11 Dec

Soluble Guanylyl Cyclase: INTRODUCTION


The actions of FSH and LH on ovarian functions are mediated in large part through increased production of the second-messenger cAMP and subsequent activation of downstream signaling pathways. Although the importance of cAMP as a second messenger influencing the ovary is well recognized, other hormones and factors acting through diverse signaling pathways also modulate ovarian functions. In this regard, a large body of evidence indicates potential roles of cGMP as an important cyclic nucleotide regulating ovarian steroidogenesis, gonadotropin-receptor expression, inhibin expression, apoptosis, and ovulation. Thus, understanding the mechanisms regulating cGMP production in the ovary has become increasingly critical.

The synthesis of cGMP from GTP is dependent on the activity of guanylyl cyclase, which exists in both particulate and soluble forms. The particulate, membrane-associated forms serve as receptors, the enzymatic activity of which is stimulated by binding of natriuretic peptides. In contrast, soluble guanylyl cyclase (sGC) is a heme-containing heterodimer consisting of an a subunit (variously reported as 73-88 kDa) and a smaller p subunit (70 kDa). Each subunit has two known isoforms (a!, a2, pb and p2), which have slightly differing activities and distributions. The a! and a2 isoforms are fairly homologous in their middle and carboxyl terminus portions, but the amino termini differ markedly. The p! and p2 subunits differ markedly at both amino and carboxyl termini, with less than 50% homology in their middle regions.

A major activator of sGC is nitric oxide (NO), which binds the heme group of sGC and markedly stimulates activity of this enzyme, increasing cGMP production. Carbon monoxide (CO) also stimulates sGC activity and may play physiological roles in triggering cGMP-dependent signaling pathways. In addition to posttranslational activation by NO and CO, sGC clearly is regulated at the message and protein levels by a number of factors, including NO, cGMP, cAMP, and estradiol (E2). Previous studies demonstrate that the sGC subunits are expressed in many organs, including gonadal tissues. Recent findings indicate that treatment of granulosa cells with a NO generator or specific activator of sGC increases cGMP accumulation, indicating sGC subunit expression in granulosa cells. Furthermore, our previous studies using an antibody that detects both sGC a and p subunits confirm the expression of sGC in rat granulosa cells (unpublished data). Whereas these observations demonstrate sGC expression in granulosa cells, to our knowledge no information is available regarding expression of sGC in other ovarian cell types or concerning regulation of sGC subunit protein levels in the ovary.

Given the growing importance of cGMP as a second messenger in the ovary and the minimal information regarding the control of cGMP production in gonadal tissues, the present study utilized immunohistochemical and im-munoblot analysis to examine the cell-specific localization and regulation of sGC a! and p! subunit protein levels in whole ovaries and cultured granulosa cells. Because the major activator of sGC, NO, is implicated in the inhibition of follicle growth and granulosa cell differentiation, we hypothesize that sGC may be down-regulated during these processes as one means of limiting the activity of NO. Furthermore, elucidation of the regulated, cell-specific manner of sGC expression in ovarian cells may provide new insights regarding the control of cGMP production and potential actions of this second messenger in regulating gonadal functions.

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