Spatiotemporal and molecular factors determining T cell stemness and differentiation in autoimmune Type 1 Diabetes 2328677

Abstract Introduction Type 1 diabetes (T1D) is a T cell—mediated autoimmune disease driven by β cell-specific CD8 T cells. How autoreactive T cells arise and sustain disease remains unclear. Using the non-obese diabetic mouse model of T1D, we previously identified a stem-like CD8 T cell population in the pancreatic lymph node (pLN) which initiates and sustains β cell destruction: stem T cells (TSC) self-renew and continuously give rise to differentiated progeny (TDIFF) that migrate to the pancreas and kill β cells. Spatial positioning of somatic stem cells is critical for their maintenance, and that niche restricted signals (i.e., WNT and NOTCH) regulate the balance between self-renewal and differentiation. However, if and how T cell stemness is associated with distinct intranodal positioning in pLN, and whether interference in migration disrupts differentiation, is unknown. Methods We conducted (i) paired single-cell RNA- and ATAC-sequencing, (ii) adoptive T cell transfer studies, (iii) high-resolution imaging, (iv) CRISPR/Cas9 mediated gene editing of autoimmune T cells in pLN to identify the molecular and functional characteristics of TSC and TDIFF. Results We discovered unique transcription factors and epigenetic programs governing autoimmune T cell stemness and differentiation. TSC and TDIFF express distinct chemokine receptors and integrins, suggesting that T cell stemness and differentiation are driven by intranodal positioning. Strikingly, WNT and NOTCH signaling were enriched in TSC, driving the expression of critical stem genes, thereby connecting T cell stemness to somatic stem cell biology. Pharmacological blockade and CRISPR/Cas9-mediated deletion of integrins and cell-cell interactions prevented autoimmune T cell differentiation and disease. Conclusion Our studies identify novel transcriptional regulators and niche-dependent signals that determine autoimmune T cell stemness and differentiation, opening novel therapeutic avenues for the prevention and treatment of T1D and other autoimmune diseases. Funding Source NIH F31DK141119, NIH R01AI173249, Juvenile Diabetes Research Foundation JDRF SRA-2023-1410-S-B, MSKCC Basic Research Innovation Award (BRIA), Hearst Foundation Topic Categories Lymphocyte Differentiation and Peripheral Maintenance (LYM)