Fig. 2.

RTA is induced by the silencing of IRF4 and regulates ISRE mediated gene expression in PEL cells. (A) Induction of RTA protein following IRF4 knockdown. Whole cell lysates fromBCBL-1 sh-IRF4 cells after 72 h treatment with various concentrations of Dox were immunoblotted with antibodies against IRF4, RTA and Actin. (B) Nuclear accumulation of RTA expression follows loss of IRF4. Nuclear fractions were prepared from cell treated with Dox for 72 h. Fractions were resolved by SDS-PAGE and immunoblotted with antibodies against IRF4, RTA, DRBP76 (nuclear marker), and Tubulin (cytoplasmic marker). (C) Effect of RTA expression on IFNβ-promoter activity. HEK293 cells were transfected with RTA or vector control along with IFNβ₁₂₅-luc and pRL-Null. Luciferase activity was measured as previously described (Forero et al., 2013). Samples were normalized to Renilla luciferase control and expressed as fold change with respect to empty vector transfected cells (value 1). (D) Quantitative RT-PCR analysis of ISG60 following ectopic expression of RTA in 293T cells. Samples were normalized to RPL32 and expressed as fold change with respect to empty vector transfected cells (value 1). (E) Analysis of ISG60 protein induction following ectopic expression of RTA in 293T cells. Samples were immunbobloted with indicated antibodies.