Interleukin-2 is a potent T cell growth factor with crucial roles in both immunity and tolerance. Genetic studies in humans and mice demonstrate a role for IL2 in autoimmune disease susceptibility, and for decades the proximal IL2 upstream regulatory region has served as a paradigm of tissue-specific, inducible gene regulation. We have used a combination of ChIP-seq, ATAC-seq and 4C-seq analyses of of chromatin and chromosome remodeling to identify a novel, long-range enhancer of the IL2 gene located 83 kb upstream of the transcription start site. This element can potently enhance IL2 transcription in recombinant reporter assays in vitro, and the native region loops to physically interact with the IL2 gene in vivo in a CD28-dependent manner. We have now edited this ~500 bp element out of the mouse genome using CRISPR/CAS9 in fertilized eggs, and CD4+ T cells from animals homozygous for deletion of the IL2-83 enhancer exhibit a ~80% reduction in IL-2 secretion in vitro. Preliminary analysis of young IL2-83 enhancer-deficient animals suggests largely normal hematopoiesis, but accumulation of activated phenotype CD4+ T cells in the peripheral lymphoid tissues. This cis regulatory element is evolutionarily conserved, and contains human SNPs associated with multiple autoimmune disorders. These results indicate that the regulatory architecture of the IL2 locus is more complex than previously appreciated, and suggest a novel molecular basis for the genetic association of IL2 polymorphism with autoimmune disease. This IL2-83 enhancer knock-out mouse represents a valuable model to study graded effects of this cytokine on activation-induced cell death, CD4 T cell differentiation, CD8 T cell memory, and regulatory T cell homeostasis in vivo.

CITATION INFORMATION: Thomas R, Mehra P, Johnson M, Grant S, Wells A. A New and Potent Distal Enhancer of the IL2 Gene. Am J Transplant. 2017;17 (suppl 3).

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