(T1118) Validating functional cardiomyocyte-specific histone-acetylated variant enhancers in human ES-derived Cardiomyocytes using a STARR-seq approach
Research Fellow National University of Singapore Singapore, Singapore
Abstract: Genome-wide association studies have identified a large number of SNPs associated with cardiovascular disease, which tend to be located in the Cis-regulatory elements of the non-coding genome such as enhancers, that can modulate the transcriptional activity of its target genes. However, the functional activity of these underlying SNPs in specific cardiac cell types can differ, and biological mechanisms of these genetic variations remain largely unexplored.
Previously we have mapped out 47,321 putative human cardiac enhancers and promoters from 70 left ventricular samples and have identified 1680 differential histone acetylome-wide association with its underlying putative SNP based on our histone acetylation quantitative trait locus (haQTL) analysis. Using self-transcribing active regulatory region sequencing (STARR-seq), we have identified 1105/1680 significantly functional cardiac haQTL of varying effect sizes in human H9-hESC derived cardiomyocytes. Additionally, 40/62 unique loci identified by colocalization of haQTLs with subthreshold loci of heart-related GWAS datasets were shown to have regulatory activity in cardiomyocytes. Interestingly, amongst the top STARR-seq hits, the variants rs5758468 and rs11663468 were also identified and validated previously to show disruption to MEF2A binding and the corresponding loss of H3K27ac peak height in cardiomyocytes. Using luciferase assay in H9 ES derived-cardiomyocytes, we have also validated the concomitant reduction in luciferase activity, in the same direction as the STARR-seq assay, but not the non-significant SNPs. Additionally, through overlapping of Hi-C, and chromatin accessibility data, the majority of SNPs within cardiac enhancers validated by STARR-seq are connected to a distal gene by activity-by-contact (ABC) model. Concordant with Posterior inclusion probability (PIP) analysis, our results have also prioritized for functional cardiomyocyte-specific SNPs, that were previously enriched in GWAS studies implicated in cardiomyocyte electrophysiology, such as Atrial Fibrillation.
Taken together, our results have adopted an unbiased approach to prioritize for functional cardiomyocyte regulatory elements, which help delineate target genes at cardiovascular GWAS loci implicated in cardiac disease and function.
Funding Source: C.J.M.L supported by grants from the Singapore National Medical Research Council (NMRC) under OF-YIRG24Jul-0014, and intramural fund under Cardiovascular Metabolic Disease Translational Research programme
