FIG. 2.

Assembly and envelopment of ASF virus require an intact ER Ca²⁺ store. Sixteen hours after infection with the BA71v strain of ASF virus, Vero cells were pulse-labeled with [³⁵S]methionine and cysteine for 15 min at 37°C and chased for 2 h in complete medium, in the presence of A23187 (1 μM), EDTA (3 mM), or thapsigargin (500 nM) as indicated. (A) The degree of envelopment of p73 was determined by the trypsin protection assay followed by immunoprecipitation and autoradiography. The bar graphs show relative protein levels calculated by densitometry. (B) The levels of membrane-bound p73 assembled into oligomers after incubation of cells alone or with A23187 (1 μM) and EDTA (3 mM) were determined by sucrose density sedimentation. The levels of p73 present were estimated by immunoprecipitation followed by SDS-PAGE under reducing conditions. The percentage distribution of p73 across the gradients was estimated by densitometry of autoradiographs. For the trypsin protection assays in the presence or absence of A23187, the averages of at least four estimations are shown with standard errors. The other experiments were performed twice, and the averages are shown.