5-Aza-dC对牛成肌细胞MyoD1启动子甲基化及mRNA表达的影响

doi:10.11843/j.issn.0366-6964.2021.09.007

Abstract

Abstract: The aim of this study was to investigate the effects of 5-Aza-2'-deoxycytidine (5-Aza-dC) on the proliferation of bovine myoblasts, promoter methylation and mRNA expression of myogenic differentiation 1 (MyoD1). In this study, the Guanling cattle myoblasts cryopreserved in the early stage were recovered and cultured. After they had grown to the logarithmic growth phase, the bovine myoblasts were treated by different concentrations of 5-Aza-dC. The flow cytometry was used to detect cell apoptosis and cycle. The methylation level of MyoD1 promoter was detected by high-throughput detection methods, the expression level of MyoD1 and related genes were detected by qRT-PCR. The results showed that 0.1 μmol·L^-1 5-Aza-dC was the optimal concentration, and the expression of anti-apoptotic factor Bcl was significantly decreased (P<0.01), the expression of pro-apoptotic factor Bax was significantly increased (P<0.01), and the expression of pro-apoptotic factor Caspase-9 was significantly increased (P<0.05); The expression of Cyclin A2 was significantly increased (P<0.05), but the expression of Cyclin B1 and Cyclin D were not significantly affected. The MyoD1 promoter methylation level in blank group and experimental group was detected, and the MyoD1 promoter methylation level in experimental group was significantly lower than that in blank group (P<0.01), while the mRNA expression level was significantly higher in experimental group than that in blank group (P<0.01). In summary, 5-Aza-dC can regulate the proliferation and apoptosis of Guanling cattle myoblasts by changing the expressions of Caspase-9, Bcl, Bax, Cyclin A2 and other genes, and can effectively reduce the methylation level of MyoD1 gene promoter (P<0.01), and significantly increase its mRNA expression (P<0.01). Low concentration of 5-Aza-dC can promote cell apoptosis and regulate MyoD1 methylation level to regulate the expression of MyoD1 in Guanling cattle. At the same time, it is speculated that the methylation level of the MyoD1 gene promoter can affect the growth and development of Guanling cattle myoblasts, which can provide a theoretical reference for screening genetic markers to assist the improvement of Guanling cattle.

Key words: Guanling cattle, 5-Aza-2'-deoxycytidine, DNA methylation, cell cycle, apoptosis

CLC Number:

LI Xiong, TIAN Niannian, SONG Linjin, CHEN Chen, XU Houqiang. Effects of 5-Aza-dC on MyoD1 Promoter Methylation and mRNA Expression in Bovine Myoblasts[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(9): 2439-2451.

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Effects of 5-Aza-dC on MyoD1 Promoter Methylation and mRNA Expression in Bovine Myoblasts

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