Figure 6

Subcellular localization of wild-type M-PMV Gag, ΔKKPKR Gag, and RSV Gag-GFP under steady state growth conditions or after treatment with LMB. HeLa cells were transfected with either pSARM-4, ΔKKPKR, or RSV Gag-GFP and left untreated or treated with LMB. The cells were fixed in methanol and the subcellular localizations of Gag were viewed by confocal microscopy using rabbit anti-Pr78 antibodies and Cy2 conjugated secondary antibodies. RSV Gag-GFP was directly visualized by fluorescence of the Gag-GFP fusion protein. Drug treatments: Wild-type M-PMV (untreated, panel 6A), wild-type MPMV (LMB treated, panel 6B) ΔKKPKR (untreated, panel 6C, ΔKKPKR (LMB treated, panel 6D), RSV Gag-GFP (untreated, panel 6E), and RSV Gag-GFP (LMB treated, panel 6F). Colocalization of wild-type M-PMV Gag, ΔKKPKR, and nuclear pores. Transfected HeLa cells were fixed with 4% paraformaldyhyde, and permiablized with 0.2% TX-100. Wild-type Gag (panels G-J), ΔKKPKR Gag (panels k-N), and nuclear pore localization were visualized by confocal microscopy using affinity purified anti-Pr78 and MAb414 antibodies, respectively, and counter stained with Cy2 anti-rabbit and Cy5 anti-mouse antibodies. 0.3 um Z-sections were stacked and orthogonal views through the cell were generated using Flowview imaging analysis software.