Roles of CD4 and coreceptors in binding, endocytosis, and proteolysis of gp120 envelope glycoproteins derived from human immunodeficiency virus type 1

Infections by human immunodeficiency virus type 1 (HIV-1) involve interacti ons of the viral envelope glycoprotein gp120 with CD4 and then with a corec eptor, R5 isolates of HIV-1 use CCR5 as a coreceptor, whereas X4 isolates u se CXCR4, It is not known whether coreceptors merely trigger fusion of the viral and cellular membranes or whether they also influence the energetics of virus adsorption, the placement of the membrane fusion reaction, and the metabolism of adsorbed gp120, Surprisingly, the pathway for metabolism of adsorbed gp120 has not been investigated thoroughly in any cells. To addres s these issues, we used purified I-125-gp120s derived from the R5 isolate B at and from the X4 isolate IIIB as ligands for binding onto human cells tha t expressed CD4 alone or CD4 with a coreceptor, The gp120 preparations were active in forming ternary complexes with CD4 and the appropriate corecepto r, Moreover, the cellular quantities of CD4 and coreceptors were sufficient for efficient infections by the corresponding HIV-1 isolates, In these con ditions, the kinetics and affinities of I-125-gp120 adsorptions and their s ubsequent metabolisms were strongly dependent on CD4 but were not significa ntly influenced by CCR5 or CXCR4, After binding to CD4, the I-125-gp120s sl owly became resistant to extraction from the cell monolayers by pH 3.0 buff er, suggesting that they were endocytosed with half-times of 1-2 h, Within 20-30 min of endocytosis, the I-125-gp120s were proteolytically degraded to small products that were shed into the media. The weak base chloroquine st rongly inhibited I-125-gp120 proteolysis and caused its intracellular accum ulation, suggesting involvement of a low pH organelle, Results supporting t hese methods and conclusions were obtained by confocal immunofluorescence m icroscopy, We conclude that the energetics, kinetics, and pathways of I-125 -gp120 binding, endocytosis, and proteolysis are determined principally by CD4 rather than by coreceptors in cells that contain sufficient coreceptors for efficient infections. Therefore, the role of coreceptors in HIV-1 infe ctions probably does not include steerage or subcellular localization of ad sorbed virus.