The mechanism for protein kinase C inhibition of androgen production and 17alpha-hydroxylase expression in a theca cell tumor model.

Abstract

In polycystic ovary syndrome (PCOS), there is increased formation of androgens by thecal cells. Moreover, PCOS ovaries have been shown to have decreased levels of c-fos transcription factor. We hypothesize that c-fos expression inhibits 17alpha-hydroxylase 17,20 lyase (CYP17) activity in the human ovary, and its decreased expression seen in PCOS may lead to elevated CYP17 transcription, resulting in increased androgen production. Our objective was to define the role of the activator protein-1 transcription factors, namely c-fos, in the regulation of CYP17 expression in theca cells. Human ovarian thecal-like tumor cells were used for all experiments. The following techniques were used: steroid quantification, mRNA extraction, microarray analysis, transfection, small interfering RNA, and immunohistochemistry. Stimulation of human ovarian thecal-like tumor cells with the protein kinase A pathway activator forskolin resulted in stimulation of C19 androgen production. In contrast, treatment with the protein kinase C pathway activator tetradecanoylphorbol acetate (TPA) resulted in decreased androgen production with a shift toward C21 progesterone production. TPA also led to complete inhibition of CYP17. Microarray data showed a 37-fold increase in c-fos after treatment with TPA. Transfection with steroidogenic factor 1 resulted in an increase in CYP17 promoter activity, which was significantly inhibited in the presence of c-fos. c-fos gene silencing led to an increase in CYP17 mRNA levels. Immunohistochemical staining for c-fos in ovaries demonstrated strong staining in granulosa cells, but not theca. The activator protein-1 transcription factor c-fos plays a role in the inhibition of CYP17 expression. The decreased levels of c-fos expression in polycystic ovaries may be responsible for increased CYP17 levels in PCOS.