HDAC11 in ovarian granulosa cells coordinates LH in the maturation of oocytes in Tan sheep

Abstract

Oocyte maturation plays an important role in supporting mammalian reproduction. Histone deacetylase 11 (HDAC11), the only member of the class IV histone deacetylase family and the smallest histone deacetylases (HDACs), has been shown to regulate oocyte maturation in mice and pigs. However, the epigenetic effects of HDACs in follicular granulosa cells in response to LH induction remain elusive in sheep. In this study, the effects of follicular somatic cell-derived HDAC11 on oocyte maturation in Tan sheep are evaluated. The expression changes of HDAC11 and related proteins are detected by means of immunofluorescence, immunohistochemistry, western blot analysis and enzyme-linked immunosorbent assay. Our results indicate that the level of HDAC11 in follicular granulosa cells as well as oocytes in Tan sheep increases with the growth and maturation of the follicles. Specific inhibition of HDAC11 by SIS17 remarkably reduces the oocyte maturation rate under LH supplementation in vitro. Accordingly, the acetylation level of H3K9 in granulosa cells is increased, while the EGF-like growth factor AREG is remarkably decreased. Furthermore, inhibition of HDAC11 markedly decreases the level of YAP1, which is a negative regulator of AREG in granulosa cells. Conclusively, HDAC11 in the granulosa cells of Tan sheep contributes to the LH induced production of AREG during oocyte in vitro maturation by decreasing the level of H3K9 acetylation and increasing the level of YAP1.

1. Introduction

The regulation of mammalian oocyte development is an extremely complex process, which involves the coordination of endocrine and epigenetic modifications in the follicular niche [1]. In domestic animals, the quality of the in vitro maturation (IVM) of oocytes determines the developmental potential of in vitro fertilized embryos and affects the pregnancy rate and birth rate after embryo transfer [2]. Luteinizing hormone (LH) acts on follicular granulosa cells before ovulation and activates downstream signaling pathways to induce cytoplasmic and nuclear maturation, ovulation, and luteal cell formation [3–5], which induces the production of EGF-like factors to promote oocyte maturation [5,6]. The EGF-like factors, including amphiregulin (AREG), epiregulin (EREG) and betacellulin (BTC), induce large gene activation networks in granulosa cells and cumulus cells in species specific manners, as well as coordinate processes such as oocyte maturation, cumulus expansion, and ovulation [7]. Related studies have shown that HDAC3 in granulosa cells of mouse is the key to negatively regulating EFG-like factor production [8]. In addition, HDAC1 and HDAC6 in granulosa cells of Tan sheep are either negative or positive in regulating oocyte maturation in vitro [9,10]. In mice, HDAC2 is mainly responsible for H4K16 deacetylation during oocyte maturation [11]. HDAC8 is localized to the spindle apparatus in porcine oocytes, and plays a role in spindle assembly by recruiting γ-tubulin, thereby driving oocyte meiotic maturation [12]. Therefore, HDACs are important to the maturation of oocytes in mammals in a cell specific manner during gonadotropin induction.

HDAC11 is highly expressed in multiple organs, and can be found in the mitochondria, cytoplasm, or nuclei, depending on the type of tissue and cell [13]. HDAC11 plays a role in the regulation of adipogenesis, lipid metabolism, metabolic inflammation, glucose tolerance, immune responses, and energy consumption [13]. According to existing reports, HDAC11 has important effects on oocyte maturation in mice [14] and pigs [15]. HDAC11 remarkably disrupts the meiotic process of mouse oocytes, leading to spindle disorders chromosomal dislocation, and impaired kinetochore-microtubule attachment and spindle-assembly checkpoint function [14]. HDAC11 is also involved in the formation of meiotic spindle, chromosome arrangement and separation, and mRNA transcription during porcine oocyte maturation by regulating α-tubulin acetylation and histone modification [15]. HDAC11 shows an unprecedented homology to class I and II catalytic domains, with low overall protein homology [16]. However, whether HDAC11 as the only member of class IV HDAC in the granulosa cells of Tan sheep plays pivotal role in oocyte maturation remains unclear.

EGF-like factors play key roles in improving oocyte meiotic maturation and cumulus expansion in mammals [7,8]. AREG is a key family member of EGF-like factors and mediates the regulation of LH cas