Abstract
BackgroundInfection with human immunodeficiency virus type-1 (HIV-1) requires binding of the viral envelope gp120 to CD4 and to the CXCR4 coreceptor. Both, gp120 and CXCR4 are subject to N-glycosylation. Here we investigated the influence of the N-linked glycans g1 and g2 present on CXCR4 for HIV-1 infection.MethodsThe two CXCR4 N-glycosylation sites g1 (NYT) and g2 (NVS) were mutated by changing the first or third amino acids N or T/S to Q and A respectively (g1; N11Q or T13A; g2, N176Q or S178A). Human osteosarcoma cells (GHOST) expressing human CD4 and the various CXCR4 glycosylation mutants were tested for infection using NL4-3-based viruses with X4, R5 or R5X4-tropism differing only in the V3 loop region.ResultsAll constructed cell lines expressing the various CXCR4 glycomutants showed similar permissiveness for the X4-monotropic virus and no change in the coreceptor specificity that allows infection of a CCR5-dependent R5-monotropic virus. Interestingly, the removal of glycan g1 significantly enhanced the permissiveness of GHOST cells for the R5X4 dualtropic virus. GHOST cells expressing the CXCR4-g1 or CXCR4-g1/2 mutants were infected at higher rates by the R5X4-dualtropic virus compared to cells expressing CXCR4-wt or CXCR4-g2 coreceptors.ConclusionOur present observations underscore a role for glycans present on the CXCR4 coreceptor in the entry process of HIV-1. The data will help to better understand the multifaceted mechanism of HIV infection and the selective forces which drive HIV-1 evolution from mono- to dual-tropism.
Highlights
Infection with human immunodeficiency virus type-1 (HIV-1) requires binding of the viral envelope gp120 to CD4 and to the CXCR4 coreceptor
We have previously shown that the lack of some N-glycan structures within the gp120 V3 loop of the HIV-1NL4–3 strain results in enhanced infectivity for CXCR4 expressing cells and in increased resistance to the inhibitor SDF1α. [15]
To generate mutants of CXCR4 differing in Nglycosylation we have introduced point mutations changing the coding sequence for asparagine, threonine or serine into the coding sequence for glutamine and alanine (Fig. 1b)
Summary
Infection with human immunodeficiency virus type-1 (HIV-1) requires binding of the viral envelope gp120 to CD4 and to the CXCR4 coreceptor. The chemokine receptor CXCR4 belongs to the seventransmembrane-domain G-protein-coupled receptor family and is one of the major coreceptors for human immunodeficiency virus type 1 (HIV-1) [1,2]. The trimolecular interaction between the HIV-1 receptor CD4, CXCR4 and viral envelope proteins (gp120/gp41) is the first step in HIV entry. The N-terminal region and extra cellular domains of CXCR4 are of particular importance for the interaction with the third variable loop (V3) of (page number not for citation purposes). While HIV-1 infection is dependent on the interaction between at least three different molecules, viral entry can be influenced by changes in the V3 region of gp120 or changes in the N-terminus and in the second extracellular loop (ecl-2) of CXCR4. While HIV-1 infection is dependent on the interaction between at least three different molecules, viral entry can be influenced by changes in the V3 region of gp120 or changes in the N-terminus and in the second extracellular loop (ecl-2) of CXCR4. [9,10,11,12]
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