GAS1 affects proliferation, apoptosis, and steroid hormone levels by regulating mitochondrial functions and the PI3K/AKT pathway in bovine granulosa cells

GAS1 is involved in the regulation of cell proliferation, apoptosis, and differentiation and has potential roles in ovarian function, ovarian luteum formation, and ovulation in mammals. However, its effect on hormone secretion in granulosa cells is not clear. In this study, through RNA sequencing, EdU, CCK8, qRT‒PCR, and flow cytometry assays, we found that GAS1 affected cell proliferation, apoptosis, redox homeostasis, steroid metabolism, and other biological processes in bovine granulosa cells. The results showed that after GAS1 inhibition, cell proliferation increased, whereas apoptosis decreased. The levels of P₄ and the P₄-related genes StAR and HSD3B1 decreased, whereas the levels of E₂ and the E₂-related gene CYP19A1 increased after GAS1 inhibition in bovine granulosa cells. The inhibition of GAS1 improved mitochondrial function by increasing ATP levels; regulating the mitochondrial homeostasis-related proteins MFN1, MFN2, DRP1, and PARKIN; and improving redox homeostasis by decreasing ROS levels and increasing CAT levels. In addition, the inhibition of GAS1 increased the level of phosphorylated PI3K/AKT, and LY294002, a PI3K/AKT pathway inhibitor, alleviated the effects of GAS1 inhibition on P₄ and E₂ levels in bovine granulosa cells. These results showed that GSA1 affects proliferation, apoptosis, mitochondrial function, and redox homeostasis and regulates P₄ and E₂ levels via the PI3K/AKT pathway in bovine granulosa cells. These results are helpful for analyzing the molecular mechanism of follicle development and provide a theoretical basis for improving follicle development and female animal fertility.