Fig. 7.

PMLIV depletion boosts EMCV production in IFN-α-treated cells. (A) Efficacy of siRNA PMLc, siRNA PMLIV, and siRNA PMLIII. U373MG cells stably expressing PMLIII or PMLIV were transfected with scramble siRNA, PMLc siRNA (common to all PML isoforms), PMLIV-specific siRNA, or PMLIII-specific siRNA (top blots). U373MG cells, transfected with scramble siRNA or PMLc siRNA, were treated 1 day posttransfection with 500 units/ml of IFN-α for 24 h (bottom blots). Western blot analysis was performed using anti-PML and antiactin antibodies. (B) Increase of PMLIII and PMLIV expression in IFN-treated U373MG cells. U373MG cells were left untreated or treated with 100 units of IFN-α. RNA was extracted in all samples, and PMLIII and PMLIV expression was tested by the real-time PCR method. Results were normalized to GAPDH. The averages plus standard deviations (error bars) for three independent assays are shown. The statistical significance (P values) was calculated using a Student's t test. (C) Knockdown of PMLIV by PMLIV-specific siRNA impairs IFN-α-induced antiviral activity. U373MG cells were transiently transfected for 12 h with siRNA scramble, siRNA PMLc, siRNA PMLIV, or siRNA PMLIII, treated with IFN-α for 24 h, and then infected with EMCV at an MOI of 1 for 14 h. Viral titers were determined as described in Materials and Methods.