Extended Data Fig. 9: WDR5 enriches at Clock/Bmal1 motifs and correlates with rhythmic regulation of Clock/Bmal1 target genes.
From: Bidirectional histone monoaminylation dynamics regulate neural rhythmicity
a, Representative IGV browser tracks of H3K4me3Q5his, H3K4me3Q5ser, WDR5, H3K4me2 and H3K4me3 at Clock/Bmal1 targets (e.g., Per1) vs. non-Clock/Bmal1 target loci (e.g., Clock and Arntl), demonstrating selective enrichment/rhythmicity of WDR5 at Clock/Bmal1 target genes. b, Assessment of gene expression (normalized reads) for genes bound by WDR5 vs. those that are not bound by WDR5 at ZT16 (the height of WDR5’s binding to chromatin across the ZT; Supplementary Table 5), indicating that WDR5 bound genes are more highly expressed vs. genes not bound by WDR5 (Wilcox Rank Sum Test, p < 2.2e-16). c, HOMER motif enrichment analysis of WDR5 bound vs. unbound loci, indicating that WDR5 significantly enriches that Clock/Bmal1 motifs (Benjamini-Hochberg, p < 0.05). d, Heatmap of gene expression (related to Fig. 3a and Supplementary Table 2) comparing WDR5 bound rhythmic genes vs. expression of genes not bound by WDR5 across the ZT, indicating that WDR5 bound genes are largely direct targets of Clock/Bmal1 (e.g., Per1/2, Dbp, Nr1d1/2, etc.), whereas circadian genes that are not bound by WDR5 are not direct targets of Clock/Bmal1 (e.g., Arntl, Npas2, etc.). e, Western blotting analyses of H3K4me3 and H3K27ac (normalized to H3/amido black staining) in TMN tissues virally transduced with AAV-GFP vs. H3.3 WT vs. H3.3Q5A. One-way ANOVAs were performed with no significant effects observed (n = 4 biological replicates/viral treatment; H3K4me3 – p = 0.4306, F2,9 = 0.9266; H3K27ac – p = 0.4646, F2,9 = 0.8357). f, Model: our genomics and biochemical data indicate that: (1) during periods of activity in TMN, WDR5 – likely in complex with MLL/SETD1 – is recruited to Clock/BMAL1 (E-Box) target genes (e.g., Per1/2, Dbp, Nr1d1/2, etc.), which are induced in their expression as a result of Clock/Bmal1 binding, and its binding is further stabilized to H3Q5ser; (2) during transitional periods towards inactivity, WDR5/MLL (SETD1) becomes destabilized at Clock/BMAL1 target genes owing, in part, to loss of H3Q5ser and stabilization of H3Q5his, the latter of which is antagonistic to WDR5 binding and H3K4 HMT activities; and (3) during periods of inactivity, H3Q5his is further reduced in its enrichment at Clock/Bmal1 targets, thereby allowing for spreading of H3K4 methylation, and the eventual re-recruitment of WDR5/MLL (SETD1) during transitions back into phases of activity. Supplementary Fig. 1 = uncropped blots. Source data provided as a Source Data file.
