In order for an HIV-1 virion to infect a host cell, the gp120 subunit of envelope, the only viral protein exposed on the surface of the virion, first attaches to the cellular receptor CD4 and subsequently to a chemokine coreceptor, either CCR5 or CXCR4. Tropism for these coreceptors is principally determined by the V3 loop, which is a 35-residue region bound by a disulphide bond and exposed on the surface of gp120. While V3 is known to be critical for coreceptor binding, V3-coreceptor interactions and the structural basis for CCR5 or CXCR4 specificity remain elusive. We hypothesized that subdomains within V3 determine coreceptor tropism. Using the dual-tropic virus HIV-1R3A, which uses both CCR5 and CXCR4, we made an extensive panel of small V3 deletions and evaluated these mutants for their effects on tropism and sensitivity to coreceptor antagonists. Small deletions on either side of the V3 base abrogated R5 tropism. The resulting X4-tropic mutants exhibited enhanced sensitivity to the CXCR4 antagonist AMD3100. Only a deletion of residues 9 to 12 in the V3 stem ablated X4 tropism. Remarkably, this R5-tropic mutant showed complete resistance to a panel of CCR5 antagonists, including vicriviroc and aplaviroc. Our findings suggest that subdomains in V3 differentially affect R5 and X4 tropism and modulate sensitivity to coreceptor antagonists.
Small molecule CCR5 inhibitors are being used clinically to treat HIV-1-infected patients, so in order to study mechanisms of resistance to this new drug class, we further characterized a variant of the drug-resistant Δ9-12 mutant, termed Δ9-12a. Since CCR5 antagonists disrupt the gp120-CCR5 extracellular loop interaction, we hypothesized that Δ9-12a is more dependent on the CCR5 amino terminus (NT). Our results indicate that Δ9-12a does have an increased reliance on the CCR5 NT, exhibits increased sensitivity to several broadly neutralizing monoclonal antibodies, including b12 and 4E10, and is capable of using aplaviroc-bound CCR5. This detailed characterization suggests that increased dependence on the CCR5 NT may represent a mechanism by which HIV-1 Envs acquire resistance to CCR5 antagonists. In addition, envelopes such as Δ9-12a may be useful for identifying new drugs that target the gpl20-CCR5 NT interaction.
|Advisor:||Hoxie, James A.|
|School:||University of Pennsylvania|
|School Location:||United States -- Pennsylvania|
|Source:||DAI-B 69/09, Dissertation Abstracts International|
|Keywords:||CCR5 antagonists, Coreceptor inhibitors, HIV-1, Tropism, V3 loop|
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