Cis-regulatory elements control gene expression in time and space

Cis-regulatory elements control gene expression in time and space and their disruption can lead to pathologies. Reporter assays allow the functional validation of enhancers and other regulatory elements, and transgenic mice provide a powerful tool to study gene regulation in vivo. However, these experiments are time-consuming and inefficient. Here, we increase the throughput of transgenic reporter assays by using a piggyBac transposon-based system, and use it to decode the regulatory landscape of atrial fibrillation, a prevalent cardiac arrhythmia. We systematically interrogated ten human atrial fibrillation associated loci, finding five new cardiac-specific enhancers. We also connected novel genes to atrial fibrillation through genome editing and three-dimensional chromatin structure analysis. Of note, we identified a bivalent regulatory element in the second intron of the CAV1 gene at the 7q31 locus, differentially acting upon four genes. Our approach also detected negative regulatory elements such as an ubiquitous silencer in the 16q22 locus that regulates ZFHX3 and can outcompete heart enhancers. Our study characterizes the function of new genetic elements that might be of relevance for the better understanding of gene regulation in cardiac arrhythmias. Thus, we have established a new framework for the efficient dissection of the genetic contribution to common human diseases. Overall design: 4C-seq experiments were perfomed on adult mouse atria to study the genomic interactions of the candidate enhancer regions. Two biological replicates per viewpoint were produced, sent to sequence and analysed independently.