The authors use tiled CRISPR activation for functional enhancer discovery across two autoimmunity risk loci, CD69 and IL2RA, and identify elements with features of stimulus-responsive enhancers, including an IL2RA enhancer that harbours a fine-mapped autoimmunity risk variant. Enhancers are gene regulatory elements that shape cell-type-specific transcriptional programs and responses to specific extracellular cues. Mapping enhancer function is challenging because of our limited understanding of the cellular context in which each enhancer contributes to gene regulation. Here, Alexander Marson and colleagues use a tiled CRISPR activation (CRISPRa) approach for functional enhancer discovery across two autoimmunity risk loci: CD69 and IL2RA. They identify several elements with features of stimulus-responsive enhancers, including an IL2RA enhancer that contains an autoimmunity risk variant. This approach should be useful for discovering functional enhancers without prior knowledge of their specific biological context. The majority of genetic variants associated with common human diseases map to enhancers, non-coding elements that shape cell-type-specific transcriptional programs and responses to extracellular cues1,2,3. Systematic mapping of functional enhancers and their biological contexts is required to understand the mechanisms by which variation in non-coding genetic sequences contributes to disease. Functional enhancers can be mapped by genomic sequence disruption4,5,6, but this approach is limited to the subset of enhancers that are necessary in the particular cellular context being studied. We hypothesized that recruitment of a strong transcriptional activator to an enhancer would be sufficient to drive target gene expression, even if that enhancer was not currently active in the assayed cells. Here we describe a discovery platform that can identify stimulus-responsive enhancers for a target gene independent of stimulus exposure. We used tiled CRISPR activation (CRISPRa)7 to synthetically recruit a transcriptional activator to sites across large genomic regions (more than 100 kilobases) surrounding two key autoimmunity risk loci, CD69 and IL2RA. We identified several CRISPRa-responsive elements with chromatin features of stimulus-responsive enhancers, including an IL2RA enhancer that harbours an autoimmunity risk variant. Using engineered mouse models, we found that sequence perturbation of the disease-associated Il2ra enhancer did not entirely block Il2ra expression, but rather delayed the timing of gene activation in response to specific extracellular signals. Enhancer deletion skewed polarization of naive T cells towards a pro-inflammatory T helper (TH17) cell state and away from a regulatory T cell state. This integrated approach identifies functional enhancers and reveals how non-coding variation associated with human immune dysfunction alters context-specific gene programs.

Discovery of stimulation-responsive immune enhancers with CRISPR activation