Identification of novel molecular mechanisms of HIV latency using forward genetic screens (360G-Wellcome-102199_Z_13_Z)

The latent viral reservoir remains the major block to HIV eradication and transcriptional repression is thought to be the predominant mechanism of silencing latent HIV.However, multiple mechanisms of repression are likely to be responsible and finding a cure for HIV is clearly dependent upon an improved understanding of the molecular mechanisms of latent HIV infection. Here, I propose to use forward genetic screens to identify novel genes required for HIV silencing. I will use an HIV-based GFP reporter construct to establish a model of HIV latency in the human near-haploid KBM7 cell line. The KBM7 cells will be mutagenised with a retroviral gene-trap vector, rare GFP-expressing clones will be isolated, and their mutation sites mapped using next generation sequencing. Candidate genes will then be validated in more representative HIV model systems and biochemically characterised to elucidate their mechanism of HIV silencing.