This project leverages high-performance computing to investigate how innate and adaptive lymphocytes regulate tissue immunity across the gastrointestinal and pulmonary systems. Building on our recent work characterizing type 3 innate lymphoid cells (ILC3s) and their ability to activate latent TGF-β1 in the human gut, we have expanded the scope of our research to include spatial, longitudinal, and multimodal profiling of lymphocyte dynamics in multiple disease contexts. The first phase of this project used NAISS resources to analyze single-cell RNA sequencing and public atlas data to define a conserved molecular program by which ILC3s activate TGF-β1 in both healthy and inflamed gut tissue. These results are featured in the manuscript “ILC3s modulate immune-epithelial interactions in the gut via TGF-β1 activation,” currently accepted pending revision in Mucosal Immunology. We now build on these findings with several new projects focused on lymphocyte-mediated tissue pathology and treatment response. In the gastrointestinal tract, we are conducting a spatially resolved, multimodal single-cell analysis of colonic tissue from Crohn’s disease patients, specifically examining how innate and adaptive lymphocytes become disorganized across epithelial, lamina propria, and submucosal layers during inflammation. In parallel, we are analyzing a longitudinal cohort of Crohn’s patients undergoing anti-TNF therapy, using paired blood and gut biopsies before and after treatment to study shifts in T cell infiltration, clonality, and phenotype in responders and non-responders. In colorectal cancer, we aim to characterize the role of B cells and plasma cells in primary tumors and peritoneal metastases using single-cell RNA sequencing and B cell receptor repertoire analysis. This work will be performed by a collaborating postdoctoral researcher and will inform future funding applications. To extend our analysis to the lung, another group member is applying these computational methods to large single-cell datasets from severe asthma patients. These data will be used to characterize disease-specific immune cell phenotypes and support biomarker discovery, with implications for patient stratification and therapy development. This project plays a central role in multiple PhD and postdoctoral research tracks within our group. It supports the doctoral dissertation work of the main applicant, John Bassett, while also serving as a shared infrastructure platform for researchers focused on asthma (Patrick Shearer), colorectal cancer (Anne Marchelot), and reproductive immunology (Emma Patey). As such, the project promotes collaborative, scalable, and reproducible analysis workflows across disease areas and organ systems. NAISS computational resources remain essential for enabling high-throughput analysis, storage, and collaborative development of pipelines for these large and complex single-cell datasets.