Constitutive cell surface association between CD4 and CCR5

Abstract

HIV-1 entry into cells involves formation of a complex between gp120 of the viral envelope glycoprotein (Env), a receptor (CD4), and a coreceptor. For most strains of HIV, this coreceptor is CCR5. Here, we provide evidence that CD4 is specifically associated with CCR5 in the absence of gp120 or any other receptor-specific ligand. The amount of CD4 coimmunoprecipitated with CCR5 was significantly higher than that with the other major HIV coreceptor, CXCR4, and in contrast to CXCR4 the CD4-CCR5 coimmunoprecipitation was not significantly increased by gp120. The CD4-CCR5 interaction probably takes place via the second extracellular loop of CCR5 and the first two domains of CD4. It can be inhibited by CCR5-and CD4-specific antibodies that interfere with HIV-1 infection, indicating a possible role in virus entry. These findings suggest a possible pathway of HIV-1 evolution and development of immunopathogenicity, a potential new target for antiretroviral drugs and a tool for development of vaccines based on Env-CD4-CCR5 complexes. The constitutive association of a seventransmembrane-domain G protein-coupled receptor with another receptor also indicates new possibilities for cross-talk between cell surface receptors.It has been known for many years that, in addition to the primary receptor CD4, HIV-1 requires cofactor molecules to enter cells (reviewed in ref. 1). It was hypothesized (2, 3) that the entry cofactors may directly associate with the complex between CD4 and the HIV-1 envelope glycoprotein (Env; gp120-gp41) and therefore serve as coreceptors. The identification of the entry cofactors as chemokine receptors (4-9) not only solved a longstanding puzzle about HIV tropism and pathogenesis but also provided new tools for understanding the mechanism of HIV entry. It was later demonstrated that gp120, CD4, and the HIV-1 coreceptor CXCR4 can be coimmunoprecipitated, suggesting that the complex between these three molecules plays a critical role in the initial stages of the entry process (10). By using a displacement assay, it was shown that in the presence of CD4, gp120 associates with the other major HIV-1 coreceptor, CCR5 (11,12). It was also found that gp120 induces CD4-CXCR4 membrane colocalization (13,14), suggesting the formation of higher order molecular complexes.Several previous observations hinted that CD4 could interact with coreceptor molecules even in the absence of gp120, but convincing evidence for the existence of such an interaction between cell surface-associated molecules was lacking (10,11,13,15). Here, we demonstrate that cell surface CD4 associates with CCR5 in the absence of gp120 or other chemokine-receptor-or CD4-specific ligands, we partially characterize regions of the two molecules that are involved in this interaction, and we show a functional correlation between this association and HIV-1 Envmediated fusion. We propose that the CD4-CCR5 interaction plays an important role in the viral entry process and in HIV-1 evolution and immunopathogenesis and could be a new target for antiv...

CD4 and CCR5 Constitutively Interact at the Plasma Membrane of Living Cells

“…Such interactions are likely to favor the association rate of gp120 with CCR5, recently proposed as being strictly dependent upon a close vicinity of CD4 and CCR5 in living cells (20). Furthermore, the observation that CD4 poorly co-immunoprecipitated with CXCR4 (19,21,22) supports the attractive hypothesis that preferential interactions between CCR5 and CD4 may be relevant to the predominance of R5-tropic HIV isolates in the course of viral infection. Nevertheless, this possibility remains debated, as it was recently inferred using coprecipitation, high-resolution deconvolution microscopy and resonance energy transfer techniques, that CD4 and CCR5 do not exist in a stable complex at the cell membrane (23,24).…”

Chemokines: attractive mediators of the immune response

“…Apparently, receptors involved in sensing chemotactic gradients are preferentially localized to the leading edge in a polarized cell. CCR5 also colocalizes with CD4 at the cell surface [9]. Viewed altogether, these data suggest that the localization of CCRs to the leading edge may allow for interaction with TCRs, thereby modulating TCR mediated signaling cascades.…”

“…GPCR-mediated transactivation of the epidermal growth factor [107] and the platelet-derived growth factor receptors [108] have been reported. Given that CCL5-CCR5 activation of Jurkat T cells correlates with the expression of CD3 [19], that CCR5 expression is associated with constitutive CD4 [9] expression and CCR5 localizes to lipid rafts upon ligand binding [4,6], activated CCR5 may function to recruit signaling intermediates of the TCR to potentiate (co)stimulation of T cells. CCL5 at micromolar doses can induce two distinct calcium fluxes in T cells: one dependent on G-proteins and the other on tyrosine kinases [15].…”

Cooperative subunit interactions within the oligomeric envelope glycoprotein of HIV-1: Functional complementation of specific defects in gp120 and gp41

“…Numerous experimental variables may contribute to complementation efficiency, including the surface densities of each of the participating proteins, the binding affinities between gp120 and each receptor, the efficiencies of subunit interactions, etc. The findings of constitutive interactions between CD4 and coreceptors may have significance in this regard (14,31), as may the recent proposal that cooperative interactions involving multiple CCR5 molecules are required for the HIV-1 infection pathway (32). Also of note is that certain complementation effects were not uniformly noted; for example, LAV-BS complemented SF162 wild type for fusion with CXCR4-expressing targets cells (Fig.…”