Fig. 5.

Increasing chromatin accessibility via drug treatment lowers the RelA induction threshold. (A) Heterochromatin fraction for clone 15.4 was quantified with a DNAse I sensitivity assay following stimulation with 400 nM TSA for 4 hours or with 5 μM 5-aza- dC for 48 hours. Quantitative PCR was performed in triplicate and normalized to the hemoglobin reference gene. Relative heterochromatin fraction was calculated by normalizing clone 15.4 with drugs, B5 and D3 to the unstimulated 15.4 control. (B) Combined flow cytometry data for 15.4 expressing iRelA in response to a range of DOX concentrations and simultaneous stimulation with 400 nM TSA for 24 hours (dark blue), 5 μM 5-aza-dC for 48 hours (red), and no drug treatment controls at 24 and 48 hours (black and light gray, respectively). iRelA dose response curves for clone B5 (green) and D3 (dark gray) without TSA or 5-aza-dC are included for comparison. (C) Relative change in induction threshold versus relative change in heterochromatin fraction for clones 15.4 (circles), 8.4 (diamonds) and E3 (triangles). Data for 15.4 are calculated from results presented in (A) and (B), and data for 8.4 and E3 are calculated from experiments presented in Fig. S10. All points are calculated by normalizing the value of heterochromatin fraction or threshold for the clone in the presence of drugs to the corresponding value for the unstimulated control clone. Data are presented as the mean ± standard deviation. Changes are labeled as significant (*) if p < 0.05. Pearson correlation coefficient R is indicated on plot.