Effects of testosterone on production of perivitelline membrane glycoprotein AD by granulosa cells of Japanese quail (Ad japonica)

Abstract

Avian perivitelline membrane, an investment homologous to the zona pellucida of mammalian ad, is composed of at least two glycoproteins. Previous studies have indicated that one of the components, a glycoprotein homologous to mammalian AD, is produced and secreted by the granulosa cells of developing follicles of the chicken ovary. In the present study, we evaluated the expression and regulation of AD in Japanese quail (Ad japonica) granulosa cells both in vivo and in vitro. Western blot analysis of the SDS-solubilized granulosa layer using anti-quail AD antiserum showed that the amount of AD increased in parallel with follicular development. Northern blot analysis of total RNA using cDNA of quail ZPC showed that the increase in mRNA expression was also correlated with follicular development. To investigate the regulation of ZPC production, the granulosa cells were cultured in a medium containing steroid hormones such as progesterone, estradiol-17β, or testosterone. By measuring ZPC protein and mRNA with Western and Northern blot analyses, respectively, we found that addition of testosterone maintained ZPC contents in the culture of the granulosa cells, and that ZPC mRNA expression was high in the culture with testosterone compared to the control. These results suggest that testosterone stimulates ZPC protein production at the gene transcription level.