Human Cytomegalovirus Mir-UL70-3p Downregulates the H2O2 Induced Apoptosis in HEK293T Cells by Targeting the Modulator of Apoptosis-1 (MOAP1)
Abstract Human Cytomegalovirus (HCMV), a prototypic member of the Beta-herpesvirinae subfamily, mainly responsible for congenital disabilities in newborns, cause opportunistic infections in immunocompromised individuals. Its seroprevalence varies across the globe ranging from 50–70 percent in developed countries to 90–100 percent in developing countries. Causing latent infections in the immunocompetent host suggests that it employs several strategies to escape the wrath of the host’s antiviral mechanisms. Apoptosis is an innate cellular response to viral infection, and downregulation of this mechanism by HCMV is a well-established phenomenon. HCMV utilizes its proteins, RNA and miRNA in regulating this response to establish a productive infection in the host. The role of HCMV miRNAs on cellular apoptosis is very interesting, where some miRNAs downregulate but a few upregulate this process. In the present study, we report the antiapoptotic activity of HCMV miRNA, miR-UL-70-3p, on H2O2 induced apoptosis in HEK293T cells. The ectopic expression of this HCMV miRNA in HEK293T cells downregulate the apoptosis, and continuing studies reveal that the proapoptotic gene, Modulator of Apoptosis Protein-1 (MOAP1), is a functional target for this miRNA. We verified the functionality of the binding site predicted in the 3'UTR of MOAP1 in our earlier studies through dual luciferase-based assays using both the wild and mutant 3'UTR’s of MOAP1. The MOAP1 protein levels were significantly downregulated by the miR-UL70-3p, suggesting that the MOAP1 mRNA was degraded after binding with the miR-UL70-3p. Further, the extent of MOAP1 mRNA and its protein inhibitions by HCMV miR-UL70-3p and siRNA of MOAP1 were compared and found that the siRNA of MOAP1 inhibits 69.52 percent of mRNA and 35.67 percent of MOAP1 protein; while the miR-UL70-3p inhibits 46.66 percent MOAP1 mRNA and 21.05 percent MOAP1 protein. Though the inhibitory activity of miR-UL70-3p is not equal to the siRNA of MOAP1, but it was significant enough in reversing the H2O2 induced apoptosis in HEK293T cells. These results confirm that hcmv-miR-UL70-3p downregulates H2O2 induced apoptosis in HEK293T cells by targeting the 3'UTR of MOAP1 mRNA.