Identification and biosynthesis of sperm receptor in quail oocyte

鸟类的卵母细胞被称为卵黄膜的半透明薄膜所包围。卵黄膜由三层组成，即内、外层和连续膜。在排卵时，卵母细胞周围只有内层卵黄膜。我们发现，排卵前卵泡的颗粒细胞含有33 kDa糖蛋白(ZPC)，这是鹌鹑卵内层的主要成分之一。尽管ZPC在受精过程中具有重要的生理意义，但在排卵周期中颗粒细胞中ZPC的产生调控细节尚不清楚。为了探讨ZPC产生的调控机制，我们用Western印迹和Northern印迹分析了ZPC在排卵周期中的表达。此外，我们还培养了含有或不含各种激素和生长因子的颗粒细胞，以研究内分泌和旁分泌对ZPC产生的调控。体内免疫印迹和Northern印迹分析表明，ZPC在生长卵泡颗粒细胞中的表达随着卵泡成熟而增加，在最大卵泡中活性最高。为探讨不同激素和生长因子对ZPC生成的影响，取三个最大卵泡的颗粒细胞，分别加入FSH、LH、EGF、IGF-I、IGF-II、TGF-α、TGF-β、孕酮、雌二醇或睾酮培养72小时。FSH对培养的第三大卵泡颗粒细胞ZPC的刺激作用最为显著，其他激素或生长因子对ZPC的刺激作用不明显。这些数据表明，鹌鹑颗粒细胞产生ZPC以响应FSH。

The avian oocyte is bounded by the thin translucent membrane called vitelline membrane. The vitelline membrane consists of three layers, the inner and outer layers and the continuous membrane. Only the inner layer of vitelline membrane surrounds the oocyte at the time of ovulation. We have found that the granulosa cells of the preovulatory follicles contain 33 kDa glycoprotein (ZPC) one of the major components of the inner layer in quail egg. Though the physiological importance of ZPC for fertilization was pointed out as the candidate of "sperm receptor" in quail, the details in the regulation of ZPC production in granulosa cells during ovulation cycle remains obscure. To investigate the regulatory mechanism of ZPC production we have studied the expression of ZPC during the ovulation cycle by Western blot and Northern blot analyses. Moreover, we cultured the granulosa cells with or without various hormones and growth factors to study the endocrine and paracrine control of ZPC production. From the Western blot and Northern blot analyses in in-vivo study, the expression of ZPC in the granulosa cells of growing follicles increased during follicular maturation with the highest activity in the largest follicle. To investigate the effects of various hormones and growth factors on the ZPC production, the granulosa cells obtained from the three largest follicles were cultured for 72 hr with FSH, LH, EGF, IGF- I, IGF- II, TGF- alpha, TGF- beta, progesterone, estradiol, or testosterone. The most prominent effects on stimulation of ZPC production was observed in the granulosa cells isolated from the third largest follicle cultured by the addition of FSH.Other hormones or growth factors caused no significant increase. These data illustrate that quail granulosa cells produce ZPC in response to FSH.