Figure 1.
Recombinant CyPA inhibits HIV-1 infection. (A) PBMCs from 6 different donors were infected with HIV-1RF in the presence of indicated concentrations of recombinant murine CyPA. At the peak of viral infection (typically day 7 to 11 after infection), RT activity was measured in culture supernatants. The results were tabulated as percentage of inhibition relative to untreated HIV-infected cultures from the same donor. The data are mean ± SEM across donors. (B) PBMCs were infected with HIV-1RF in the presence of indicated concentrations of recombinant murine CyPA. Two hours after infection, cells were washed and viral DNA was analyzed by PCR by using primers that amplify the R/U5 region of the LTR. Amplified fragments were revealed by Southern hybridization, and the amount of product was quantified on Packard Direct Imager (bar graph on the right). A fragment of the α tubulin gene was amplified from the same samples to control for possible differences in the amount of total DNA. Dilutions of 8E5 cells containing the indicated number of HIV-1 genomes were amplified in parallel as standards. Results are shown for one representative experiment out of two performed. (C) Primary differentiated human macrophages were infected with HIV-1ADA in the presence of the indicated concentrations of CyPA. Two hours after infection, cells were washed and treated with trypsin. Viral RNA then was analyzed by RT-PCR by using primers specific for the pol gene. Results of one out of two independent experiments are shown on the left. Results of both experiments were quantified on a Packard Direct Imager and plotted on the right as mean ± SE. Dilutions of 8E5 cells were amplified in parallel, as above, to provide a standard curve.