Project Summary Interleukin-2 is a potent T cell growth factor with crucial roles in both immunity and self-tolerance. Genome-wide association studies (GWAS) in humans have shown that genetic variation at the IL2 and IL2RA loci influence susceptibility to multiple immune-mediated diseases including allergic asthma, systemic lupus erythematosus, and inflammatory bowel disease (IBD), identifying this as a key molecular axis that controls immune activity. However, mechanistic basis of disease risk for these polymorphisms is poorly understood. We have new evidence that the cis-regulatory architectures of IL2 and IL2RA extend much further from the gene than previously appreciated, encompassing regions harboring known disease-associated variants, and we hypothesize that polymorphisms within these elements control the level and timing of IL-2 and IL-2RA expression to control the balance of tolerance vs. inflammation. In this application, we propose a comprehensive screen for potential cis-regulatory elements that interact with the IL2 and IL2RA genes using state-of-the-art epigenomic approaches like ATAC-seq and Capture-C-seq. We will establish how these elements contribute to IL2 and IL2RA gene expression using powerful, CRISPR/CAS9-based genome editing approaches. To study the impact of disease-associated genetic variation at these regulatory elements on immune function in in vivo systems, we will analyze the responses of genetically characterized subjects curated from the Benaroya Research Institute biorepository, and create mice in which orthologous murine Il2 and Il2ra enhancer sequences have been replaced with human risk alleles, assessing the disease susceptibility in models of allergic asthma, SLE and IBD. Our studies will provide comprehensive maps of the transcriptional architecture of IL2 and IL2RA, insights into the molecular basis for the genetic association of IL2 and IL2RA with autoimmune disease, and may guide the design of new approaches for the treatment of organ transplant rejection and inflammatory disease.