Figure 3

Regulation of BTG2 mediated–NFκB activation by IκBα degradation. (A) To confirm the effect of BTG2 on the activation of NFκB and its regulation of MnSOD induction, the mixture of five short interfering RNAs to BTG2 (siBTG2#1 - siBTG2#5) was transfected to HeLa cells for 48 h and then expression of MnSOD was analyzed by RT-PCR. The expression was downregulated by the concentration of siBTG2 dependently, indicating the regulation of MnSOD expression by endogenous BTG2. The sense and the antisense primer sequences of siBTG2 were described in the Additional file 6. (B) Immunoblot analysis revealing the regulation of IκBα degradation by BTG2. Knockdown of endogenous BTG2 expression by transfection of HeLa cells with 100 nM siBTG2 abolished BTG2 protein expression, in contrast to the accumulation of IκBα. (C) Transfection of siBTG2 blocked the effect of exogenous BTG2^/TIS21 on IκBα degradation, indicating the same effect of the exogenous BTG2^/TIS21 and the endogenous BTG2 genes on the regulation of NFκB activation. (D) To further evaluate the effect of BTG2^/TIS21 gene on the IκBα degradation and the upregulation of MnSOD expression, HeLa cells were infected with lentivirus containing IκBα mutant (Ser^{32, 36}Ala) prepared in our laboratory. Overexpression of the mutant reduced upregulation of MnSOD transcription stimulated by BTG2 gene. (E) Complete failure of the BTG2-mediated degradation of IκBα mutant (Ser^{32, 36}Ala), examined by immunoblot analysis.