Figure 4: Tβ4 interacts with SWI/SNF enhancing activation of the adult epicardium. | Nature Communications

Figure 4: Tβ4 interacts with SWI/SNF enhancing activation of the adult epicardium.

From: BRG1-SWI/SNF-dependent regulation of the Wt1 transcriptional landscape mediates epicardial activity during heart development and disease

Figure 4

(a,b) Immunoprecipitation experiments with Tβ4-EGFP-transfected cells using anti-BRG1 (a) or anti-Tβ4 (b) antibodies. The original uncropped images of gels are shown in Supplementary Fig. 11. (ce) Duolink in situ analysis using anti-rabbit PLUS and anti-mouse MINUS PLA probes specific for Tβ4 and BRG1 antibodies respectively, in embryonic (c), intact adult (d) and post-MI hearts (e). (fh) Duolink in situ analysis using anti-rabbit PLUS and anti-mouse MINUS PLA probes against BRG1 and BAF155. (ik) re-ChIP with anti-Tβ4, anti-BAF180 or anti-BRG1 antibodies using chromatin pulled-down with anti-BRG1 (i,j) or anti-BRM (k) to examine Tβ4-SWI/SNF complex formation in E11.5 (i) and adult post-MI, primed hearts (j,k). (l) ChIP-qPCR data from chromatin derived from post-MI, Tβ4 knockout (KO) adult hearts showing minimal binding of BRG1 in Wt1 ECRs. (m) Wt1 and Raldh2 expression analysis by qRT-PCR compared to sham-operated animals. (n) ChIP-qPCR data from chromatin generated from primed- (+Tβ4), and injured- (MI) adult Tβ4 knockout hearts revealing BRG1 strong enrichment in Wt1 ECRs. (o) Wt1 and Raldh2 expression analysis by qRT-PCR compared to sham-operated animals. At least three independent ChIP experiments per antibody per sample group (n=10 fetal hearts; n=3 adult hearts) were performed. ChIP results are presented as fold enrichment over input, whereas re-ChIP results are present in fold enrichment over the level of ChIP with negative control IgG antibody. (p) List of representative BRG1-enriched epicardial genes shared between PBS- and Tβ4-primed, -injured (MI) adult hearts and representative UCSC browser snapshots of the Aldh1a2 and Tbx18 loci (highlighted in blue in the list of genes) derived from the Brg1 ChIP-seq experiments. All error bars are data±s.d. Significant differences (P values) were calculated using two-tailed Student’s t-test (*P≤0.05; **P≤0.01). ep, epicardium; my, myocardium; ses, subepicardial space. All scale bars 50 μm.

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