Expression Patterns of Key Factors of Wnt/β-catenin and Notch Signaling Pathway in Bovine (Bos taurus) Follicular Granulosa Cells
DANG Wen-Qing1, HE Min1, QU Yu-Han1, ZHANG Kai1, YANG Xiao-Feng2, QIN Xiao-Wei3, HAN Qi1, GUO Xiang-Yu1, LYU Li-Hua1,*
1 College of Animal Science, Shanxi Agricultural University, Taigu 030801, China; 2 Department of Biology, Xinzhou Normal College, Xinzhou 034000, China; 3 Department of Reproductive Medicine, Shanxi Provincial People's Hospital, Taiyuan 030012, China
Abstract：The proliferation, apoptosis and steroid hormone synthesis of granulosa cells play important roles in follicle development, and these functions are regulated by signaling pathways. In this study, the transcriptome sequencing of granulosa cells from the first large follicle F1 and the second large follicle F2 in bovine (Bos taurus) first follicle wave pre-deviation (PD) and onset of diameter deviation (OD) were separately performed to obtain the expression patterns of genes in Wnt/β-catenin and Notch signaling pathways, cell cycle, apoptosis and steroid hormone synthesis, and to validate the expression of key components of the 2 pathways. The results showed that the genes of key components of the 2 pathways with high expression in PDF1, PDF2, ODF1 and ODF2 follicular granulosa cells were wingless-type MMTV integration site family, member 10a (Wnt10a), Wnt2b, frizzled3 (Fzd3), low density lipoprotein-related receptor 8 (LRP8), dishevelled 3 (Dvl3), Axin1, glycogen synthase 3β (GSK3β), catenin beta 1 (CTNNB1), transcription factor 7-like 2 (TCF7L2), lymphoid enhancer factor1 (LEF1), B-cell CLL/Lymphoma 9 (BCL9), Notch2, Dll4 and Hes related with YRPW motif2 (Hey2). Cell cycle-related genes with high expression were cyclin2 (CCND2), cyclindependent kinase4 (CDK4), cyclin-dependent kinase inhibitor 1A (CDKN1A), CDKN1B. The apoptosis of granulosa cells was mainly regulated by BCL2-associated X (Bax) in the mitochondrial apoptosis pathway and executed by Caspase3 and Caspase6. The expression of the steroid genes were 3β-hydroxysteroid dehydrogenase (3β-HSD), cytochrome P450 family 11 subfamily A member 1 (CYP11A1), and CYP19A1 were elevated, and E2 synthesis ability of OD follicular granulosa cells was increased compared with that of PD follicles. Wnt2 protein was localized in follicular granulosa cells in deviation process and its expression was significantly higher in OD follicular granulosa cells (P<0.05); Wnt2b protein was localized in follicular granulosa cells and theca cells and its expression was extremely significant lower in OD follicular granulosa cells (P<0.01). The mRNA expression of CTNNB1, LEF1, Notch2 and Hey2 were all extremely significantly increased in OD follicular granulosa cells (P<0.01). This study showed that the gene expression in F1 and F2 follicular granulosa cells during the deviation of bovine first follicular wave was specific, and the gene expression of key components in Wnt/β-catenin and Notch signaling pathway had the same trend, which provides a reference basis for the molecular regulatory mechanism of granulosa cell function during follicular development.
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