TNP1 Gene Modulates Proliferation, Apoptosis, and Testosterone Synthesis in Bovine (Bos taurus) Leydig Cells
WANG Zhen1, WANG Xin1, MENG Xiang1, KOU Meng-Yuan1, ZHANG Yun-Hai1,2,3, LIU Hong-Yu1,2,3, SONG Ning1,2,3,*
1 College of Animal Science and Technology, Anhui Agricultural University, Hefei 230036, China; 2 Anhui Province Key Laboratory of Local Livestock and Poultry Genetic Resource Conservation and Germplasm Innovation, Hefei 230036, China; 3 Research Institute of Beef Cattle Industry, Anhui Agricultural University, Hefei 230036, China
Abstract：The key to bull (Bos taurus) fertility lies in the development of testes and sperm production, with testosterone synthesis and secretion by Leydig cells influencing testicular physiological function. The transition protein 1 (TNP1) gene is highly expressed in adult bull testes; however, its impact on Leydig cells remains unclear. In this study, bovine Leydig cells were isolated and cultured, and overexpression and interference of TNP1 were conducted. The effects of TNP1 on cell proliferation, apoptosis, and testosterone synthesis were assessed by qRT-PCR, Western blot, 5-ethynyl-2'-deoxyuridine (EdU) staining, flow cytometry, and ELISA. The results showed that overexpression of TNP1 significantly upregulated the expression of proliferation-related genes proliferating cell nuclear antigen (PCNA) and cyclin-dependent kinase 2 (CDK2)(P<0.05); extremely significantly increased the proportion of cells in an active proliferative state; downregulated the expression of apoptosis-related gene B-cell lymphoma-2 associated X protein (BAX) (P<0.01), extremely significant reduced the apoptosis rate; upregulated the expression of testosterone synthesis-related genes cytochrome P450 family 17 subfamily A member 1 (CYP17A1), 17 β-hydroxysteroid dehydrogenase 3 (HSD17B3), and steroidogenic acute regulatory protein (STAR) as well as testosterone levels (P<0.01). Conversely, interference of TNP1 extremely significantly downregulated the expression of PCNA and CDK2 (P<0.01), decreased the proportion of cells in an active proliferative state; upregulated the expression of BAX (P<0.01), significantly increased the apoptosis rate; downregulated the expression of CYP17A1, HSD17B3, and STAR as well as testosterone levels (P<0.05). These results indicated that TNP1 promoted proliferation and testosterone synthesis while inhibiting apoptosis in bovine Leydig cells. This study provides a foundation for exploring the mechanism by which TNP1 regulates bovine Leydig cells and offers a theoretical basis for enhancing the reproductive performance of bulls.
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