Expression and Functional Analysis of the BCL2-Associated Agonist of Cell Death (BAD) Gene in the Sheep Ovary During the Reproductive Cycle.

Xiaohan Cao,Xiaoyun He,Xiaoyu Li,Mingxing Chu,Wenping Hu,Qiuyue Liu,Ran Di,Xiangyu Wang,Lulu Lu,Xianyin Zeng

doi:10.3389/fendo.2018.00512

Abstract

Most ewes in China are seasonally polyestrous with normal ovulatory cycles, which is controlled by photoperiod (length of the daily light phase). These ewes are estrous in the short-day season and anestrus in the long-day season and cannot mate during anestrus. Thus seasonal breeding limits both diversification and intensification of production. If sheep can estrus all round year, it can be mated twice per year, which can greatly improve the economic benefits. To change seasonal estrus at the genetic level and cultivating new sheep breeds, it is important to understand the molecular mechanisms of seasonal breeding trait in sheep. The BCL2-associated agonist of cell death (BAD) gene being a regulator of cellular apoptosis was identified by our previous RNA-Seq, which is associated with follicular development in mammalian ovaries. However, the mechanism how BAD can regulate estrus in sheep was poorly understood. In this study, we characterized ovine BAD, including full-length mRNA cloning and protein sequence prediction, as well as BAD expression profile in Small-tailed Han (STH) sheep. The highest expression levels of BAD were observed in sheep hypothalamus, lung, and pituitary, while the lowest expression was in liver. Functional analysis of BAD was performed in primary granulosa cells of sheep. The concentration of P4 was significantly increased after RNAi interference of BAD, while P4 level was shown to be opposite after BAD overexpression in vitro. It has been found that BAD can reduce progesterone levels by promoting ovarian GC apoptosis, which might be involved in regulating the estrus cycle in sheep.

Highlights

Seasonality in reproduction is characteristic of most wild and domesticated species of mammals and birds
Among the differentially expressed genes detected by RNAsequencing during anestrus and the breeding season (unpublished data, the RNA-sequencing data reported have been deposited to the Genome Sequence Archive of the BIG Data Center 34 under accession number PRJCA000881.), B-cell lymphoma-2 (BCL2)-associated agonist of cell death (BAD) expression was significantly upregulated during proestrus (p < 0.01; Figure 1A) in Small-tailed Han (STH) sheep
Despite extensive research on the role of apoptosis in normal ovarian development and function, limited functional studies have been performed on the role of BAD in granulosa cells (GCs) apoptosis during the transition from a non-breeding to a breeding state among seasonally breeding mammals

Summary

Introduction

Seasonality in reproduction is characteristic of most wild and domesticated species of mammals and birds. Temperature, and ambient photoperiod (PP; length of the daily light phase) can facilitate or inhibit reproductive function seasonally. The PP serves as the primary factor in the regulation of reproductive timing in long- and short-day breeding mammals and most bird species [1, 2]. Upstream signals mainly go through the hypothalamic–pituitary–ovarian axis to regulate gonadotropin and ovarian steroid levels, which in turn influence reproductive activities. Ewes are estrous in the short-day season and anestrus in the long-day season.

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Results

Conclusion