Figure 6

Differentiation impairs Bach1 displacement from the promoter region of HO-1 in response to H₂O₂. Undifferentiated and 4d-ATRA differentiated cells have been treated for 3 h with 500 μM H₂O₂. t-BHQ (50 μM) has been used as positive control able to displace Bach1, allowing Nrf2 to bind. (a) ChIP analysis of Bach1 binding to ARE sequences in the E1 promoter of HO-1. Statistical analysis: n = 3, *p < 0.01vs control; ^#p < 0.05 vs control; ^§p < 0.05 vs H₂O₂ undiff. (b) ChIP analysis of Nrf2 binding to ARE sequences in the E1 promoter of HO-1. Statistical analysis: n = 3, *p < 0.05 and **p < 0.01 vs control. (c) Negative control using IgG. As indicated no bands are detected in samples immunoprecipited with normal rabbit/goat IgG in comparison to samples immunoprecipitated by using rabbit Anti Nrf2 or goat Anti Bach1 antibodies. The intensity of PCR products amplified from immunoprecipitated samples are normalized on the intensity of bands obtained from the amplification of pre-cleared DNA (input). The bands show one representative experiment of three. Full-length gels are presented in supplementary information.