No abstract

The major surface glycoprotein of feline immunodeficiency virus (FIV) specifically binds to a 43-kDa glycoprotein expressed on the surface of a subset of T cells in peripheral blood mononuclear cells and IL-2-dependent T cell lines. Binding to this molecule, in conjunction with CXC chemokine receptor (CXCR) 4, is required for productive infection of these cells by primary isolates of FIV. Here, we demonstrate that the 43-kDa molecule is CD134, a receptor for FIV recently identified independently [Shimojima, M., et al. (2004) Science 303, 1192-1195]. Furthermore, we show that CD134 is specifically up-regulated on CD4 + T cells that have been activated by treatment with IL-2 and Con A. CD8 + T cells remained negative for CD134 expression regardless of the activation state. Binding of the FIV major surface glycoprotein on activated CD4 + T cells was observed through direct interaction with CD134 whereas, on activated CD8 + T cells, the binding was CD134-independent and mediated by CXCR4 and, to a lesser extent, heparan sulfate proteoglycans. However, this CD134-independent interaction was not sufficient to render CD8 + T cells permissive to FIV infection, as FIV replicated primarily in activated CD4 + T cells and not in cells negative for CD134 expression. Altogether, our results substantiate that CD134 acts as a primary binding receptor for FIV and explain the specific targeting and depletion of the CD4 + T cell population observed during the course of infection independent of the use of CD4 as a binding receptor/coreceptor.

The calcium-dependent lectin, DC-SIGN, binds to human immunodeficiency virus (HIV) (and simian immunodeficiency virus) gp120 and mediates the binding and transfer of HIV from monocyte-derived dendritic cells (MDDCs) to permissive T cells. However, it has been recently reported that DC-SIGN binding to HIV gp120 may be carbohydrate independent. Here, we formally demonstrate that gp120 binding to DC-SIGN and MDDCs is largely if not wholly carbohydrate dependent. Endo-␤-N-glucosaminidase H (EndoH) treatment of gp120-Fc under conditions that maintained wild-type CD4 binding-and the full complement of complex glycans-significantly decreased (>90%) binding to DC-SIGN expressing cell lines, as well as to MDDCs. Any residual binding of EndoH-treated gp120-Fc to DC-SIGN was completely competed off with mannan. Mutational analysis indicated that no single glycosylation site affected the ability of gp120-Fc to bind DC-SIGN. To further guide our efforts in mapping the DC-SIGN binding sites on gp120, we used two well-characterized HIV inhibitory agents (2G12 monoclonal antibody and cyanovirin) that bind to high-mannose sugars on gp120. We showed that 2G12 and DC-SIGN bound to nonoverlapping sites in gp120 because (i) 2G12 did not block soluble gp120 or virion binding to DC-SIGN, (ii) 2G12 bound to gp120-Fc that was prebound to cell surface DC-SIGN, and (iii) gp120-Fc mutants that lack glycosylation sites involved in 2G12's epitope were also fully capable of binding DC-SIGN. These data were substantiated by the inability of cyanovirin to block gp120-Fc binding to DC-SIGN. Cyanovirin has been shown to effectively compete for 2G12 binding to gp120. Indeed, high concentrations of cyanovirin dramatically enhanced gp120-Fc binding to cell surfaces in the presence or absence of DC-SIGN. We provide evidence that this enhancement may be due to cyanovirin's ability to bridge gp120 to mannosylated cell surface proteins. These results have implications for antiviral therapeutics and for ongoing efforts to finely map the glycan structures on gp120 responsible for DC-SIGN binding.

HIV-1 has maximized its utilization of syndecans. It uses them as in cis receptors to infect macrophages and as in trans receptors to infect T-lymphocytes. In this study, we investigated at a molecular level the mechanisms that control HIV-1-syndecan interactions. We found that a single conserved arginine (Arg-298) in the V3 region of gp120 governs HIV-1 binding to syndecans. We found that an amine group on the side chain of this residue is necessary for syndecan utilization by HIV-1. Furthermore, we showed that HIV-1 binds syndecans via a 6-O sulfation, demonstrating that this binding is not the result of random interactions between basic residues and negative charges, but the result of specific contacts between gp120 and a well defined sulfation in syndecans. Surprisingly, we found that Arg-298, which mediates HIV-1 binding to syndecans, also mediates HIV-1 binding to CCR5. We postulated that HIV-1 recognizes similar motifs on syndecans and CCR5. Supporting this hypothesis, we obtained several lines of evidence that suggest that the 6-O sulfation recognized by HIV-1 on syndecans mimics the sulfated tyrosines recognized by HIV-1 in the N terminus of CCR5. Our finding that CCR5 and syndecans are exploited by HIV-1 via a single determinant echoes the mechanisms by which chemokines utilize these two disparate receptors and suggests that the gp120/ chemokine mimicry may represent a common strategy in microbial pathogenesis.The dominant cell surface heparan sulfate proteoglycans (HSPGs) 3 are the syndecans. Syndecans are transmembrane receptors highly expressed on adherent cells (i.e. epithelial cells, endothelial cells, and macrophages), but poorly expressed on suspension cells (i.e. T-lymphocytes) (1-4). The syndecan family is composed of four members, syndecan-1 to -4. Their ectodomain bears linear heparan sulfate chains, which are composed of a repetition (30 -400 repeats) of a sulfated disaccharide motif (5). The sulfation pattern of the heparan sulfates dictates the ligand specificity of syndecans. Syndecans via their heparan sulfates function as co-factors in cell-cell adhesion, in linking cells to ligands in the extracellular matrix, and in the binding and activation of cellular growth factors (5). Syndecans also function as receptors for HIV-1. We and others demonstrated that pretreatment of HIV-1 target cells (i.e. CD4ϩ T cell lines, CD4ϩ HeLa cells, and CD4ϩ CHO cells or macrophages) with heparinase, an enzyme that removes heparan sulfates from the ectodomain of syndecans, significantly reduces HIV-1 infectivity (1, 6 -11). Zhang et al. (11), using CHO cells that either express HSPGs (CHO-K1) or lack HSPGs (pgsA745), obtained evidence that HSPGs favor HIV-1 infection in a gp120-dependent manner. Previous work suggests that the requirement for syndecans and HSPGs in HIV-1 infection is particularly accentuated when target cells express low levels of entry receptors (CD4 and CCR5/CXCR4) such as CD4ϩ HeLa cells and macrophages (1). However, why and how HIV-1 uses sulfated syndecans to optimally infect these t...

CD134 is a primary binding receptor for feline immunodeficiency virus (FIV), and with CXCR4 facilitates infection of CD4(+) T cells. Human CD134 fails to support FIV infection. To delineate the regions important for defining virus specificity of CD134, we exchanged domains between human and feline CD134. The binding site for FIV surface glycoprotein (SU) is located in domain 1, in a region distinct from the natural ligand (CD134L)-binding site. Mutagenesis showed that Asp60 and Asp62 are required for interaction with FIV, and modeling studies localized these two residues to the outer edge of domain 1. Substitutions S60D and N62D, in conjunction with H45S, R59G and V64K, imparted both FIV SU binding and receptor function to human CD134. Finally, we demonstrated that soluble CD134 facilitates infection of CD134(-) CXCR4(+) target cells in a manner analogous to CD4 augmentation of HIV infection.