Effect of activation of FPR on monocytotropic HIV-1–envelope fusion and viral infection.

(A) HOS/CD4/CCR5 cells stably contransfected with FPR cDNA, treated with MIP-1α (1 μg/mL, 1 minute, 37°C) or fMLF (10⁻⁶mol/L, 60 minutes, 37°C) were examined for CCR5 phosphorylation with the use of antiphosphoserine antibody for immunoprecipitation followed by immunoblotting with anti-CCR5 antibody. (B) HeLa cells infected with recombinant vaccinia encoding the envelope protein from HIV-1BAL 31 were fused with HOS/CD4/CCR5 cells with (FPR/CCR5/CD4) or without FPR (mock/CCR5/CD4). Chemokines (10 μg/mL) or fMLF were added when env-expressing HeLa cells and target cells were mixed. (C) Human monocytes cultivated for 48 hours in the absence of fetal bovine serum were also fused with HeLa cells expressing the HIV-1BAL 31 env in the presence or absence of fMLF. Results are the mean (± SD) from 2 separate experiments. *Denotes significantly reduced β-galactosidase. (D) Expression of HIV-1 p24 protein following HIV-1 infection of macrophages. rhM-CSF–induced peripheral blood macrophages were exposed to fMLF for 1 hour followed by infection with HIV_JRFL. Four hours later, the cells were washed and placed in culture for 6 days; the supernatants were then collected for analysis of p24 by ELISA. * Significantly reduced p24 production. Similar results were obtained with HOS cells transfected with CD4, CCR5, as well as the fMLF receptor FPR (data not shown).