p38 Mitogen-Activated Protein Kinase Is Critical for Synergistic Induction of the FSHβ Gene by Gonadotropin-Releasing Hormone and Activin through Augmentation of c-Fos Induction and Smad Phosphorylation

Abstract

GnRH and activin independently and synergistically activate transcription of the FSH beta-subunit gene, the subunit that provides specificity and is the limiting factor in the synthesis of the mature hormone. This synergistic interaction, as determined by two-way ANOVA, is specific for FSHbeta and may, therefore, contribute to differential expression of the two gonadotropin hormones, which is critical for the reproductive cycle. We find that the cross-talk between the GnRH and activin signaling pathways occurs at the level of p38 MAPK, because the synergy is dependent on p38 MAPK activity, which is activated by GnRH, and activin cotreatment augments p38 activation by GnRH. Both the Smad and activator protein-1 binding sites on the FSHbeta promoter are necessary and sufficient for synergy. After cotreatment, Smad 3 proteins are more highly phosphorylated on the activin-receptor signaling-dependent residues on the C terminus than with activin treatment alone, and c-Fos is more highly expressed than with GnRH treatment alone. Inhibition of p38 by either of two different inhibitors or a dominant-negative p38 kinase abrogates synergy on FSHbeta expression, reduces c-Fos induction by GnRH, and prevents the further increase in c-Fos levels that occurs with cotreatment. Additionally, p38 is necessary for maximal Smad 3 C-terminal phosphorylation by activin treatment alone and for the further increase caused by cotreatment. Thus, p38 is the pivotal signaling molecule that integrates GnRH and activin interaction on the FSHbeta promoter through higher induction of c-Fos and elevated Smad phosphorylation.

Frequency-Dependent Regulation of Follicle-Stimulating Hormone β by Pulsatile Gonadotropin-Releasing Hormone Is Mediated by Functional Antagonism of bZIP Transcription Factors

“…The answer may rest in either the differential activation of signaling systems within the gonadotrope and/or synthesis, modification, and degradation of either transcription factors or regulatory proteins. It is well established that members of the mitogen activated protein kinase (MAPK) family of signaling proteins are stimulated by GnRH in vivo (50), in cultured rat pituitary fragments (22) and in L␤T2 cells (9,31) to contribute to regulation of gonadotropin subunit gene expression. In particular, activation of phosphorylated extracellular signal-regulated kinases (pERK) 1/2 is more rapid and more sustained in L␤T2 cells perifused with low, rather than high, GnRH pulse frequencies, suggesting that pERK is important in the preferential increase of FSH␤ transcription observed at low GnRH pulse frequency (27).…”

Leukemia Inhibitory Factor Represses GnRH Gene Expression via cFOS during Inflammation in Male Mice

“…Cells were plated into 12-well plates and transfected using Fugene 6 reagent (Roche Applied Science), as described previously [66-69]. Wells were transfected with 500 ng of reporter plasmid, 100 ng of β-galactosidase reporter plasmid driven by the Herpesvirus thymidine kinase promoter, as an internal control for the efficiency of the transfection, and 200 ng of expression vectors or empty vector control, as indicated in the figure legends.…”

Mechanisms of FSH synthesis: what we know, what we don't, and why you should care

“…Nucleotides conserved across species are marked with an asterisk (*) and gaps (À) have been introduced to facilitate the alignment. Gonadotropin-releasing hormone also stimulates the p38 MAPK pathway, and inhibitors of this kinase attenuate GnRH induction of ovine, murine, and human Fshb/FSHB reporters in LbT2 cells (165,166,170). To our knowledge, a role for p38 in GnRH induction of the rat promoter in cell lines has not been reported.…”

“…Importantly, delivery of pulsatile GnRH to GnRH-deficient male rats (i.e., castrated and testosterone treated) similarly stimulates ERK1/2 phosphorylation in the pituitary in vivo, with slower pulses stimulating greater increases than faster pulses (163,164). Chemical inhibition of MAPK kinase 1 (MEK1; MAP2K1), the upstream ERK1/2 kinase, attenuates GnRH-stimulated ovine, murine, rat, and human Fshb/FSHB promoter activities in LbT2 cells (158,(165)(166)(167)(168)(169)(170). Similarly, pulsatile GnRH stimulates Fshb mRNA levels and FSH release in male rat primary pituitary cultures, and these effects are attenuated by MEK1 inhibition (163).…”