Investigating the role of WT1 in Brugada syndrome pathophysiology

Abstract Background Brugada syndrome (BrS) is a heritable arrhythmic disorder presumably caused by conduction slowing in the right ventricular (RV) outflow tract (RVOT). A recent genome-wide association study found an association between common variants on chromosome 11 near WT1, tagged by the SNP rs72905083, and BrS susceptibility. Purpose To study whether WT1 affects cardiac conduction and could therefore represents the underlying association of the chr11 locus with BrS susceptibility. Methods We conducted in silico analyses of omics datasets to identify the likely causal gene at the chr11 BrS locus. Next, we conducted electrophysiological studies in heterozygous young adult (4 months old) Wt1-deficient (Wt1+/-) mice and wild-type littermates to explore the effect of Wt1 on cardiac conduction. To study the potential role of Wt1 in the setting of reduced conduction reserve, we compared aged Scn5a haploinsufficient mice (Scn5a+/-;Wt1+/+, 17 months) with littermates haploinsufficient for both Wt1 and Scn5a (Scn5a+/-;Wt1+/-). Optical mapping was performed in Langendorff-perfused hearts, where conduction was further challenged by acute ajmaline administration or by pacing at the effective refractory period (ERP). Transcriptomic analyses were conducted in Wt1+/- and wild-type mice by RNAseq of PCM1-sorted cardiomyocytes. Results Analysis of omics datasets uncovered WT1 as the likely causal gene since WT1 was the closest gene to, and in fact overlapped, the BrS-associated region; promoter capture Hi-C data in human tissue showed chromatin contact between the promoter of WT1 and the associated region; and H3K27ac ChIP-seq signals (i.e. enhancer activity) of putative cardiac enhancers located within the associated region were correlated with WT1 transcript abundance across multiple tissues (Z-score = 4.32, p=1.57e-05). No differences were observed in ECG parameters or optically mapped epicardial conduction between Wt1+/- mice and wild-type littermates. However, comparison of aged Scn5a+/-;Wt1+/- and Scn5a+/-; Wt1+/+ hearts, in the setting of acute ajmaline challenge or pacing at the ERP, showed faster conduction in RV/RVOT in the presence of Wt1 haploinsufficiency (genotype effect: p=0.057 and p=0.041 for RV and RVOT epicardial conduction, respectively, during ajmaline challenge; p=0.034 for RV epicardial conduction during pacing at ERP). RNAseq of RV/RVOT cardiomyocytes revealed higher Scn5a expression in Wt1+/- vs wild-type (p=0.006). Conclusion We identified WT1 as the gene that most likely underlies the effect of the chr11 BrS susceptibility locus. Functional data revealed protective effects of reduced Wt1 expression in the setting of reduced conduction reserve. Transcriptomic data suggest that this occurs through a transcriptional effect on Scn5a expression.