猪流行性腹泻病毒通过miR-133c-3p/BCL2L2轴调控细胞凋亡

doi:10.11843/j.issn.0366-6964.2021.08.016

Abstract

Abstract: The purpose of this study was to investigate the role of miR-133c-3p in the process of cell apoptosis caused by porcine epidemic diarrhea virus (PEDV) infection, and to explore its mechanism. In this study, PEDV-infected MARC-145 cells were used as a model to detect the expression differences of 6 apoptosis-related microRNAs during PEDV infection. The expression levels of 6 apoptosis-related microRNAs were measured by RT-qPCR during PEDV infection. The apoptosis of PEDV-infected, miR-133c-3p transfected MARC-145 cells was detected by flow cytometry. The effect of miR-133c-3p on cell apoptosis was determined by flow cytometry. The target gene of miR-133c-3p was predicted by bioinformatics method. The binding of miR-133c-3p and the 3'UTR of target gene was determined by the luciferase reporter genet. The expression levels of BCL-w and PEDV protein were determined by Western blot when miR-133c-3p was over-expressed. The cell apoptosis rate was determined by flow cytometry when knock-down of BCL-w. The results showed that PEDV infection could induce apoptosis of MARC-145 cells, and the expression of microRNAs related to apoptosis, such as miR-133c-3p and miR-149-5p, were upregulated (P<0.05 or P<0.001), the expression of miR-138-3p was down-regulated (P<0.05). The apoptosis-related microRNAs miR-133c-3p and miR-149-5p were up-regulated (P<0.05 or P<0.001), and miR-138-3p was down-regulated (P<0.05). Among these, the expression of miR-133c-3p was upregulated almost 5 folds (P<0.01). The cell apoptosis rate was significantly increased after overexpression of miR-133c-3p (P<0.01) and the cell apoptosis rate was reduced after knock-down of miR-133c-3p (P<0.05). Bioinformatics methods were used to predict the binding sites between the miR-133c-3p and the 3'UTR of BCL2L2 gene. The result of luciferase reporter gene experiment showed that miR-133c-3p could bind to 3'UTR of BCL2L2 gene (P<0.01). The expression of BCL-w in cells was significantly down-regulated after over-expression of miR-133c-3p (P<0.01). PEDV could inhibit the expression level of BCL-w. Knock-down of BCL-w could induce cell apoptosis. PEDV infection could down-regulate the expression of BCL-w by up-regulating the expression level of miR-133c-3p, thereby promoting cell apoptosis.

Key words: porcine epidemic diarrhea virus, miR-133c-3p, BCL2L2, apoptosis

CLC Number:

S852.659.6

ZHENG Hongqing, WU Xujin, ZHU Xiaofu, YIN Baoying, GAO Junhua, LI Yanzhi, ZHI Chanping. Porcine Epidemic Diarrhea Virus Regulates Cell Apoptosis by miR-133c-3p/BCL2L2 Axis[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(8): 2233-2243.

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Porcine Epidemic Diarrhea Virus Regulates Cell Apoptosis by miR-133c-3p/BCL2L2 Axis

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