Functional characterization of Vpus from Cameroonian HIV-1 N strains.

<p>(<b>A</b>) FACS analysis of 293T cells cotransfected with tetherin, CD4, CD1d or NTB-A expression vectors and pCGCG plasmids expressing eGFP alone (lanes 2 and 3) or together with the indicated <i>vpu</i> allele (lanes 4 to 9). eGFP expression levels used to calculate receptor downmodulation and the mean fluorescence intensities (MFIs) are indicated. (<b>B</b>) Effect of various Vpus on infectious virus release. 293T cells were cotransfected with HIV-1 ΔVpu NL4-3 (2 µg), pCGCG vectors coexpressing eGFP and Vpu (500 ng), and a construct expressing HU tetherin (125 ng). Viral supernatants were obtained two days later and used to measure the quantity of infectious HIV-1 in the culture supernatants by infecting TZM-bl indicator cells. Shown is the infectious virion yield relative to that obtained in the absence of tetherin (100%). The results were confirmed in two independent experiments and each symbol indicates infectious virus yield in the presence of one of the 25 <i>vpu</i> alleles analyzed. (<b>C–F</b>) Vpu-dependent reduction of (<b>C</b>) tetherin, (<b>D</b>) CD4, (<b>E</b>) CD1d and (<b>F</b>) NTB-A surface expression in 293T cells. Shown is the reduction in the levels of receptor cell surface expression relative to those measured in cells transfected with the eGFP only control vector. Each symbol represents n-fold downmodulation of the indicated receptor molecule by one individual <i>vpu</i> allele examined. Shown are average values derived from three experiments. <i>Vpu</i> alleles derived from HIV-1 M are color coded red, N blue (except for the DJO0131 Vpu highlighted in green) and SIVcpz black; those shown in panel A are indicated by open symbols in panels B to F.