Fig. 2: RFC4 is a de novo direct transcriptional target of Notch1 signaling.

a, b Effect of activating Notch signaling by overexpression of NICD1, JAG1, or inhibiting Notch signaling by treatment of a γ-secretase inhibitor DAPT on both protein and mRNA levels of RFC4 in A549, H1975, and LC1 cells. Representative images of three independent reproducible experiments are shown. c, d ChIP analysis following H3K27ac immunoprecipitation shows the interaction between H3K27ac and the promoter region of the RFC4 gene in response to overexpression of NICD1 or treatment of DAPT. IgG immunoprecipitation was used as a negative control. e The effect of silencing RBP-Jκ or overexpressing RBP-Jκ together with treatment of DAPT on protein levels of RFC4 in A549, H1975, and LC1 cells. Representative images of three independent reproducible experiments are shown. f The effect of silencing RBP-Jκ and overexpressing RBP-Jκ together with treatment of DAPT on mRNA levels of RFC4 in A549, H1975, and LC1 cells. g Schematic diagram of potential binding site for RBP-Jκ in the promoter region of RFC4. h ChIP enrichment assay shows binding of RBP-Jκ to the predicted binding site in the promoter region of RFC4 in stimulation of JAG1. IgG immunoprecipitation was used as a negative control. i The effects of overexpressing NICD1 together with RBP-Jκ depletion on luciferase activities of the reporter constructs spanning wild type or mutant predicted putative binding site for RBP-Jκ in the promoter region of RFC4. Data in panels b–d, f, h and i are presented as mean ± SD derived from three independent experiments. Two-way ANOVA multiple comparison analysis was used for statistical analysis. Source data are provided as a Source Data file.