Investigation of the impact of miR-34a knock-out in HeLa cells.

Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus that infects approximately 10-20 million persons worldwide. A small percentage of HTLV-1-infected persons develop an aggressive hematologic neoplasm termed adult T-cell leukemia (ATL). Studies from our laboratory showed that HTLV-1-infected cells express increased levels of miR-34a-5p compared to normal T-cells and suggested that this microRNA may provide a survival advantage in the context of HTLV-1-infection. This proposed role contrasts with the generally accepted identification of miR-34a-5p as a tumor suppressor microRNA. To extend these studies, our laboratory obtained HeLa clones knocked-out (KO) for miR-34a expression using the CRISPR-Cas9 technique. The present thesis describes a characterization of the miR-34a-KO clones with control HeLa cells by comparing their proliferation rates, cell cycle, metabolism, and gene expression profiles. Results indicated that ablation of miR-34a: (1) reduced cell viability as detected using the MTT assay; (2) increased the levels of the NAD-dependent deacetylase SIRT1; and (3) modified the expression of >100 mRNAs as detected by RNAseq. These findings suggest that further analysis of the miR-34a-KO clones may uncover new roles for miR-34a-5p in the context of HTLV-1 infection.