Antibodies to several conformation-dependent epitopes of gp120/gp41 inhibit CCR-5-dependent cell-to-cell fusion mediated by the native envelope glycoprotein of a primary macrophage-tropic HIV-1 isolate

The beta-chemokine receptor CCR-5 is essential for the efficient entry of primary macrophage-tropic HIV-1 isolates into CD4(+) target cells. To study CCR-5-dependent cell-to-cell fusion, we have developed an assay system based on the infection of CD4(+) CCR-5(+) HeLa cells with a Semliki Forest virus recombinant expressing the gp120/gp41 envelope (Env) from a primary clade B HIV-1 isolate (BX08), or from a laboratory T cell line-adapted strain (LAI), In this system, gp120/gp41 of the ''nonsyncytium-inducing,'' primary, macrophage-tropic HIV-1(BX08) isolate, was at least as fusogenic as that of the ''syncytium-inducing'' HIV-1(LAI) strain, BX08 Env-mediated fusion was inhibited by the beta-chemokines RANTES (regulated upon activation, normal T cell expressed and secreted) and macrophage inflammatory proteins 1 beta (MIP-1 beta) and by antibodies to CD4, whereas LAI Env-mediated fusion was insensitive to these beta-chemokines. In contrast soluble CD4 significantly reduced LAI, but not BX08 Env-mediated fusion, suggesting that the primary isolate Env glycoprotein has a reduced affinity for CD4. The domains in gp120/gp41 involved in the interaction with the CD4 and CCR-5 molecules were probed using monoclonal antibodies, For the antibodies tested here, the greatest inhibition of fusion was observed with those directed to conformation-dependent, rather than linear epitopes. Efficient inhibition of fusion was nut restricted to epitopes in any one domain of gp120/gp41. The assay was sufficiently sensitive to distinguish between antibody-and beta-chemokine-mediated fusion inhibition using serum samples from patient BX08, suggesting that the system may be useful for screening human sera for the presence of biologically significant antibodies.