miR-128 Directly Represses L1 Retrotransposition, Cellular Targets in L1 Retrotransposition and HIV-1 Replication
microRNAs (miRs) are small endogenously encoded RNAs that post-transcriptionally repress gene expression by degradation or translational repression of target mRNAs. miRs are involved in normal development and homeostasis; dysregulation of miRs is tied to the development and progression of many diseases including cancer. Increasing evidence shows that miRs may have evolved to defend the human genome against mutagenesis by retroelements such as L1 retrotransposons or HIV-1.
L1 retrotransposons account for approximately 17% of the human genome and are the only autonomous transposons that replicate by a copy-paste mechanism. L1 retrotransposition is associated with mutagenesis and genomic instability, two factors implicated in cancer and various genetic disorders.
HIV-1 is a retrovirus that infects millions of people worldwide and can lead to fatal acquired immune deficiency syndrome (AIDS). Current anti-retroviral therapies are effective at reducing viral loads to non-detectable levels and prolonging the lifespan of patients; however, HIV-1 may remain latent in the genome and can be reactivated later.
The focus of this dissertation is to elucidate the mechanisms of miR-128-mediated repression of L1, its repression of cellular TNPO1 and hnRNP A1 that are involved in L1 retrotransposition, as well as cellular TNPO3 involved in HIV-1 replication. We identify a novel miR, miR-128, involved in L1 retrotransposition. We demonstrate that miR-128 represses L1 RNA, protein, and de novo retrotransposition by a direct binding mechanism in multiple cell lines including cancer initiating cells and induced pluripotent stem cells (iPSCs) (Chapter 2). miRs are highly pleiotropic with a single miR targeting multiple mRNAs in the same pathway. We show that TNPO1 is a cellular target of miR-128, and is necessary for efficient replication of L1 retrotransposons and nuclear import of L1 ORF1p (Chapter 3). We provide evidence that hnRNP A1 is also a direct target of miR-128 and repression of hnRNP A1 significantly reduces L1 RNA, protein and de novo L1 retrotransposition (Chapter 4). Lastly, we identify TNPO3 which is known to be involved in HIV-1 replication as a direct target of miR-128. Reduction of TNPO3 by miR-128 over-expression significantly reduces HIV-1 replication using a single cycle HIV-1 luciferase reporter virus (Chapter 5).