis

本研究旨在了解促性腺激素诱导鳗鱼精子发生的分子机制。将含有激活素BETAB亚单位基因的表达载体导入哺乳动物CHO细胞后，可分泌出一个22 kDa的蛋白。纯化的重组蛋白(鳗鱼激活素B)具有红系分化因子活性。在培养液中加入鳗鱼激活素B，在15天内以与11-酮睾酮相同的方式诱导精原细胞增殖，产生晚期B型精原细胞。综上所述，支持细胞在促性腺激素和11-酮睾酮的作用下产生的激活素B在鱼类睾丸的生精调控中起着重要作用。我们还从鳗鱼睾丸的cDNA文库中克隆了编码细胞激活素I和II受体的cDNAs。Northern印迹分析表明，在注射促性腺激素之前，激活素I和II受体的mRNA转录产物在鳗鱼的睾丸中表达。原位杂交显示这些受体定位于生殖细胞(精原细胞和精母细胞)和支持细胞，这为激活素B可能直接作用于生殖细胞提供了证据。已从鳗鱼精巢中克隆了几个编码G1/S细胞周期蛋白的基因(A1、A2、D1、D2、E1、E2)。在激素诱导生精之前，这些G1/S细胞周期蛋白转录本均不存在于鳗鱼的睾丸中。注射促性腺激素后，这些细胞周期蛋白在睾丸中的表达顺序如下。：E1(G1/S)*A2，B1，B2*A1(精母细胞)。

The purpose of this study is to understand the molecular mechanisms of gonadotropin-induced spermatogenesis in the Japanese eel (Anguilla japonica). With introduction of the expression vector containing activin betaB subunit cDNA into mammalian CHO cells, these cells secreted a 22kDa protein into the medium. Purified recombinant protein (eel activin B) had erythroid differentiation factor activity. Addition of eel activin B to the culture medium induced proliferation of spermatogonia, producing late type B spermatogonia, within 15 days in the same manner of 11-ketotestosterone. Taken together, it is concluded that activin B produced by Sertoli cells under the influence of gonadotropin and 11-ketotestosterone plays an important role in the regulation of spermatogenesis in fish testes. We have also cloned cDNAs encoding ell activin type I and II receptors from an eel testis cDNA library. Northern blot analysis reveals that mRNA transcripts of activin type I and II receptors are expressed in testes of eels prior to gonadotropin injection. In situ hybridization shows that these receptors are localized in germ cells (spermatogonia and spermatocytes) and Sertoli cells, providing evidence that activin B may act directly on the germ cells. Several cDNAs encoding G1/S cyclins (A1, A2, D1, D2, E1, E2) have been cloned from eel testes. None of these G1/S cyclin transcripts were present in testes of eels before hormonal induction of spermatogenesis. The following sequence of expression of these cyclins was observed in testes after gonadotropin injection. : E1 (G1/S) *A2, B1, B2*A1 (spermatocytes).