In this era in which antiretroviral drugs are being used for both treatment and prevention of AIDS, there is an urgent need for discovery of new drug targets and novel antiretroviral agents. Virus assembly is a complex stage of the HIV-1 life cycle that contains poorly understood drug targets. These targets are understudied, in part because of unresolved controversies in the HIV-1 assembly field. The field has been dominated by the self-assembly model, which is based on the finding that HIV-1 Gag assembles spontaneously in an idealized in vitro system that does not recapitulate the intracellular environment. In contrast, this application is based on a model proposing that the intracellular environment presents barriers to assembly, which the virus overcomes using cellular enzymes. This model is supported by studies demonstrating that during immature capsid assembly, HIV-1 Gag progresses through a stepwise, energy-dependent pathway of assembly intermediates composed of cellular proteins, including two cellular enzymes that facilitate assembly, the ATPase ABCE1 and the RNA helicase DDX6. Thus, while HIV-1 can assemble spontaneously in an idealized in vitro system, studies in cells indicate that HIV-1 capsid assembly is facilitated by at least two host enzymes. Further evidence in favor of the host-catalyzed ABCE1 assembly pathway comes from the discovery by Prosetta Antiviral of novel antiretroviral compounds that target this pathway. This summer Bristol Myers Squibb announced a partnership with Prosetta Antiviral to develop these novel antiretroviral compounds. Despite this breakthrough in the area of drug development, the ABCE1 pathway remains poorly understood. Studies in Aim 1 will test whether ABCE1 acts as an adaptor for recruiting cellular machinery and will also define the Gag-ABCE1 binding site, which appears to involve the highly conserved major homology region of Gag.
Aim 2 will help establish a unified model of assembly by determining whether the Gag-containing complex that first associates with HIV-1 genomic RNA, identified by Paul Bieniasz's lab, corresponds to an ABCE1- containing assembly intermediate.
Aim 3 will uses viruses generated by Eric Hunter's group to examine whether Gag polymorphisms that arise in vivo can alter viral host interactions, leading to increased virus assembly kinetics, greater virus production, and higher viral loads in infected individuals. Identifying viral-host interactions that impact virus production could lead t strategies for targeting such interactions in the future. In sum, this "bench to bedside" application will advance our understanding of a cellular facilitator of assembly that binds to Gag, help reach a consensus model of HIV-1 capsid packaging, and provide insights into how viral-host interactions in assembly affect pathogenesis.
HIV-1 capsid assembly is a poorly understood but critical stage of the HIV-1 life cycle. Studies proposed here will advance understanding of a poorly understood pathway of viral-host interactions that are critical for immature HIV-1 capsid assembly, and are targeted by drug-like compounds that inhibit HIV-1 replication and are currently being advanced by pharma.
|Robinson, Bridget A; Reed, Jonathan C; Geary, Clair D et al. (2014) A temporospatial map that defines specific steps at which critical surfaces in the Gag MA and CA domains act during immature HIV-1 capsid assembly in cells. J Virol 88:5718-41|
|Lingappa, Jaisri R; Reed, Jonathan C; Tanaka, Motoko et al. (2014) How HIV-1 Gag assembles in cells: Putting together pieces of the puzzle. Virus Res 193:89-107|