CircRNA-Mediated Inhibin-Activin Balance Regulation in Ovarian Granulosa Cell Apoptosis and Follicular Atresia.

Mengnan Ma,Yi Zhang,Jinbi Zhang,Huiming Wang,Jingge Liu,Zengxiang Pan

doi:10.3390/ijms22179113

Abstract

Ovarian granulosa cells (GC) play an essential role in the development and atresia of follicles. Emerging studies suggest that non-coding RNAs are involved in the regulation of GC apoptosis. Here, we aimed to analyze the function of ssc-circINHA-001, coded by the first exon of the inhibin subunit α gene (INHA), in resisting GC apoptosis and follicular atresia by enhancing the expression of the inhibin subunit β A (INHBA) through a cluster of miRNAs. A higher expression of ssc-circINHA-001 in healthy follicles compared to early atretic follicles was detected by qRT-PCR. Its circular structure was confirmed by RNase R treatment and reversed PCR. The function of ssc-circINHA-001 in GC resistance to apoptosis was detected by in vitro transfection of its si-RNA. Furthermore, the dual-luciferase reporter assay suggested that ssc-circINHA-001 adsorbed three miRNAs, termed miR-214-5p, miR-7144-3p, and miR-9830-5p, which share the common target INHBA. A low expression of ssc-circINHA-001 increased the levels of the free miRNAs, inhibited INHBA expression, and thus raised GCs apoptosis through a shift from the secretion of activin to that of inhibin. Our study demonstrated the existence of a circRNA–microRNAs–INHBA regulatory axis in follicular GC apoptosis and provides insight into the relationship between circRNA function and its coding gene in inhibin/activin balance and ovarian physiological functions.

Highlights

The quantity of ovarian follicles determines the reproductive rate of livestock [1].Ovarian granulosa cell (GC) apoptosis is a physiological phenomenon that occurs during all stages of follicular development
Our previous deep circular RNA (circRNA) sequencing analysis showed that the expression level of ssc-circINHA-001, a candidate circRNA produced from the first exon (50 UTR) region of the INHA gene, decreased during follicular atresia [9]
To further elucidate the role of this circRNA in follicular atresia, the expression levels of ssc-circINHA-001 in healthy follicles (HF) and atretic follicles (AF) were measured by qRT-PCR, and a significant reduction was observed in AF (Figure 1A)

Summary

Introduction

The quantity of ovarian follicles determines the reproductive rate of livestock [1]. Ovarian granulosa cell (GC) apoptosis is a physiological phenomenon that occurs during all stages of follicular development. Inhibin A and B are heterodimers comprising an α- and a β(A/B)-subunit, respectively. Inhibin A and B arise from three gene products, including the α-subunit gene (INHA), the βA-subunit gene (INHBA), and the βB-subunit gene (INHBB). Activins are homodimers or heterodimers of only the β subunits (βA, βB, βC, βD, and βE). Inhibins and activins counteract each other, and the balance between the amounts of α- and β-subunits will determine how much inhibin versus activin is formed, e.g., increased levels of α-subunit will favor the formation of inhibin, while increased levels of β-subunit will favor the formation of activin [3].

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