FIG. 3.
Kinetics of translation and processing of p1a-22. (A) Pulse-label translation. Proteins were radiolabeled with [35S]methionine at 7 h postinfection, and samples were withdrawn from 10 to 120 min and used for immunoprecipitation by B4 antiserum, followed by electrophoresis and fluorography on SDS–5 to 18% gradient polyacrylamide gels. Lysates of mock-infected cells were immunoprecipitated by B4 antiserum (lane mock), and lysates of infected cells were immunoprecipitated with B4 preimmune serum (lane Inf-pre). Molecular mass standards (in kilodaltons) are to the left of the gel, and the location of p1a-22 is shown to the right of the gel. (B) Translation with different durations of labeling and with constant chase. Proteins were labeled in MHV-infected DBT cells for periods from 10 to 120 min and then chased with media containing excess unlabeled methionine and cycloheximide for an additional 90 min. The locations of markers and p1a-22 are shown. (C) Pulse-chase translation. Proteins were radiolabeled for 45 min and then chased in media lacking [35S]Met but containing a 10-fold excess of unlabeled methionine for 10 min (10") to 16 h (16′). All samples were immunoprecipitated and analyzed as described above for panel A. All labeling is as described for panel A. The location of the 200-kDa protein is indicated by the small arrow, and open arrowheads indicate the locations of proteins seen only after prolonged chase.