RNAi-mediated RNF20 knockdown increases interferon regulatory factor 1 gene ( IRF1 ) transcription. (a, b) Reverse transcription quantitative PCR (RT Q-PCR) to quantitate IRF1 mRNA and pre-mRNA expression in stably selected 2fTGH cells expressing a non-silencing small hairpin (sh)RNA or RNF20-shRNA that were uninduced or treated with IFNγ. IRF1 expression was normalized to ACTB and presented as fold change relative to the uninduced, non-silencing shRNA condition. Error bars are standard error (n = 4). (c) Western blot of extracts prepared from stably selected 2fTGH cells expressing non-silencing shRNA (NS) or RNF20 shRNA or non-transfected cells (NT) with α-RNF20. Signal transducer and activator of transcription 1 (STAT1) served as loading control. (d) Western blot of extracts prepared from transiently transfected 2fTGH cells expressing pcDNA3.0 or FLAG tagged RNF20 pcDNA3.0 with α-FLAG and α-RNF20. STAT1 served as loading control. (e) RT Q-PCR of transiently transfected 2fTGH cells expressing pcDNA3.0 or FLAG tagged RNF20 pcDNA3.0 as in (a). Error bars are standard error (n = 2). **P = 0.01, *P = 0.05. Blot quantifications were performed using ImageJ.