FIG. 4.

A VP22 mutant with both the N- and C-terminal sites altered is phosphorylation negative. (A) Extracts were made from COS-1 cells transfected with plasmids expressing either WT or pGE168 protein. These extracts were analyzed by Western blotting using antibody AGV30 (W. blot) and by immunoprecipitation with AGV30 followed by in vitro phosphorylation in the absence (− CKII) or presence (+ CKII) of CKII. (B) COS-1 cells transfected with plasmids expressing either WT (Wt) or pGE168 (168) proteins or mock transfected (M) were labelled in vivo with [³²P]orthophosphate. Extracts were made and analyzed by Western blotting with AGV30 (W. blot), and immunoprecipitation with AGV30 was followed by SDS-PAGE and autoradiography. The positions of radiolabelled protein which coprecipitates with both WT and pGE168 proteins (●) and heavily radiolabelled material at the top of the gel which coprecipitates with both WT and pGE168 proteins (◂) are indicated.