Ethier Jean-François, Farnworth Paul G, Findlay Jock K, Ooi Guck T
Prince Henry's Institute of Medical Research, Clayton, Victoria 3168, Australia.
Mol Endocrinol. 2002 Dec;16(12):2754-63. doi: 10.1210/me.2002-0014.
Activin stimulates expression of GnRH receptor (GnRHR) and FSH beta-subunit in gonadotropes. Inhibin antagonizes activin actions on the gonadotropes, but its molecular mechanism of action remains poorly understood. It has been suggested that inhibin exerts its antagonistic effects by competing with activin for the binding of the activin receptor complex. Betaglycan has recently been identified as an inhibin-binding accessory protein in this process. Because both inhibin and TGFbeta bind betaglycan, we examined whether TGFbeta can modify inhibin's antagonism of activin-induced transcription in gonadotrope cells. Two activin-responsive reporter constructs were used, the first containing 5.5 kb of the ovine FSHbeta promoter (oFSHbetaluc), and the second containing three copies of the activin-responsive sequence of the GnRHR promoter (3XGRAS-PRL-lux). These constructs were transfected into the gonadotrope cell line LbetaT2. The oFSHbetaluc and 3XGRAS-PRL-lux activities stimulated by 0.5 nM activin A were decreased by up to 50% in a dose-dependent manner by inhibin A. TGFbeta(1) and TGFbeta(2) (0-4 nM), alone or in the presence of activin A, did not significantly affect the promoter elements. However, with increasing doses of TGFbeta(1) or TGFbeta(2), inhibin A antagonism of activin A activity was partly or completely reversed. Competition studies with radiolabeled inhibin A showed that TGFbeta(1) and TGFbeta(2) competed with [(125)I]inhibin for the binding to LbetaT2 cells (IC(50) = 280 pM and 72 pM, respectively). Immunoprecipitation studies of [(125)I]inhibin A cross-linked receptor complexes confirmed that TGFbeta(1) and TGFbeta(2) competed with inhibin A for the binding of betaglycan. These results suggest that TGFbeta competition with inhibin for binding to betaglycan interferes with inhibin's suppression of activin-induced FSHbeta and GnRHR promoters in LbetaT2 cells. We propose that under certain circumstances, TGFbeta may facilitate activin biological activity by hindering the access of inhibin to its coreceptor betaglycan.
激活素可刺激促性腺激素细胞中促性腺激素释放激素受体（GnRHR）和促卵泡激素β亚基的表达。抑制素可拮抗激活素对促性腺激素细胞的作用，但其分子作用机制仍知之甚少。有人提出，抑制素通过与激活素竞争激活素受体复合物的结合来发挥其拮抗作用。β聚糖最近被确定为这一过程中与抑制素结合的辅助蛋白。由于抑制素和转化生长因子β（TGFβ）都能结合β聚糖，我们研究了TGFβ是否能改变抑制素对激活素诱导的促性腺激素细胞转录的拮抗作用。我们使用了两种激活素反应性报告基因构建体，第一种含有5.5 kb的绵羊促卵泡激素β启动子（oFSHbetaluc），第二种含有促性腺激素释放激素受体启动子激活素反应序列的三个拷贝（3XGRAS-PRL-lux）。将这些构建体转染到促性腺激素细胞系LbetaT2中。0.5 nM激活素A刺激的oFSHbetaluc和3XGRAS-PRL-lux活性，被抑制素A以剂量依赖性方式降低了多达50%。单独或在激活素A存在的情况下，TGFβ（1）和TGFβ（2）（0 - 4 nM）对启动子元件没有显著影响。然而，随着TGFβ（1）或TGFβ（2）剂量的增加，抑制素A对激活素A活性的拮抗作用部分或完全被逆转。用放射性标记的抑制素A进行的竞争研究表明，TGFβ（1）和TGFβ（2）与[¹²⁵I]抑制素竞争与LbetaT2细胞的结合（IC₅₀分别为280 pM和72 pM）。对[¹²⁵I]抑制素A交联受体复合物的免疫沉淀研究证实，TGFβ（1）和TGFβ（2）与抑制素A竞争β聚糖的结合。这些结果表明，TGFβ与抑制素竞争结合β聚糖会干扰抑制素对激活素诱导的LbetaT2细胞中促卵泡激素β和促性腺激素释放激素受体启动子的抑制作用。我们提出，在某些情况下，TGFβ可能通过阻碍抑制素与其共受体β聚糖的结合来促进激活素的生物学活性。
