Fig. 3.

Cap snatching is essential for viral expression. (A) Decapping activity of Gag to form m⁷Gp-Gag adduct. L-A virions (lane 1) or M1 virions with WT (lane 2) or mutant (Arg154, lane 3) Gag protein were incubated for decapping with 5-nt cap donor labeled at the γ-phosphate with ³²P. Then proteins were separated in an SDS gel and detected by Coomassie blue staining (Left) or by autoradiography (Right). M; protein standards for mobility. (B) Detection of capped viral transcripts in vivo. RNA was extracted from cells containing M1 supported by L-A plasmid expressing WT or mutant Gag (Arg154) or from L-A plasmid-cured cells (−) as control. Capped RNA was immunoprecipitated with (Anti-cap) or without (−) anti-cap antibody and hybridized with M1 (+) strand-specific (Upper), L-A (+) strand-specific (Middle), or 25S rRNA-specific (Lower) probe. As control, total RNA was processed in parallel (Total RNA). (C) Killer assays.