Abstract
17β-estradiol (E2) regulates hormonal release as well as proliferation and cell death in the pituitary. The main nitric oxide receptor, nitric oxide sensitive- or soluble guanylyl cyclase (sGC), is a heterodimer composed of two subunits, α and β, that catalyses cGMP formation. α1β1 is the most abundant and widely expressed heterodimer, showing the greater activity. Previously we have shown that E2 decreased sGC activity but exerts opposite effects on sGC subunits increasing α1 and decreasing β1 mRNA and protein levels. In the present work we investigate the mechanisms by which E2 differentially regulates sGC subunits' expression on rat anterior pituitary gland. Experiments were performed on primary cultures of anterior pituitary cells from adult female Wistar rats at random stages of estrous cycle. After 6 h of E2 treatment, α1 mRNA and protein expression is increased while β1 levels are down-regulated. E2 effects on sGC expression are partially dependent on de novo transcription while de novo translation is fully required. E2 treatment decreased HuR mRNA stabilization factor and increased AUF1 p37 mRNA destabilization factor. E2-elicited β1 mRNA decrease correlates with a mRNA destabilization environment in the anterior pituitary gland. On the other hand, after 6 h of treatment, E2-BSA (1 nM) and E2-dendrimer conjugate (EDC, 1 nM) were unable to modify α1 or β1 mRNA levels, showing that nuclear receptor is involved in E2 actions. However, at earlier times (3 h), 1 nM EDC causes a transient decrease of α1 in a PI3k-dependent fashion. Our results show for the first time that E2 is able to exert opposite actions in the anterior pituitary gland, depending on the activation of classical or non-classical pathways. Thus, E2 can also modify sGC expression through membrane-initiated signals bringing to light a new point of regulation in NO/sGC pathway.
Highlights
Nitric oxide sensitive- or soluble guanylyl cyclase, the main intracellular receptor of nitric oxide is comprised of two subunits, a and b, of which several isoforms (a1, a2, a2i, b1 and b2) have been described. a1b1 is the most abundant and widely expressed heterodimer, showing the greater activity [1].The major female hormone, 17b-estradiol (E2), is a key regulator of pituitary physiology involved in hormonal release as well as proliferation and cell death in anterior pituitary gland [2,3,4].Previous studies from our laboratory show that acute E2 treatment exerts an inhibitory effect on sGC activity by downregulating sGC b1 subunit in anterior pituitary gland
To investigate which kind of E2 receptor (ER) is involved in such effect, anterior pituitary cell cultures were incubated with membrane-impermeable E2 compounds: bovine serum albumin-conjugated E2 (E2-BSA) and estrogen dendrimer conjugate (EDC) [25] using the free steroid (E2) as control
Our results show that 1 nM E2-BSA was unable to reproduce 1 nM free E2 effects on sGC subunits mRNA expression at the same concentration (Figure 1)
Summary
Previous studies from our laboratory show that acute E2 treatment exerts an inhibitory effect on sGC activity by downregulating sGC b1 subunit in anterior pituitary gland. This treatment increases sGC a1 expression from both, immature and adult rats [5,6]. The E2 effects on anterior pituitary sGC were observed after in vivo and in vitro treatment and during estrous cycle. These observations support a direct effect of E2 on sGC regulation and a differential and independent regulation on both subunits. Previous evidence [7,8,9] further sustains that under certain conditions, a1 and b1 can be independently expressed
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