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. Author manuscript; available in PMC: 2011 Sep 15.
Published in final edited form as: Free Radic Biol Med. 2010 Jun 8;49(6):976–983. doi: 10.1016/j.freeradbiomed.2010.05.030

Figure 4. Sag disruption causes IκBα accumulation and blocks NF-κB activation.

Figure 4

Subconfluent ES cells with genotype of Sag+/+ (AB2) and Sag−/− (AB1) was subjected to Western blotting analysis (A) or exposed to 10 Gy radiation, followed by Western blotting analysis at indicated time points (B). A NFκB luciferase reporter construct (pNifty plasmid, Invivogene) [55] was transiently transfected, along with Renilla construct, into AB1 or AB2 ES cells via electroporation. Cells were seeded into 96-well plates and cultured for 24 hr, and left untreated or treated with TNFα (10 ng/ml) (C), or exposed to 10 Gy radiation (D) for 8 hr and 24 hr, respectively, followed by luciferase assay. The results, from three independent transfections with six replicates per treatment, are presented as relative luciferase units after normalization with transfection efficiency with Renilla. The paired Student t test was used for statistical analysis. (*, p<0.05; **p < 0.01). Subconfluent cells were left untreated or exposed to 10 Gy radiation for 8 or 24 hrs, followed by Western blotting analysis using indicated antibodies (E).