CD4 Interacts Constitutively with Multiple CCR5 at the Plasma Membrane of Living Cells

Aurélie-Marie Baker,Bernard Lagane,Gérald Gaibelet,Laurence Salomé,Françoise Bachelerie,Fabrice Dumas,Aude Saulière,Marie Fourage,Serge Mazères,André Lopez

doi:10.1074/jbc.m705617200

Abstract

The entry of human immunodeficiency virus into target cells requires successive interactions of the viral envelope glycoprotein gp120 with CD4 and the chemokine receptors CCR5 or CXCR4. We previously demonstrated, by Förster resonance energy transfer experiments, the constitutive association of CD4 and CCR5 at the surface of living cells. We therefore speculated that this interaction may correlate with compartmentalization of CD4 and CCR5 within the plasma membrane. Here, we characterize the lateral distribution, the dynamics, and the stoichiometry of these receptors in living cells stably expressing CD4 and/or CCR5 by means of fluorescence recovery after photobleaching at variable radii experiments. We found that (i) these receptors expressed alone are confined into 1-microm-sized domains, (ii) CD4-CCR5 associations occur outside and inside smaller domains, and (iii) these interactions involve multiple CCR5 molecules per CD4.

Highlights

The entry of human immunodeficiency virus into target cells requires successive interactions of the viral envelope glycoprotein gp120 with CD4 and the chemokine receptors CCR5 or CXCR4
The fusion induced by human immunodeficiency virus type 1 (HIV-1)6 is a multistep process that requires for the virus to bind to two different host cell surface receptors
Our recent study carried out using a steady state Forster resonance energy transfer (FRET) approach, at the surface of living cells, demonstrated the constitutive interaction of CD4 and CCR5, reinforced by gp120

Summary

A FLUORESCENCE RECOVERY AFTER PHOTOBLEACHING AT VARIABLE RADII APPROACH*

Current models suggest an initial interaction between the trimeric viral envelope protein gp120 and the primary CD4 receptor on target cells This results in a conformational change of gp120 that exposes a co-receptor binding site. Our recent study carried out using a steady state Forster resonance energy transfer (FRET) approach, at the surface of living cells, demonstrated the constitutive interaction of CD4 and CCR5, reinforced by gp120. This finding indicated that a ternary complex between gp120, CD4, and CCR5 would form before the fusion process occurred [10]. For the first time, we highlight the dynamics of this prefusion complex within the membrane of living cells

EXPERIMENTAL PROCEDURES

Findings

RESULTS

DISCUSSION