Acute Signaling by the LH Receptor Is Independent of Protein Kinase C Activation

Abstract

LH receptor activation leads to the phosphorylation/activation of p42/44 MAPK in preovulatory granulosa cells. As the LH receptor can activate both adenylyl cyclase and phospholipase C, we hypothesized that the LH receptor could elicit phosphorylation of p42/44 MAPK through activation of protein kinase A (PKA) and/or protein kinase C (PKC). Preovulatory granulosa cells in serum-free primary cultures were treated with ovulatory concentrations of human chorionic gonadotropin (hCG), an LH receptor agonist, with or without various inhibitors. The PKA inhibitor H89 as well as the myristoylated PKA inhibitor peptide PKI strongly inhibited hCG-stimulated p42/44 MAPK phosphorylation, whereas the PKC inhibitor GF109203X had no effect on p42/44 MAPK phosphorylation. LH receptor-stimulated phosphorylation of cAMP response element-binding protein (CREB), histone H3, and MAPK kinase (MEK) was also strongly inhibited by H89 and not by GF109203X. The extent of PKC activation was assessed in preovulatory granulosa cells using three criteria: translocation of PKC isoforms to the membrane fraction, phosphorylation of a known PKC substrate, and autophosphorylation of PKC delta on an activation-related site. By all three criteria PKCs were partially activated before hCG stimulation, and hCG treatment failed to elicit further PKC activation, in vitro or in vivo. Taken together, these results indicate that, under primary culture conditions where physiological levels of signaling proteins are present, hCG signals to activate MEK, p42/44 MAPK, CREB, and histone H3 in a predominantly PKA-dependent and PKC-independent manner. Unexpectedly, PKCs were partially activated in the absence of LH receptor activation, and LH receptor activation did not elicit further detectable PKC activation.

Paracrine regulation of growth hormone gene expression by gonadotrophin release in grass carp pituitary cells: functional implications, molecular mechanisms and signal transduction

“…subsequent activation of the AC/cAMP/PKA pathway . Recently, functional coupling of LH receptors to MAPK-and PI3K-dependent mechanisms has also been reported in granulosa cells (Salvador et al 2002) and Leydig cells (Hirakawa et al 2002). In this study, hCG treatment not only elevated steady-state GH mRNA levels but also increased cAMP production in carp pituitary cells.…”

Aromatase expression in the ovary: Hormonal and molecular regulation

“…Remarkably, LH causes a rapid and transient activation of ERK1/2 in granulosa cells expressing low levels of LH receptors; whereas, if high levels of LH receptor are present, a delayed and more sustained activation of ERK1/2 take place [77]. The early increase in ERK1/2 phosphorylation is PKA dependent and PKC independent [75]. In contrast the delayed effect is PKA and PKC dependent and seems to be mediated by an increase in epidermal growth-like factors (EGF-1) [77].…”

“…In granulosa cells, ERK1/2 phosphorylation is stimulated by LH [74][75][76]. Remarkably, LH causes a rapid and transient activation of ERK1/2 in granulosa cells expressing low levels of LH receptors; whereas, if high levels of LH receptor are present, a delayed and more sustained activation of ERK1/2 take place [77].…”

Mutations of the lutropin/choriogonadotropin receptor that do not activate the phosphoinositide cascade allow hCG to induce aromatase expression in immature rat granulosa cells

“…At low receptor densities the LHR and FSHR stimulate cAMP accumulation, and elicit a transient increase in Akt and ERK1/2 phosphorylation but they do not stimulate inositol phosphate accumulation (this paper and Alam et al, 2004;Andric and Ascoli, 2006;Bebia et al, 2001;Donadeu and Ascoli, 2005;Gonzalez-Robayna et al, 1999;Gonzalez-Robayna et al, 2000;Hunzicker-Dunn and Maizels, 2006;Salvador et al, 2002;Zeleznik et al, 2003). At these low receptor densities the LHR and FSHR also stimulate epiregulin and aromatase expression (this paper and Andric and Ascoli, 2006;Bebia et al, 2001;Donadeu and Ascoli, 2005;Fitzpatrick et al, 1997;Fitzpatrick and Richards, 1993;Zeleznik et al, 2003).…”

“…At low gonadotropin receptor densities only a rapid and transient increase in ERK1/2 phosphorylation is observed (Andric and Ascoli, 2006;Cottom et al, 2003;Salvador et al, 2002;Seger et al, 2001), whereas at high receptor densities an additional, delayed increase in phosphorylation becomes obvious (Andric and Ascoli, 2006). Here we show that the two LHR mutants that do not activate the phosphoinositide cascade also do not support a robust increase in the early or late increase in ERK1/2 phosphorylation.…”