ERK1/2-dependent gene expression in the bovine ovulating follicle

Yasmin Schuermann,Juliana Ferst,Rogério Ferreira,Bernardo Gasperin,Raj Duggavathi,Ejimedo Madogwe,Paulo B Gonçalves,Vilceu Bordignon,Monique T Rovani

doi:10.1038/s41598-018-34015-4

Abstract

Ovulation is triggered by gonadotropin surge-induced signaling cascades. To study the role of extracellular signal-regulated kinase 1/2 (ERK1/2) in bovine ovulation, we administered the pharmacological inhibitor, PD0325901, into the preovulatory dominant follicle by intrafollicular injection. Four of five cows treated with 50 µM PD0325901 failed to ovulate. To uncover the molecular basis of anovulation in ERK1/2-inhibited cows, we collected granulosa and theca cells from Vehicle and PD0325901 treated follicles. Next-generation sequencing of granulosa cell RNA revealed 285 differentially expressed genes between Vehicle and PD0325901-treated granulosa cells at 6 h post-GnRH. Multiple inflammation-related pathways were enriched among the differentially expressed genes. The ERK1/2 dependent LH-induced genes in granulosa cells included EGR1, ADAMTS1, STAT3 and TNFAIP6. Surprisingly, PD0325901 treatment did not affect STAR expression in granulosa cells at 6 h post-GnRH. Granulosa cells had higher STAR protein and theca cells had higher levels of STAR mRNA in ERK1/2-inhibited follicles. Further, both granulosa and theca cells of ERK1/2-inhibited follicles had higher expression of SLC16A1, a monocarboxylate transporter, transporting substances including β-hydroxybutyrate across the plasma membrane. Taken together, ERK1/2 plays a significant role in mediating LH surge-induced gene expression in granulosa and theca cells of the ovulating follicle in cattle.

Highlights

The process of ovulation is dependent upon the luteinizing hormone (LH) surge to elicit a plethora of signaling cascades required for the release of a fertilizable oocyte and formation of a corpus luteum (CL) needed for pregnancy[1,2]
Mouse studies have revealed that the absence of extracellular signal-regulated kinase 1 and 2 (ERK1/2) signalling reduces the expression of LH-regulated genes including: the early growth response 1 (Egr1) transcription factor, as well as genes required for follicular rupture (a disintegrin and metalloproteinase with thrombospondin motifs 1 (Adamts1) and prostaglandinendoperoxide synthase 2 (Ptgs2)), cumulus expansion (pentraxin 3(Ptx3) and tumor necrosis factor (TNF) alpha induced protein 6 (Tnfaip6)), oocyte maturation (amphiregulin (Areg)) and luteinization markers (steroidogenic acute regulatory protein (Star) and cytochrome P450 family 11 subfamily a member 1 (Cyp11a1))[10,11,12]
Transrectal ultrasonography five days after the gonadotropin releasing hormone (GnRH) treatment revealed that all cows treated with Vehicle, 1 μM and 10 μM doses of PD0325901 successfully ovulated, while only one of five cows treated with 50 μM PD0325901 ovulated, this cow had low levels of circulating progesterone, suggesting her CL was not functional (Fig. 1)

Summary

Introduction

The process of ovulation is dependent upon the luteinizing hormone (LH) surge to elicit a plethora of signaling cascades required for the release of a fertilizable oocyte and formation of a corpus luteum (CL) needed for pregnancy[1,2]. The LH-dependent changes in the gene expression program is required for follicle rupture, cumulus cell expansion, oocyte maturation, and luteinization[3]. Mouse studies have revealed that the absence of ERK1/2 signalling reduces the expression of LH-regulated genes including: the early growth response 1 (Egr1) transcription factor, as well as genes required for follicular rupture (a disintegrin and metalloproteinase with thrombospondin motifs 1 (Adamts1) and prostaglandinendoperoxide synthase 2 (Ptgs2)), cumulus expansion (pentraxin 3(Ptx3) and TNF alpha induced protein 6 (Tnfaip6)), oocyte maturation (amphiregulin (Areg)) and luteinization markers (steroidogenic acute regulatory protein (Star) and cytochrome P450 family 11 subfamily a member 1 (Cyp11a1))[10,11,12]

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Conclusion