FIG 3.

CHD1 silencing reduces influenza virus RNP activity. (A) HEK293T cells were infected with lentiviruses expressing CHD1 silencers or a control lentivirus (shTT) and were used for CAT RNP reconstitution (see Materials and Methods). At 48 h postreconstitution, the amount of CAT protein in total-cell extracts was analyzed by ELISA. CAT activity was normalized in each case to the amount of GFP expression, which was used as a transfection control. Activity in control lentivirus-transfected cells was considered 100%. Mock, untransfected cells; negative control, the plasmid expressing PB1 was omitted. (B) Aliquots of samples for which results are shown in panel A were analyzed by Western blotting for RNP, GFP, and β-actin. (C) Aliquots of samples for which results are shown in panel A were subjected to primer extension analysis (see Materials and Methods). Asterisks indicate nonspecific bands in uninfected cells (−). (D) Quantification of data from panel C. Bars represent means ± standard errors of the means. Significance was determined using an unpaired Student t test with Welch's correction; 3 independent experiments were performed. **, P < 0.01; ***, P < 0.001.