HIV‐1 Capsid Binding Host Factor CPSF6 is Post Transcriptionally Regulated by the Cellular MicroRNA miR‐125b

Abstract

The cleavage and polyadenylation specificity factor 6 (CPSF6) is a cellular protein involved in RNA cleavage and polyadenylation. Emerging evidence also suggest that CPSF6 plays a key role during HIV‐1 infection by guiding integration of the viral DNA into gene‐dense regions of the host genome. Since viral DNA integration is a critical step of HIV‐1 infection, the role of CPSF6 in the virus lifecycle is being intensely investigated. Surprisingly, the cellular mechanisms that regulate CPSF6 expression are largely unknown. In this study, we report a post‐transcriptional mechanism of regulation of CPSF6. Our initial bioinformatics analysis revealed that the 3′ untranslated region (3′UTR) of Cpsf6 contains a binding site for the cellular miRNA miR‐125b that is strikingly conserved across different mammalian species. Since miRNAs negatively regulate protein expression, we carried out knock‐down and over‐expression studies of miR‐125b. Results from these experiments revealed that miR‐125b expression is negatively associated with CPSF6 protein levels. Interestingly, HIV‐1 infection resulted in the down‐regulation of miR‐125b concurrent with induction of CPSF6. To probe that CPSF6 expression is post‐transcriptionally regulated by miR‐125b, we cloned the 3′UTR regions of Cpsf6 mRNA into a luciferase reporter. Results from luciferase assay provide evidence that miR‐125b expression negatively regulates Cpsf6 3′UTR activity. Accordingly, mutations in the miR‐125b seed sequences abrogated the regulatory effect of the miRNA on Cpsf6 3′UTR. Pull‐down studies demonstrated that miR‐125b physically interacts with the Cpsf6 mRNA. Continuing studies probe the necessity of productive HIV‐1 infection for negatively regulating miR‐125b expression as well as to elucidate the pathway through which HIV‐1 infection can knockdown miR‐125b expression. Collectively, these findings establish a post‐transcriptional mechanism of CPSF6 expression and describe a novel function of miR‐125b in virus‐host interaction.Support or Funding InformationThis work is partly supported by grants DA024558, DA30896, DA033892 and DA021471 from NIDA/NIH to CD. We also acknowledge the RCMI Grant G12MD007586, the Vanderbilt CTSA grant UL1RR024975, the Meharry Translational Research Center (MeTRC) CTSA grant (U54 RR026140 from NCRR/NIH, the U54 grant MD007593 from NIMHD/NIH, and Tennessee CFAR grant (P30 AI110527).