Résumé
Objective To investigate the effect of SARS-CoV-2 membrane protein on the processing of the 3' untranslated region (UTR) of the mRNA precursor (pre-mRNA) in host cells. Methods Based on the cell model of human lung epithelial cells A549, over-expression of the SARS-CoV-2 membrane protein was performed. The RNA-Seq high-throughput sequencing technique and bioinformatics methods was employed to analyze the systematic characterization of alternative polyadenylation (APA) events in host cells. Genes with significant APA events were uploaded to the Metascape database for functional enrichment analysis. In addition, alternative 3'UTR length of genes with APA events was verified by RT-qPCR. Then the target protein expression level was detected by Western blot. Results A total of 813 genes that were significant dynamic APA events in host cells that over-expressed SARS-CoV-2 membrane protein. These genes were enriched in cell biologicial processes such as the mitotic cell cycle and regulation of cellular response to stress. We further screened AKT1, which encodes a critical regulator involved in the above biological process, showing a 3'UTR lengthening in IGV software. RT-qPCR verified the trend of 3'UTR length changes of AKT1. Western blot showed the increased level of phosphorylated AKT1 protein in over-expressed group of M protein. Conclusion SARS-CoV-2 membrane protein potentially affects the 3' processing of host pre-mRNAs. AKT1, which is involved in a variety of viral biological processes, with 3'UTR lengthening, and its protein function was activated intracellularly. © 2022 Institute of Biophysics,Chinese Academy of Sciences. All rights reserved.
Résumé
Objective To investigate the effect of SARS-CoV-2 membrane protein on the processing of the 3' untranslated region (UTR) of the mRNA precursor (pre-mRNA) in host cells. Methods Based on the cell model of human lung epithelial cells A549, over-expression of the SARS-CoV-2 membrane protein was performed. The RNA-Seq high-throughput sequencing technique and bioinformatics methods was employed to analyze the systematic characterization of alternative polyadenylation (APA) events in host cells. Genes with significant APA events were uploaded to the Metascape database for functional enrichment analysis. In addition, alternative 3'UTR length of genes with APA events was verified by RT-qPCR. Then the target protein expression level was detected by Western blot. Results A total of 813 genes that were significant dynamic APA events in host cells that overexpressed SARS-CoV-2 membrane protein. These genes were enriched in cell biologicial processes such as the mitotic cell cycle and regulation of cellular response to stress. We further screened AKT1, which encodes a critical regulator involved in the above biological process, showing a 3'UTR lengthening in IGV software. RT-qPCR verified the trend of 3'UTR length changes of AKT1. Western blot showed the increased level of phosphorylated AKT1 protein in over-expressed group of M protein. Conclusion SARS-CoV-2 membrane protein potentially affects the 3' processing of host pre-mRNAs. AKT1, which is involved in a variety of viral biological processes, with 3'UTR lengthening, and its protein function was activated intracellularly.