Gp120 undergoes disulfide-reduction prior to, or concomitant with, binding to CXCR4. All cells except for those in lane 7 were pretreated with soluble gp120 III_B followed by labeling with biotin maleimide (MPB) to label free -SH groups. Lane 1, HOS-CD4^–/CXCR4^– cells (devoid of CD4 and CXCR4); lane 2, HOS-CD4^–/CXCR4^– cells pretreated with 5 mM DTNB to block PDI prior to MPB labeling; lane 3, HOS-CD4⁺/CXCR4^– cells (devoid of only CXCR4); lane 4, HOS-CD4^+/CXCR4–cells pretreated with 5 mM DTNB prior to MPB labeling; lane 5, HOS-CD4⁺/CXCR4⁺ cells (expressing both CD4 and CXCR4); lane 6, HOS-CD4⁺/CXCR4⁺ cells pretreated with 5 mM DTNB prior to MPB labeling; lane 7, HOS-CD4⁺/CXCR4⁺ cells that were not pretreated with gp120 (A). Residual supernatants after immunoprecipitation were analyzed for actin content with an anti-actin PAb (B). Data shown are representative of 2 experiments.