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CircRNA.0007127 triggers apoptosis through the miR-513a-5p/CASP8 axis in k-562 cells

Journal of Zhejiang University-Science B(Biomedicine & Biotechnology) 2022年第9期23卷 732-746页

作者： Xiajing LI Yiyu ZHANG Ning WANG Zhaohu YUAN Xiaojie CHEN Qicong CHEN Hui DENG Xinxin TONG Honglin CHEN Yuyou DUAN Yaming WEI School of Medicine South China University of TechnologyGuangzhou510000China Department of Blood Transfusion the Second Affiliation Hospital of South China University of TechnologyGuangzhou510000China Guangdong Engineering Research Center of Precise Transfusion Guangzhou510000China Department of Blood Transfusion Shenzhen Longhua Central HospitalShenzhen518000China Laboratory of Stem cells and Translational Medicine Institutes for Life SciencesSchool of MedicineSouth China University of TechnologyGuangzhou510000China School of Biomedical Sciences and Engineering Guangzhou International CampusSouth China University of TechnologyGuangzhou510000China

Background:Circular RNAs(circR NAs)are covalently closed single-stranded RNAs with multiple biological functions.CircRNA.0007127 is derived from the carbon catabolite repression 4-negative on TATA-less(CCR4-NOT)complex subunit2(CNOT2),which was found to regulate tumor cell apoptosis through caspase pathway.Methods:Potential circR NA.0007127 target microRNAs(miRNAs)were analyzed by miRanda,TargetScan,and RNAhybrid software,and the miRNAs with binding sites for apoptosis-related genes were screened.The roles of circR NA.0007127 and its downstream target,microR NA(miR)-513a-5p,were validated by quantitative real-time polymerase chain reaction(qPCR),flow cytometry,mitochondrial membrane potential,immunofluorescence,western blot,and caspase-8(CASP8)protein activity in vitro in HO-induced k-562 cells.The circRNA.0007127-miR-513a-5p and CASP8-miR-513a-5p interactions were verified by luciferase reporter assays.Results:Silencing circRNA.0007127 decreased cell apoptosis by inhibiting CASP8 pathway activation in k-562 cells.Compared with the control group,the expression of CASP8 was reduced by 50%and the 43-kD fragment of CASP8 protein was significantly reduced(P≤0.05).The luciferase reporting assay showed that circRNA.0007127 combined with miR-513a-5p or CASP8,with extremely significant differences(P≤0.001).The overexpression of miR-513a-5p inhibited the gene expression level of CASP8in a human myeloid leukemia cell model(75%change)and the level of a 43-kD fragment of CASP8 protein(P small-interfering RNA(siRNA)and the miR-≤0.01).The rescue experiment showed that cotransfection with circRNA.0007127513a-5p inhibitor increased CASP8 gene expression and the apoptosis rate,suggesting that the miR-513a-5p inhibitor is a circRNA.0007127siRNA antagonist.Conclusions:CircRNA.0007127 regulates k-562 cell apoptosis through the miR-513a-5p/CASP8 axis,which can serve as a novel powerful molecular target for k-562 cells.

关键词： CircRNA.0007127 miR-513a-5p Caspase-8(CASP8) Apoptosis k-562 cells

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