Abstract
The restriction factor BST2 (tetherin) prevents the release of enveloped viruses from the host cell and is counteracted by HIV-1 Vpu. Vpu and BST2 interact directly via their transmembrane domains. This interaction enables Vpu to induce the surface down-regulation and the degradation of BST2, but neither of these activities fully accounts for the ability of Vpu to enhance virion release. During a study of naturally occurring Vpu proteins, we found that a tryptophan residue near the Vpu C terminus is particularly important for enhancing virion release. Vpu proteins with a W76G polymorphism degraded and down-regulated BST2 from the cell surface, yet they inefficiently stimulated virion release. Here we explore the mechanism of this anomaly. We find that Trp-76 is critical for the ability of Vpu to displace BST2 from sites of viral assembly in the plane of the plasma membrane. This effect does not appear to involve a general reorganization of the membrane microdomains associated with virion assembly, but rather is a specific effect of Vpu on BST2. Using NMR spectroscopy, we find that the cytoplasmic domain of Vpu and Trp-76 specifically interact with lipids. Moreover, paramagnetic relaxation enhancement studies show that Trp-76 inserts into the lipid. These data are consistent with a model whereby Trp-76 anchors the C terminus of the cytoplasmic tail of Vpu to the plasma membrane, enabling the movement of Vpu-bound BST2 away from viral assembly sites.
Highlights
HIV-1 Vpu displaces bone marrow stromal antigen 2 (BST2) from sites of viral assembly
As BST2 is induced by interferon in response to HIV infection in vivo [32], we first evaluated the phenotypes of the Vpu-W76G mutant in the setting of the high levels of BST2 expression induced by ␣-interferon in vitro
The combined mutant Vpu-S52N,S56N ϩ W76G was as impaired for virion release as the no Vpu mutant (⌬Vpu), indicating that the S52N,S56N and W76G mutations have additive effects and that these regions of Vpu likely contribute to its function by distinct mechanisms
Summary
HIV-1 Vpu displaces bone marrow stromal antigen 2 (BST2) from sites of viral assembly. To elaborate the mechanism of the displacement effect, we show using NMR spectroscopy that the Vpu cytoplasmic domain interacts with lipids, and we support a model in which Trp-76 anchors the C-terminal region of the protein to the lipid bilayer This interaction was enhanced in the presence of lipid head groups of net positive charge, phenylalanine did not substitute functionally for tryptophan at this position; this result suggests that the ability of tryptophan to bind lipid cannot be explained fully by a cation- interaction but likely depends on the hydrophobic nature of the residue. We propose a refined topological model for the counteraction of BST2 by Vpu: tryptophan-mediated anchoring of the Vpu C terminus to the lipid bilayer enables Vpu to displace BST2 from virion-assembly sites
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