A pleiotropic hypoxia-sensitive EPAS1 enhancer is disrupted by adaptive alleles in Tibetans [Paired ATAC-seq & RNA-seq]

Background & Aims: A major focus of human genetics research has been to identify locally adapted alleles in global populations. In Tibetans, noncoding alleles in EPAS1 – who’s protein product HIF-2 is a key driver of the response to hypoxia – carry some of the strongest signatures of positive selection found in humans, yet their functional mechanism has never been systematically examined. Here we report the discovery of three enhancers within EPAS1, whose activity is significantly disrupted by Tibetan alleles in one or more key organs (endothelium, kidney, and heart). We further characterize one of these enhancers (ENH5) whose activity was found to be not only allele-specific, but also hypoxia-dependent in all three cell types. Deletion of this enhancer results in downregulation of EPAS1 and HIF-2 targets in acute hypoxia as well as a blunting of the transcriptional response to sustained hypoxia. In vivo deletion of the orthologous ENH5 in mice results in dysregulation of gene expression across multiple tissues. We propose that pleiotropic adaptive effects of the Tibetan alleles in EPAS1 underlie the strong selective signal at this gene. ATAC-sec/RNA-sec in HAECs: Paired ATAC-seq and RNA-seq were performed on primary human aortic endothelial cells (HAECs). Triplicate samples of cells cultured for 24 hours in either normoxia (21% O2) or hypoxia (1% O2), were harvested in parallel and processed into ATAC-seq libraries using the Nextera DNA library prep kit (FC-121-1030) or RNA-seq libraries using TruSeq RNA Library Preparation Kit v2, Set B (Illumina, RS-122-2002)