Regulation of Human Immunodeficiency Virus Type 1 Envelope Glycoprotein Fusion by a Membrane-Interactive Domain in the gp41 Cytoplasmic Tail

Abstract

Truncation of the human immunodeficiency virus (HIV) or simian immunodeficiency virus (SIV) gp41cytoplasmic tail (CT) can modulate the fusogenicity of the envelope glycoprotein (Env) on infected cells and virions. However, the CT domains involved and the underlying mechanism responsible for this "inside-out" regulation of Env function are unknown. HIV and SIV CTs are remarkably long and contain amphipathic alpha-helical domains (LLP1, LLP2, and LLP3) that likely interact with cellular membranes. Using a cell-cell fusion assay and a panel of HIV Envs with stop codons at various positions in the CT, we show that truncations of gp41 proximal to the most N-terminal alpha helix, LLP2, increase fusion efficiency and expose CD4-induced epitopes in the Env ectodomain. These effects were not seen with a truncation distal to this domain and before LLP1. Using a dye transfer assay to quantitate fusion kinetics, we found that these truncations produced a twoto fourfold increase in the rate of fusion. These results were observed for X4-, R5-, and dual-tropic Envs on CXCR4-and CCR5-expressing target cells and could not be explained by differences in Env surface expression. These findings suggest that distal to the membrane-spanning domain, an interaction of the gp41 LLP2 domain with the cell membrane restricts Env fusogenicity during Env processing. As with murine leukemia viruses, where cleavage of a membrane-interactive R peptide at the C terminus is required for Env to become fusogenic, this restriction of Env function may serve to protect virus-producing cells from the membrane-disruptive effects of the Env ectodomain.

Surface Exposure of the HIV-1 Env Cytoplasmic Tail LLP2 Domain during the Membrane Fusion Process

“…The HIV-1 gp41 CT consists of the "Kennedy" polypeptide sequence (aa 731-752) and the lentivirus lytic peptide (LLP1-2, aa 773-862), which contains three highly conserved ␣-helix domains as follows: LLP1 (aa 833-862), LLP2 (aa 773-793), and LLP3 (aa785-807) (11). The HIV-1 gp41 CT has many functions, including interaction with the plasma membrane, decrease of bilayer stability, alteration of membrane ionic permeability, and mediation of cell killing (10,(12)(13)(14)(15)(16). Recent studies have shown that the gp41 CTs of HIV and simian immunodeficiency virus correlate with their infectivity and Env-mediated cell-cell fusion (10,15).…”

“…The HIV-1 gp41 CT has many functions, including interaction with the plasma membrane, decrease of bilayer stability, alteration of membrane ionic permeability, and mediation of cell killing (10,(12)(13)(14)(15)(16). Recent studies have shown that the gp41 CTs of HIV and simian immunodeficiency virus correlate with their infectivity and Env-mediated cell-cell fusion (10,15). The mutations or truncations of gp41 CT can affect HIV-1 Env-mediated cell-cell fusion, presumably because of modulation of the fusogenicity of Env (17,18).…”

“…However, in some other studies, certain point mutations and particularly truncations of gp41 CT were shown to induce faster fusion kinetics (19 -22). Recently, Wyss et al (10) have also demon-* This work was supported, in whole or in part, by National Institutes of Health Grant AI46221. This work was also supported by the Emphases Project of NSFC-30530680 and 973-2006CB504203 (to Y. H. C.) and the Outstanding Overseas Chinese Scholars Fund of Chinese Academy of Sciences Grant 2005-2-6 (to S.…”

“…However, existing experimental and computational evidence suggest that the Kennedy sequence might lie outside the cell, because the antibodies against the Kennedy sequence can bind to virus-infected cells and neutralize HIV-1 infectivity (23,24). Our study focuses on the gp41 CT LLP1-2 region, especially the LLP1 and LLP2 regions that have been directly proven to be associated with the membrane fusion process (10,15), and aims to determine whether LLP1-2 directly or indirectly participates in the fusion reaction and, if so, to understand the underlying mechanism(s) of this event. Here we report, for the first time, that during the HIV-1 fusion process, the LLP1-2 region of gp41 CT, in particular the LLP2 domain, may be transiently exposed on the surface of the effector cell in the presence of the target cell.…”

HIV-1 envelope glycoprotein-mediated fusion and pathogenesis: Implications for therapy and vaccine development

“…Using a cell-cell fusion assay and a panel of HIV Envs with stop codons at various positions in the CT, we showed that truncations of gp41 proximal to the most N-terminal alpha helix, LLP2 (lentivirus lytic peptide), increase fusion efficiency and expose CD4-induced epitopes in the envelope ectodomains [34]. These effects were not seen with a truncation distal to this domain and before LLP1.…”