Molecular mechanisms of activin B-induced spermatogenesis

本研究的目的是了解促性腺激素诱导日本鳗鲡精子发生的分子机制。将含有激活素β B亚基cDNA的表达载体导入哺乳动物CHO细胞后，这些细胞分泌22 kDa的蛋白到培养基中。纯化的重组蛋白（鳗鱼激活素B）具有红系分化因子活性。添加鳗鱼激活素B到培养基中诱导精原细胞增殖，产生晚期B型精原细胞，在15天内以相同的方式11-酮睾酮。综上所述，支持细胞在促性腺激素和11-酮睾酮的影响下产生的激活素B在鱼类睾丸精子发生的调节中起着重要的作用。我们还从鳗鱼睾丸cDNA文库中克隆了编码细胞激活素I型和II型受体的cDNA。北方印迹分析表明，激活素I型和II型受体的mRNA转录本在注射促性腺激素之前的鳗鱼睾丸中表达。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，为激活素B可能直接作用于生殖细胞提供了证据。从鳗鱼睾丸中克隆了几个编码G1/S周期蛋白（A1、A2、D1、D2、E1、E2）的cDNA。这些G1/S细胞周期蛋白转录本都不存在于激素诱导精子发生前的鳗鱼睾丸。在注射促性腺激素后的睾丸中观察到这些细胞周期蛋白的表达顺序如下。：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).