Assembly and Disassembly Processes in the HIV-1 Life Cycle - PROJECT SUMMARY HIV-1 must assemble and disassemble into different morphologies throughout its life cycle to infect new cells and replicate. Some examples of this include assembly and budding of the immature virion, maturation of the viral core, and uncoating. Inhibiting or otherwise disrupting these transitions, which are critical for viral infectivity, is an effective strategy in the design of antiretroviral therapies. For example, a class of drugs known as allosteric integrase inhibitors disrupts maturation by preventing packaging of viral RNA in the mature core resulting in a loss of infectivity. A full understanding of the process by which viral RNA is packaged would greatly aid in a mechanistic understanding of how these drugs work. This mechanistic understanding would also aid in the design of future therapeutics targeting RNA packaging just as would for drugs targeting budding and uncoating. In this work, I propose to computationally investigate these three morphological transitions in HIV, focusing on the role of factors beyond the capsid domain proteins including RNA, host cell factors, and potential therapeutics. After expression by an infected cell, Gag polyproteins assemble at the plasma membrane into the 100nm, quasi- spherical immature virion that buds outward from the plasma membrane. The formation and budding of the virion were thought to be driven by interactions of the Gag CA domain, however, recent experiments paint a more complex picture. I will investigate the role of membrane and membrane bending IBAR protein IRSp53 in budding and assembly of the immature virion. I will incorporate IRSp53 into a coarse-grained (CG) model, which could not previously generate the curvature necessary for budding to occur. The maturation of the virion begins when HIV-1 protease cleaves the Gag polyprotein, freeing its different domains such as the capsid (CA) domain. CA then assembles into the mature, conical capsid which packages the ribonucleoprotein complex (RNP) composed of viral RNA, Integrase (IN), and nucleocapsid (NC) protein among other components. Using CG models of different resolution, I will investigate how the polymer-like RNP affects assembly of the mature focusing on the physical properties necessary for encapsulation of the RNP. The final CG model will be built based off experimental data showing contacts between RNA, NC, and IN with and without IN. Following import of the mature capsid into the nucleus, the mature capsid uncoats and releases the viral DNA for integration. Uncoating occurs as a result of reverse transcriptase transcribing viral RNA into more rigid DNA. This creates on outward pressure on the capsid which leads to makes it unstable. I will investigate the pathway of uncoating using a UCG model of reverse transcription that changes the rigidity of the RNP and a bottom-up CG model of the mature capsid. This model will include host factors such as cleavage and polyadenylation specificity factor 6 (CPSF6) as well as potentially therapeutic molecules that rigidify the capsid, possibly inhibiting the rupture of the capsid and release of DNA.
