ABSTRACT Eosinophilic inflammation is a key feature of many human pathologies, and therapeutic targeting of eosinophils is of clinical interest, as illustrated by the new class of eosinophil-depleting drugs in development for allergic and inflammatory diseases. The full potential and consequences of these drugs are active areas of medical research, particularly as eosinophil depletion does not uniformly cause symptom reduction. Despite their post- mitotic state and short circulating lifespan, eosinophils can persist for days to weeks in certain tissue environments. Increasingly, there is evidence of a positive role of long-lived, “tissue-resident” eosinophils in tissue structure development and maintenance that counters the historic view of eosinophils as only having pro-inflammatory functions. Given the prevalence of eosinophilic inflammatory conditions and development of eosinophil-depleting drugs, there is a significant need to understand 1) which factors enable eosinophil differentiation, 2) how differentiated eosinophils interact with the surrounding tissue, and 3) what molecular events prolong survival of an otherwise short-lived post-mitotic cell. This project will elucidate the regulation and function of tissue-resident eosinophils at steady state and during allergic disease. We hypothesize that eosinophils respond to pro-survival signals in the epithelial environment by engaging cell cycle machinery to prevent apoptosis and support rapid gene expression during specialization. The three research areas will 1) target cell cycle machinery to manipulate eosinophil specialization in mucosal tissue, 2) delineate pre- and post-transcriptional events in eosinophil re-specialization, and 3) define and inhibit pathogenic functions of specialized eosinophils. These studies are technically and conceptually innovative in that they use high- sensitivity immunoassays, 3D organotypic tissue models, next-generation sequencing, and chromatin looping technologies to expand on paradigm-shifting evidence of a dynamic, long-lasting role of eosinophils in tissue. The technical approach is creative and designed to address eosinophil functions that are poorly understood but central to understanding and treating eosinophilic diseases, type 2 allergic diseases, autoimmune pathologies, and certain cancers. This research is ideally suited to the NIAID DP2 award due to its potentially transformative effect on the understanding of granulocyte biology. The applicant is well suited to lead the proposed work given her technical and analytical skill, research productivity, and success in creative problem-solving and collaborative project development. The DP2 award will augment the resources accompanying her newly independent position and allow Dr. Dunn to apply impactful new technologies to a clinically relevant but understudied area of eosinophil biology that has broader implications for the converging fields of clinical immunology, personalized medicine, and nuclear str...