Skip to main content
. 2011 Jul;85(14):7353–7362. doi: 10.1128/JVI.00141-11

Fig. 1.

Fig. 1.

Depletion of the endogenous pool of Spry2 inhibits HIV-1 Gag release. (A) Qualitative analysis of Spry2 mRNA expression. COS-1 cells were treated with siRNA against Spry2 or against a nonspecific target (C) 24 h prior to transfection with DNA encoding Gag-GFP. A total of 48 h post-DNA transfection, total RNA was harvested, and cDNAs were generated. The blot shows Spry2 mRNA expression following treatment of COS-1 cells with control or targeted siRNA. (B) Analysis of Spry2 depletion by qPCR. cDNA levels were quantified by qPCR analysis using primers specific to Spry2. The data were normalized to the housekeeping gene h60S levels. The results represent the average of three independent experiments. (C) Analysis of Spry2 depletion by Western analysis. Cell lysates prepared as described in Materials and Methods were centrifuged to remove particulate material and then analyzed by Western blotting for exogenous Spry2 protein expression using antibody directed to the Spry2 C-terminal region. (D) Effect of Spry2 depletion on VLP release from cells. Gag proteins in cell lysates and VLPs in media from samples treated with control (lanes 1 to 3) or targeted siRNA (lanes 4 to 6) were isolated as described in Materials and Methods and analyzed by SDS-PAGE and Western blotting. (E) Semiquantitative analysis of VLP release. The panel shows the ratio of the Gag signal in VLP isolated from the media to the sum of the Gag signal in VLPs plus the cell lysate.