Androgens Regulate Ovarian Gene Expression Through Modulation of Ezh2 Expression and Activity

Abstract

A substantial amount of evidence suggests that androgen signaling through classical androgen receptors is critical for both normal and pathologic ovarian physiology. Specifically, we and others have shown that, in mouse granulosa cells, androgen actions through both extranuclear and nuclear androgen receptor signaling are critical for normal follicle development and ovulation. Here, we show that androgens through the PI3K/Akt pathway rapidly (within minutes) phosphorylate and inhibit activity of the Polycomb group protein enhancer of zeste homolog 2 (Ezh2). Over the course of 24 to 48 hours, androgens then induce expression of the microRNA miR-101, which targets Ezh2 messenger RNA (mRNA), leading to a nearly complete loss of Ezh2 protein expression. This long-term androgen-induced loss of Ezh2 actions ultimately results in sustained reduction of the H3K27me3-repressive mark in the promoter region of the Runt-related transcription factor-1 (Runx1) gene, a luteinizing hormone (LH)-induced transcription factor essential for ovulation, leading to increased Runx1 mRNA expression. Accordingly, blocking androgen-induced inhibition of Ezh2 in vivo adversely affects LH-induced Runx1 mRNA expression and subsequent ovulation. Importantly, although estrogen treatment of granulosa cells similarly causes rapid activation of the PI3K/Akt pathway and short-term phosphorylation of Ezh2, it does not induce miR-101 expression and thereby does not reduce overall Ezh2 expression, demonstrating the androgen specificity of long-term Ezh2 suppression. Thus, this study provides insight regarding how androgen-induced extranuclear kinase signaling and intranuclear transcription through Ezh2 modifications may influence the expression pattern of genes, ultimately affecting various downstream physiological processes.

Androgen Action in the Ovary

“…The classic mode of androgen action, as for most steroids, involves binding of androgen to AR in the cell cytoplasm and translocation of the hormone-receptor complex to the nucleus where it binds to a specific sequence in the promotor of the relevant target gene and promotes gene transcription ( 31 , 32 ). Whilst there is clear evidence that this pathway is operational in both physiological and pathological actions of androgens, recent work suggests that the pathway(s) of androgen signaling are more complex and involve rapid effects that do not involve classic nuclear receptor action on transcription ie non-genomic actions ( 22 , 33 , 34 ). These non-genomic actions have been highlighted in the work of Sen and colleagues who describe transactivation of the epidermal growth factor receptor (EGFR) by androgen.…”

“…Androgen can enhance both synthesis and action of insulin-like growth factors (IGFs) ( 18 , 28 ). IGFs signal primarily though the PI3Kinase (PI3K) pathway and recently, Sen and colleagues have provided evidence that androgens also directly activate the PI3K signaling pathway, leading to a complex cascade of events that involve, in an initial and rapid effect, phosphorylation, and thereby inhibition, of the polycomb group protein enhancer of zeste homolog 2 (Ezh2) ( 34 ). Ezh2 appears to be a factor in regulation of LH action in the preovulatory follicle.…”

“…Ezh2 appears to be a factor in regulation of LH action in the preovulatory follicle. In the longer term (24–48 h) the micro RNA miR-101 is induced which, in turn, greatly reduces the expression of Ezh2 protein ( 34 ). These findings give further support to their hypothesis that androgen action is likely to involve both genomic and non-genomic events that are closely co-ordinated.…”

The Effect of Folic Acid Supplementation in the Ovary and Upon Embryo Development

“…EZH2 is negatively regulated by phosphorylation via PI3K/AKT signalling pathway (Cha, 2005). A number of hormones such as E2, and androgen receptors (ER and AR respectively), and LH hormone (See Figure 1.6) can influence EZH2 activity through this mechanism (Cha, 2005;Bredfeldt et al, 2010;Ma et al, 2017). Moreover, among the targets of PRC2 and EZH2 are the differentiation-specific genes such as members of the GATA, SOX, FOX, POU family and components of Wnt, TGF-β, NOTCH, FGF and retinoic acid signalling (Simon and Lange, 2008).…”

“…These changes were seen at the same time as changes in Fshr and Oct4, which suggests they may be connected. Moreover, since EZH2 expression and activity is regulated by hormone levels (E2, androgens and LH), any alteration in hormone levels could affect EZH2-target genes associated with proliferation in the ovary (Bredfeldt et al, 2010;Ma et al, 2017). More detailed analyses are necessary to determine the methylation status and the binding sites of these epigenetic regulators of the gene expression.…”