DNA methylation mediated RSPO2 to promote follicular development in mammals

Abstract

In female mammals, the proliferation, apoptosis, and estradiol-17β (E2) secretion of granulosa cells (GCs) have come to decide the fate of follicles. DNA methylation and RSPO2 gene of Wnt signaling pathway have been reported to involve in the survival of GCs and follicular development. However, the molecular mechanisms for how DNA methylation regulates the expression of RSPO2 and participates in the follicular development are not clear. In this study, we found that the mRNA and protein levels of RSPO2 significantly increased during follicular development, but the DNA methylation level of RSPO2 promoter decreased gradually. Inhibition of DNA methylation or DNMT1 knockdown could decrease the methylation level of CpG island (CGI) in RSPO2 promoter and upregulate the expression level of RSPO2 in porcine GCs. The hypomethylation of −758/−749 and −563/−553 regions in RSPO2 promoter facilitated the occupancy of transcription factor E2F1 and promoted the transcriptional activity of RSPO2. Moreover, RSPO2 promoted the proliferation of GCs with increasing the expression level of PCNA, CDK1, and CCND1 and promoted the E2 secretion of GCs with increasing the expression level of CYP19A1 and HSD17B1 and inhibited the apoptosis of GCs with decreasing the expression level of Caspase3, cleaved Caspase3, cleaved Caspase8, cleaved Caspase9, cleaved PARP, and BAX. In addition, RSPO2 knockdown promoted the apoptosis of GCs, blocked the development of follicles, and delayed the onset of puberty with decreasing the expression level of Wnt signaling pathway-related genes (LGR4 and CTNNB1) in vivo. Taken together, the hypomethylation of −758/−749 and −563/−553 regions in RSPO2 promoter facilitated the occupancy of E2F1 and enhanced the transcription of RSPO2, which further promoted the proliferation and E2 secretion of GCs, inhibited the apoptosis of GCs, and ultimately ameliorated the development of follicles through Wnt signaling pathway. This study will provide useful information for further exploration on DNA-methylation-mediated RSPO2 pathway during follicular development.

The Dynamic Transcription Profiles of Proliferating Bovine Ovarian Granulosa When Exposed to Increased Levels of β-Hydroxybutyric Acid

“…The normal development of GCs is necessary to ensure a normal follicular cycle and maintaining the balance between the proliferation and apoptosis of GCs is vitally important for oocyte maturation and embryonic development (39). When this balance is disturbed, follicular development will be either retarded or even terminated (40). It is generally considered that in vitro cell models are particularly valuable for studying steroidogenesis and folliculogenesis, especially for humans and large ruminants (41,42).…”

A regulatory network controlling ovarian granulosa cell death

“…In this study, we further revealed a regulatory relationship between these factors in sow GCs and constructed a small regulatory network for controlling GC apoptosis and follicular atresia, with these factors as hub genes. This is different from previous studies on the regulatory mechanism of sow follicular atresia and GC apoptosis, which mainly focused on a single factor [ 45 , 46 ], regulatory axis [ 47 ], or signaling pathway [ 48 , 49 ]. Our findings define, for the first time, a small regulatory network that affects follicular atresia and GC apoptosis in sows, providing a new perspective for revealing the mechanism of complex processes such as GC apoptosis and follicular atresia.…”

Molecular mechanism of modified Huanglian Wendan decoction in the treatment of polycystic ovary syndrome

“…[ 29 , 30 ] CCND1 regulates cyclinD1 protein and promotes cytoplasmic proliferation, Cyclin D1 and its inhibitors are associated with GC proliferation at specific follicular stages, CCND1 promoted E2 secretion of PCOS ovarian GC, increased the expression level of CYP19A1, inhibited the expression level of Caspase 3 in apoptotic GC, and reduced CASP3 cleavage. [ 31 , 32 ] CASP3 is a regulatory factor of apoptosis signaling pathway, the early ovarian stage of PCOS patients is characterized by increased number of follicles, and the apoptosis rate of sinus follicles is related to increased expression of CASP3 in GC. Hyperandrogenemia in the late stage is closely related to excessive GC apoptosis.…”