Determinants of HIV-1 Restriction by the A3G Complex
Mario Santiago, J. David Gladstone Institutes
Biomedical and Clinical Sciences
Background: The dynamic interplay between viral proteins and innate host factors represent promising avenues for novel antiviral intervention strategies. Recently, our laboratory found that A3G is a potent restriction factor for HIV-1 infection of restingT-cells, and this antiviral activity correlated with its recruitment into low- (LMM) and high-molecular mass (HMM) complexes (Chiu et al. 2005 Nature 435(7038):108-14).The LMM complex functions as a potent antiretroviral barrier in unstimulated T cells, but upon T cell activation, A3G is recruited into an enzymatically-inactive HMM ribonucleoprotein complex that is incapable of restricting single-round HIV-1 infection. Intriguingly, the ability of theA3G LMM complex to block HIV-1 infection in resting T-cells does not seem to solely depend on its deaminase activity.
Methods: To investigate the structural and functional determinants ofA3G that govern HMM complex formation and HIV-1 restriction, we propose to generate a panel of A3G mutants that takes into account the potential functional redundancy imposed by A3G's double-domain structure. These 'double-mutants' will then be characterized for their ability to form HMM complexes and mediate the HIV-1 post-entry block in permissive cells. These properties will then be correlated with RNA binding, deaminase activity, dimer formation and ability to form cytoplasmic bodies.
Expected Results: We hope to identify A3G mutations that will disrupt HMM complex formation. Such mutants that constitutively express in the LMM form may effectively restrict HIV-1 infection in permissive cells. If so, the properties of these mutants that facilitate the antiviral activity will be addressed by further mutagenesis.
Conclusions: Unraveling the structural domains of A3G that govern HMM complex formation and HIV-1 restriction may provide a much-needed molecular blueprint for exploiting this innate restriction factor as a therapeutic agent against HIV-1.