HIV Env suppresses anti–CD3-activated CD4⁺ T-cell proliferation. (A) 293T cells expressing IIIB, 89.6, YU-2, SF162, and JR-Fl Envs or no Env (poly(A, C) RNA transfected) were cocultured with CFSE-labeled PBMCs with or without sCD4. CD4⁺ T cells were activated with anti-CD3 mAb (0.5 μg/mL), stained with anti-CD4 mAb, and monitored for proliferation 4 days after coculture by flow cytometry. (B) Negatively selected purified CD4⁺ T cells were cocultured with 293T cells expressing IIIB Env or no Env and activated using anti-CD3 mAb (0.5 μg/mL). Fusion assays demonstrated surface expression of functional Env. Encoding mRNA-transfected 293T cells were mixed with QT6 cells expressing CD4 and CXCR4 (C) or CCR5 (D), and fusion of the 2 cell populations was measured by luciferase production. Quadrants are labeled with the percent of events in each. Experiments shown are representative of 3 independent experiments.