A multi-enhancer hub at the Ets1 locus controls T cell differentiation and allergic inflammation through 3D genome topology

Multi-enhancer hubs are spatial clusters of enhancers which have been recently characterized across numerous developmental programs. Yet, the functional relevance of these three-dimensional (3D) structures is poorly understood. Here we show that the multiplicity of enhancers interacting with the transcription factor Ets1 is essential to control the precise expression level of this gene in response to cellular cues, and the failure to do so can lead to allergic diseases. Focusing on T cells as a model, we identified a highly connected multi-enhancer hub at the Ets1 locus, comprising a noncoding regulatory element that is a hotspot for sequence variation associated with allergic diseases. We deleted this hotspot and found that the multi-enhancer connectivity is dispensable for T cell development but required for CD4+ T helper (Th1) differentiation in response to changes in the cytokine milieu. Mice lacking this hotspot are thus protected from Th1-mediated colitis but demonstrate an overt allergic response to house dust mites, a T cell-mediated response which is dampened by Th1 cells. Mechanistically, the multi-enhancer hub controls the expression level of Ets1 that is dispensable for the active enhancer landscape but required for the Th1-specific genome topology through recruitment of CTCF. Together, we establish a paradigm for the functional and mechanistic relevance of multi-enhancer hubs controlling cellular competence to respond specifically to an inductive cue.