A deep mutational scan of the HIV-1 capsid core.

About This Grant

Abstract. Human immunodeficiency virus type 1 (HIV-1) facilitates viral egress and ingress via the activity of the polyprotein, Gag. Gag is the sole facilitator of the identification, organization, and sequestration of the viral genome from the host cytoplasm to the plasma membrane, nucleating the assembly of a nascent virion. To produce infectious progeny, this process must occur with remarkable efficiency and specificity, despite the tremendous diversity among viral genomes. After budding, Gag is cleaved into its constituent domains by the viral protease, facilitating the intra-virion assembly of the conical viral capsid (CA) core (Core) and the compaction of the viral RNA genome by the viral nucleocapsid (NC). Upon entry, the Core is released into the cytoplasm and must translocate to the nucleus, traverse the nuclear pore, facilitate reverse transcription, while also shielding the viral genome and its reverse transcription products from the host immune system. The precise determinants and genetic constraints of infectivity by HIV-1 Cores remain incompletely identified and even less is known about HIV-1’s lentiviral relatives. The main goal of this project is to apply deep mutational scanning methodologies to understand the basic biology of the HIV-1 Core and potential heterogeneity therein. In the laboratory setting, single nucleotide substitutions within protein coding and non-coding regions of the HIV-1 genome have been demonstrated to have dramatic effects on viral fitness. Despite this, HIV-1 produces enough infectious progeny to remain viable in most infected individuals. And despite remarkable improvements in our ability to detect mutations in trace amounts of viral genetic material, the precise consequences of mutations within the HIV-1 genome have yet to be comprehensively investigated at scale. Therefore, Specific Aim 1 will seek to Identify the genetic determinants of HIV-1 Core assembly and infectivity. Similarly, Specific Aim 2 will extend this analysis to interrogate divergent lentiviral Cores, to characterize the more general constraints of lentiviral Core form, function, and adaptation to new intracellular barriers to infection. The results of this work could have implications for our basic knowledge of the cell biology of HIV-1 infection, the clinical treatment of HIV-1 infection, the prediction of drug-resistance mutations by HIV-1, and the development of next-generation lentiviral vectors for various applications in the laboratory and clinic. With the mentorship of my sponsor and thesis committee, the leadership of the Tri-I MD-PhD program, and the support of this fellowship, I believe that conducting the work described here will prepare leave me well prepared to take the next steps in pursuing a career as a physician-scientist.