3D genome organization of the Ets1-Fli1 locus controls allergic responses

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The goal of this proposal is to study how perturbation in three-dimensional (3D) genome folding alters CD4+ T cell function, mediating allergic disorders. T cell identity depends on not only the linear genome sequence that embeds millions of regulatory elements, but also the 3D chromatin architecture that orchestrates the spatial localization of the regulatory elements with their target genes. Recent advances in our understanding of nuclear organization indicate that single-nucleotide polymorphisms associated with immune-mediated diseases may impact gene regulation through altered 3D genomic structure and reorganization of large genomic regions in the disease relevant cell types. However, the link between sequence variation, cellular context, 3D genome folding, and aberrant gene expression in majority of immune-mediated complex diseases remains largely unknown. Our objective is to determine the molecular processes through which 3D genome organization in T cells is linked to allergic disorders. We formulated this objective based on four unexpected observations: (A) Our algorithmic definition of groups of densely interacting multi-enhancer elements, which we called 3D cliques, revealed that a locus harboring the Ets1 and Fli1 genes is hyperconnected in T cells. (B) This unique 3D genome architecture is conserved in human T cells coinciding with multiple polymorphisms associated to type 2 immune diseases including allergy, asthma, and atopic dermatitis. (C) We generated a novel strain of mice by deleting a non-coding sequence homologous to the allergy-associated polymorphic region in the human genome, ~250kbp downstream of the Ets1 promoter. This genetic deletion left T cell development intact but led to major defects in CD4+ T helper 1 (Th1) differentiation. Th1 cells are responsible for the control of intracellular pathogens such as bacteria and dampen Th2 responses to allergens. Hence, limited Th1 differentiation due to genetic modification of the Ets1-Fli1 3D clique may cause allergic responses. (D) We modeled the type 2 immune responses in vivo using house dust mites. In the lung tissues of mice with a deletion in the non-coding sequence in the Ets1-Fli1 3D clique, we detected a dramatic increase in allergic responses characterized by a significant accumulation of eosinophils and Th2 cells and a reduction in Th1 cells. These unpublished data provide us with compelling evidence that our engineered mouse strain is a model for understanding the role of noncoding regulatory elements and 3D genome folding in type 2 immune diseases. However, detailed cellular and molecular mechanisms through which genetic deletion in the Ets1-Fli1 locus causes overt allergic responses remain to be understood. Moreover, the generalizability of our 3D clique analysis to additional pathogenic regulatory nodes remains to be examined. This work is significant because it is the first-ever mechanistic investigation providing a connection between genome architecture and type 2 immune diseases.