Fig. 5.

MafG regulates Nrf2 signaling. (A) Reporter gene activity assay. Co-expressing wild type MafG regulated Nrf2 induced ARE-luciferase activities in a bi-directional way. In contrast, co-expressing MafG2p mutant markedly inhibited Nrf2 induced ARE-luciferase activities in a dose dependent way. Hela cells were transfected with 1 µg pHM6-Nrf2, 0.5 µg plasmid expressing ARE-Luc together with 0, 1, 10, 25 and 100 ng plasmids expressing wild type (wt) or 2p mutant (mt) MafG. Twenty four hours after transfection, cells were harvested. Luciferase activity was measured and normalized to protein concentration. Single and double asterisks indicate statistical significance (t-test) of p<0.05 and p<0.01, respectively. (B) RT-PCR analysis of the transcription of phase II genes. 3 µg pcDNA3.1-Nrf2-V5 plasmids were expressed alone or co-expressed with 1 µg pcDNA3.1-Myc-MafG or pcDNA3.1-Myc-MafG2p in HeLa cells. Total RNAs were extracted using RNeasy method and reversed transcribed (RT). Same amount of RT samples were amplified by poly chain reaction (PCR) for 40 cycles and resolved in 1% agarose gel and visualized by ethidium bromide incorporation exited by UV light. The densitometric values of RT-PCR products were labeled underneath. (C) Western blotting results showed that MafG and MafG2p could enhance and attenuate Nrf2-induced HO-1 and NQO1 expression, respectively.