FIG 3.

HIV-1 group M Nefs antagonize the long isoform, but not the short isoform, of human tetherin. (A and B) Jurkat-TAg cells expressing long versus short isoforms of human tetherin (JTAg L- versus S-tetherin cells) were infected with HIV-1.Δvpu.IeG-Nef recombinants expressing the indicated Nef alleles, and virus release was measured by a p24 antigen capture ELISA (A) and by Western blot analysis of culture supernatants and cell lysates (B). To maximize infection, cells were infected by spinoculation for 2 h with concentrated virus (2 μg/ml p24) in the presence of Polybrene (8 μg/ml). The error bars represent the standard deviations of the means for independent infections, and mean virus release was compared to that in JTAg L-tetherin cells infected with a virus expressing NL4-3 Nef by unpaired t tests (**, P ≤ 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001; ns, not significant). (C) Histograms show tetherin (BST-2) staining on the surface of cells infected with recombinant viruses expressing the indicated Nef alleles relative to cells infected with a nef-minus control virus or stained with an isotype control antibody (shaded). Virus-infected cells were identified by gating on viable, p55⁺ CD4^low cells. The data in panels A to C are representative of results from three independent experiments.