Nitric oxide sensitive guanylyl cyclase alfa1 and beta1 subunits are differentially regulated by classical and non-classical estrogen pathways in anterior pituitary gland

JIMENA P CABILLA, SONIA A RONCHETTI, SILVANA I NUDLER, FERNANDA A QUINTEROS, BEATRIZ H DUVILANSKI.

Congreso; The Endocrine Society's 93nd. Annual Meeting,; 2011

The Endocrine Society

17b-estradiol (E2) regulates hormonal release as well as pituitary cells’ proliferation and death. The main nitric oxide receptor, soluble guanylyl cyclase (sGC), is a heterodimer composed by two subunits a and b and catalyzes cGMP formation. Previously we have shown that E2 exerts opposite effects on these sGC subunits, increasing both a1 mRNA and protein levels and decreasing b1 expression in vitro and in vivo. In the present work we investigate the mechanisms by which E2 through classical and/or non classical pathways regulates sGC subunits’ expression on primary cultures of anterior pituitary cells. After 6 h, actinomycin D (2 mM) partially abolished E2-elicited a1 mRNA and protein augment and b1 levels decrease (relative units (RU) as % of control (C): E2, a1: 148±11**, b1: 72±8*; Act D, a1: 91±7, b1: 94±6; E2+Act D, a1: 121±9#, b1: 86±8; *p<0.05, **p<0.01 vs C; #p<0.05 vs E2, n=3). Cycloheximide (10 mg/mL) fully abolished E2 actions on a1 and b1 mRNA expression (RU as % of C; E2, a1: 162±13**, b1: 48±7**; CHX, a1: 98±5, b1: 102±4; CHX+E2, a1: 99±4##, b1: 100±5#; **p<0.01 vs C; #p<0.05, ##p<0.01 vs E2, n=3). E2 decreased HuR mRNA stabilization factor and increased AUF1p37 mRNA destabilization factor expression (% of respective C, HuR: 46±5**, AUFp37: 142±2**, **p<0.01, n=3). After 6 h of treatment, E2-BSA (1 nM) and E2-dendrimer conjugate (EDC, 1 nM) were unable to modify a1 or b1 mRNA levels, showing that intracellular receptor is involved in these E2 actions. However, at earlier times (3 h), 1 nM EDC decreased a1 mRNA levels and slightly increased b1 levels (RU as % of C; E2, a1: 115,6±10*, b1: 67±17*; empty dendrimer, a1: 95±24, b1: 114±5; EDC, a1: 59±10#, b1: 88±11; *p<0.05 vs C, #p<0.01 vs E2, n=4), opposite to E2 effects at longer times of exposure. Here we show for the first time that E2 activation of membrane or cytoplasmic pathways leads to different effects on sGC subunits expression. Upon membrane receptors activation, E2 causes a transiently decrease of a1 subunit expression, contrary to that observed at longer times. At this time (6 h), a1 levels are increased, E2 effects on sGC expression is partially dependent on de novo transcription and de novo translation is fully required. E2-elicited b1 mRNA downregulation correlates with mRNA destabilization environment in anterior pituitary gland. Our results show that E2 has an opposite effect on sGC expression depending on the activation of classical or non-classical pathways.