Fig. 5

Interferon regulatory factor (IRF)-1 transactivates SMN and SMNc gene expression. A172 cells were transfected with 16 µg of the vector alone pCMV or of the expression plasmid pCMV/IRF-1. Total cellular RNA and protein extracts were prepared at 24 hr and 48 hr post-transfection and analyzed for SMN and SMNc expression. (A) by reverse transcription-polymerase chain reaction (RT-PCR) analysis and (B) by Western blots analysis. In (A), lane (0) represents the negative control that was the product of PCR, where RNA was omitted. (C) A172 cells were transfected with the SMN-chloramphenicol transferase (CAT) plasmid, alone or with pCMV/IRF-1 as described in “Materials and Methods.” When indicated, cells were treated 24 hr after transfection with interferon (IFN)-β (1000 U/ml) or IFN-γ (1000 U/ml). CAT activity was determined after 48 hr. The SMN promoter response (fold induction) is the ratio of CAT activity in stimulated or co-transfected cells to that in unstimulated cells, which is defined as 1.0. Values are means of four independent experiments. Identical results were obtained with the SMNc-CAT plasmid.