LH-Induced Neuregulin 1 (NRG1) Type III Transcripts Control Granulosa Cell Differentiation and Oocyte Maturation

Abstract

Epidermal growth factor (EGF)-like factors [amphiregulin (AREG), betacellulin, and epiregulin] are induced by LH and activate the EGF receptor (ERBB1)/ERK1/2 pathway in granulosa cells and cumulus cells of preovulatory follicles to impact ovulation. However, the expression and roles of other ERBB family members and their ligands have not been explored in detail. Herein, we document that two transcripts of the neuregulin (Nrg1) gene are expressed in granulosa cells, and that the type III Nrg1 is induced during ovulation in an ERK1/2 and C/EBPβ-dependent manner. Western blotting shows that intact (75 kDa) and secreted (45 kDa) forms of neuregulin 1 (NRG1) are present in the ovary. NRG1 likely binds to ERBB3/ERBB2 complexes that are expressed in granulosa cells and cumulus cells. In cultured granulosa cells, NRG1 selectively stimulates the phosphorylation of AKT/PKB compared to ERK1/2. However, when granulosa cells were cultured with NRG1 and AREG, the phosphorylation of ERK1/2 was markedly enhanced as compared with that by AREG alone. Cotreatment with NRG1 and AREG also increased progesterone production. When cumulus-oocyte complexes (COCs) were cultured with both NRG1 and AREG, the matured oocytes exhibited significantly higher developmental competence as compared with that of oocytes cultured with AREG alone. Collectively, these results document that the expression of type III NRG1 is induced in granulosa cells during ovulation and that NRG1 enhances AREG-induced ERK1/2 phosphorylation in both granulosa cells and cumulus cells. The NRG1 pathway has two roles: one is to enhance AREG-induced progesterone production in granulosa cells, and the other is to regulate oocyte maturation by a cumulus cell-dependent mechanism.

Characterization of microRNAs differentially expressed during bovine follicle development

“…Many of the signaling pathways overrepresented by these targets corresponded to growth factors putatively involved in follicular cell proliferation and steroidogenesis and prevention of premature luteinization including HGF, IGF1, TGFb, estrogen, neurotropin/TRK, PEDF, and NGF (Parrott et al 1993, Salas et al 2006, Glister et al 2010, Mani et al 2010, Garcia-Rudaz et al 2011, Chuderland et al 2013 and others involved in oocyte maturation including neuregulin, GDNF, and EGF (Conti et al 2006, Noma et al 2011, Linher-Melville & Li 2013, the fine-tune regulation of which is of key importance during the final stages of follicle development. Also present were several core cell cycle regulatory pathways (cell cycle regulation by BTG family proteins, estrogen-mediated S phase entry, and G1/S checkpoint regulation), a finding which is consistent with the notion that the development of the dominant follicle involves a substantial decrease in granulosa cell proliferation (Lussier et al 1987, Gaytan et al 1996.…”

Molecular cloning and expression analysis of neuregulin 1 (Nrg1) in the hypothalamus of Huoyan goose during different stages of the egg-laying cycle

“…In addition, Nrg1 was shown to be expressed in ovarian granulosa cells after the LH surge and may impact luteinization and oocyte maturation (Noma et al, 2011;Kawashima et al, 2014). In poultry, the reproductive endocrine system and the reproductive activity are strictly controlled by the hypothalamic-pituitary-gonadal axis (Padmanabhan et al, 2002).…”

Targeted Disruption of Nrg1 in Granulosa Cells Alters the Temporal Progression of Oocyte Maturation

“…COCs were isolated from these follicles by needle puncture and collected by pipette. Nonexpanded COCs were selected, and groups of 50 COCs were cultured in a 50-L drop of defined medium containing 1% fetal bovine serum with or without 100 ng/mL AREG and/or 1 or 10 ng/mL NRG1 in a 35-mm dish in the presence of 1 or 4 mM hypoxanthine (HX) (Sigma-Aldrich) (11). The meiotic status (disappearance of germinal vesicle [GV] or emission of the first polar body) was observed every 2 hours for up to 16 hours.…”

“…Twenty oocytes were lysed with 10 L of SDS sample buffer. Western blot analyses were performed according to our previous study (11). In brief, extracts (10 g of protein) or oocyte lysates were resolved by SDS-polyacrylamide gel (12.5%) electrophoresis and transferred to polyvinylidene difluoride membranes (GE Bioscience).…”

“…Nrg1 is expressed in the brains of embryonic and adult mice (9,10) but is also induced in mural granulosa cells of periovulatory follicles (11). LH activation of ERBB2 and ERBB3 occurs as early as 2 hours after human chorionic gonadotropin (LH) injections in vivo and thus concomitantly with the induction of Nrg1 type III expression in mural granulosa cells but not in cumulus cells (11,12), indicating that NRG1 might affect early events in ovulation and/or oocyte maturation. This is supported by the observation that NRG1 can suppress the spontaneous resumption of meiosis that occurs when COCs are isolated from preovulatory follicles in the absence of any hormone treatments and can improve the developmental competence of oocytes in in vitro fertilization (IVF) (11).…”