FIG 4.

Effects of PA-X on the activity and expression of the RNP complex. (A) Polymerase activity of the RNP in 293T, CEF, and DEF cells. Cells were transfected in triplicate with a luciferase reporter plasmid (p-Luci for 293T cells; paviPolIT-Luc for CEF and DEF) and a Renilla internal control plasmid, together with plasmids expressing CK10 PB2, PB1, and NP and a plasmid expressing either CK10 PA or a CK10 PA-derived mutant. After 24 to 36 h, luciferase activities in cell lysates were measured. The polymerase activity values were normalized to the Renilla luciferase activity. Values shown are means ± SD for three independent experiments and were standardized to those for CK10 PA (taken as 100%). (B) Expression of RNP components from cells cotransfected with plasmids expressing CK10 PB2, PB1, and NP and a plasmid expressing either CK10 PA or a CK10 PA-derived mutant. Levels of viral protein expression were determined by Western blotting. The level of expression of each RNP component relative to the expression of the control, β-actin, was determined by densitometry using BandScan software, version 5.0. Data are representative of the results of three independent experiments and were standardized to those for r-CK10 (taken as 100%).