Luteinizing Hormone Signaling in Preovulatory Follicles Involves Early Activation of the Epidermal Growth Factor Receptor Pathway

Abstract

LH activates a cascade of signaling events that are propagated throughout the ovarian preovulatory follicle to promote ovulation of a mature egg. Critical to LH-induced ovulation is the induction of epidermal growth factor (EGF)-like growth factors and transactivation of EGF receptor (EGFR) signaling. Because the timing of this transactivation has not been well characterized, we investigated the dynamics of LH regulation of the EGF network in cultured follicles. Preovulatory follicles were cultured with or without recombinant LH and/or specific inhibitors. EGFR and MAPK phosphorylation were examined by immunoprecipitation and Western blot analyses. By semiquantitative RT-PCR, increases in amphiregulin and epiregulin mRNAs were detected 30 min after recombinant LH stimulation of follicles and were maximal after 2 h. LH-induced EGFR phosphorylation also increased after 30 min and reached a maximum at 2 h. EGFR activation precedes oocyte maturation and is cAMP dependent, because forskolin similarly activated EGFR. LH-induced EGFR phosphorylation was sensitive to AG1478, an EGFR kinase inhibitor, and to inhibitors of matrix metalloproteases GM6001 and TNFalpha protease inhibitor-1 (TAPI-1), suggesting the involvement of EGF-like growth factor shedding. LH- but not amphiregulin-induced oocyte maturation and EGFR phosphorylation were sensitive to protein synthesis inhibition. When granulosa cells were cultured with a combination of neutralizing antibodies against amphiregulin, epiregulin, and betacellulin, EGFR phosphorylation and MAPK activation were inhibited. In cultured follicles, LH-induced MAPK activation was partially inhibited by AG1478 and GM6001, indicating that this pathway is regulated in part by the EGF network but also involves additional pathways. Thus, complex mechanisms are involved in the rapid amplification and propagation of the LH signal within preovulatory follicles and include the early activation of the EGF network.

Cross-talk between G Protein-coupled and Epidermal Growth Factor Receptors Regulates Gonadotropin-mediated Steroidogenesis in Leydig Cells

“…These observations indicate that cAMP and PKA are both necessary and sufficient for short term EGFR-mediated steroidogenesis. Interestingly, cAMP/PKA signaling may be a conserved mechanism for regulating LH and EGF receptor cross-talk, as cAMP appears to mediate gonadotropin-induced trans-activation of the EGFR during oocyte maturation (23). How PKA activates the EGFR is still unclear.…”

“…maturation, and steroidogenesis, suggesting that extracellular signaling is essential for EGFR trans-activation (15,22,23). In MA-10 mouse Leydig cells, MMP inhibitors also reduce phosphorylation of the EGFR (13,14).…”

Gene expression profiling of bovine preovulatory follicles: gonadotropin surge and prostanoid-dependent up-regulation of genes potentially linked to the ovulatory process

“…Regulation of bovine preovulatory follicular ADAM17 and AREG mRNAs following the LH surge Extensive evidence in rodents supports a key role of EGFlike growth factors in mediating LH action in preovulatory follicles (Ben-Ami et al 2006, Conti et al 2006, Hsieh et al 2007, Panigone et al 2008. Moreover, ADAMs, such as ADAM10 function as sheddases and play critical roles in releasing such EGF receptor ligands (Blobel 2005).…”

Cumulus cell gene expression following the LH surge in bovine preovulatory follicles: potential early markers of oocyte competence

“…In fact, LH was reported to be important to oocyte final maturation, ovulation, as well as optimal fertilization and early embryo development . This is achieved through activation of intrafollicular signaling and gene expression pathways leading to CCs expansion, meiosis resumption, oocyte final maturation, and ovulation in most mammalian species including mouse (Panigone et al 2008), pig (Kawashima et al 2008), cow (Hyttel et al 1997, and human (Filicori 1999, Feuerstein et al 2007. Although it has been established that LH triggers key functions in both oocyte and cumulus, there are questions about the presence of LH receptor proteins in CCs membrane, suggesting an indirect induction pathway (Peng et al 1991) probably through the EGFR (epidermal growth factor receptor) pathway (Panigone et al 2008, Reizel et al 2010.…”