PROJECT SUMMARY – CORE C Efficient and reproducible detection of immune cell states is paramount to our understanding of the immune responses in the context of health and disease, such as cancer and autoimmunity. Immune function is highly dependent upon costimulatory signals where their role has become apparent in the study of tumor infiltrating lymphocytes with PD-1/PD-L1 expression, as well as in autoimmune disease particularly with activation of autoreactive T cells and costimulatory receptor CD226 and its ligand CD155 which is shared with TIGIT. Achieving a mechanistic understanding by which PD-1/PD-L1 and TIGIT/CD155 axes regulate T cell and myeloid cell function will enable refinement of the therapeutic use and target of these costimulatory pathways. The Single Cell and Spatial Multi-omics Core (Core C) at Yale University will provide support for novel transcriptional, epigenetic, and spatial profiling of single cells as well as enriched immune cell types across all three projects of the Program Project Grant. Our core has established systematic pipelines for processing single cell RNA-seq and ATAC-seq from low-input samples, from single cells or single nuclei, as well as for spatial transcriptomics using DBiT-seq, a single cell spatial technology newly developed at Yale which will be used in both experimental models and in human tissues across the three projects. This core provides capabilities and a pipeline for the preparation, sequencing and single cell analysis and data integration across species. Our experimental and computational resource will facilitate (1) the investigations of immunity in mouse models that are deficient for TIGIT, PD1, or both, (2) the characterization of cells or nuclei or sections human tissues from patients with multiple sclerosis and glioblastoma, and (3) the single cell, integrative analysis of all data generated in Projects 1-3. Overall, the outcome will be a better understanding of co-stimulatory and co-inhibitory mechanisms, and its function in regulating inflammasome activation in T cells and myeloid cells in human diseases and disease models. This may potentially lead to new avenues for therapy for autoimmunity and cancer.