Fig. 6: Binding of PIF4 on the G-boxes enhances CDF2-binding strength on DNA.

a, Binding strengths for CDF2 peaks varied significantly with the numbers of G-box motifs (CACGTG, Kruskal–Wallis test, P = 4.588 × 10⁻¹⁵). b, Binding strengths for peaks with different numbers of DOF-binding motifs (AAAAG, Kruskal–Wallis test, P = 1.964 × 10⁻⁷). Box plots in panel a and b show the minimum, 25th percentile, median, 75th percentile and maximum of data points. Letters in panels a and b show significant differences among groups (adjusted P < 0.05) using pairwise Wilcox tests. Groups that share at least one identical letter are not significantly different. c, Gel-shift analysis of the interactions between single PIF4^bHLH or CDF2^DOF or combination of PIF4^bHLH and CDF2^DOF WT proteins with WT or mutant DNA probes. d, Gel-shift analysis of the interactions between single PIF4^bHLH or CDF2^DOF or combination of PIF4^bHLH WT or PIF4^{bHLH (Mu1)} mutant proteins and CDF2^DOF WT protein with WT or mutant DNA probes. e, Interactions between single PIF4^bHLH or CDF2^DOF or combinations of PIF4^bHLH WT and CDF2^DOF WT or CDF2^{DOF (Mu1 and Mu8)} mutant proteins with WT or mutant DNA probes analyzed by EMSA. EMSA assays in c, d and e were performed three times with similar results. f, Proposed model for the role of the PIF4–CDF2 module in regulating gene transcription in the light. Upon chromatin opening, the E-box and DOF-binding motifs are accessible. Binding of the G-boxes by PIF4, and the DOF-binding motifs near those G-boxes by CDF2 occurs. Interaction between PIF4 and CDF2 can occur when they are bound to DNA. Binding of PIF4 to the G-boxes strengthens CDF2 binding and allows it to bind to DNA independently of the DOF-binding sites. Therefore, PIF4 recruits CDF2 to the YUCCA8 promoter. After binding of the PIF4–CDF2 module to chromatin, RNA polymerase II is recruited by PIF4 via the Mediator complex to induce gene transcription.