Fig. 4: SETD5 is recruited to primed enhancers and diminishes prior to enhancer activation. | Nature Communications

Fig. 4: SETD5 is recruited to primed enhancers and diminishes prior to enhancer activation.

From: Spatiotemporal dynamics of SETD5-containing NCoR–HDAC3 complex determines enhancer activation for adipogenesis

Fig. 4

a Genome-wide distribution of SETD5-V5 before (0 h) and 6 h of differentiation in 3T3-L1 preadipocytes. TTS, transcription termination site. b Venn diagram (left) and heatmap (middle) representations of genomic regions occupied by SETD5-V5, H3K4me1, and H3K27ac. Heatmap analysis showing ChIP-seq data for SETD5-V5, H3K4me1, and H3K27ac at a 20-kb region centered on SETD5-V5 binding regions. Pie chart (right) shows SETD5-V5 binding on H3K27 acetylated regions repressed by SETD5. c Genome browser representation for SETD5-V5, H3K4me1, H3K27ac, C/EBPβ, and C/EBPδ on Cebpa and Pparg genes in 3T3-L1 preadipocytes. Tracks 3–11 are the same data as Supplementary Fig. 1d or Fig. 2e. d ChIP-qPCR analysis of SETD5-V5 on Cebpa and Pparg genes during the early adipogenesis. 3T3-L1 preadipocytes transduced with SETD5-V5 were subjected to ChIP-qPCR analysis using anti-V5 antibody. Data are mean ± SEM (0, 6, 12, 48 h n = 4; 24 h n = 3; independent experiments). e Motif analysis of SETD5-V5 binding sites at 6 h of differentiation. Motif analysis was performed by Homer bioinformatics resources3. p-values were calculated as one-tailed. f Co-immunoprecipitation assay showing the interaction of SETD5-V5 with C/EBPβ. 3T3-L1 preadipocytes transduced with SETD5-V5 at the indicated time of differentiation were cross-linked with ethylene glycerol bis(succinimidyl succinate) and subjected to immunoprecipitation using anti-C/EBPβ antibody. Immunoprecipitates were decrosslinked and examined by immunoblotting using anti-V5 and anti-C/EBPβ antibodies. Representative of two independent experiments. g ChIP-qPCR analysis of SETD5-V5 on Cebpa and Pparg genes under knockdown of C/EBPβ and C/EBPδ. 3T3-L1 preadipocytes transduced with SETD5-V5 were transfected with control siRNA or siRNA targeting to Cebpb and Cebpd and subjected to ChIP-qPCR analysis using anti-V5 antibody. Data are mean ± SEM of four independent experiments. d, g One-way ANOVA with Tukey’s multiple comparisons test. *p < 0.05; **p < 0.01. Source data are provided as a Source data file.

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