Biomechanical stiffness is essential for CD8+ T cell activation and proliferation 2258840

Abstract Introduction The cytoskeleton maintains cellular shape and structure, and serves as a biomechanical regulator of cellular responses. Although CD8+ T cell activation is well characterized, the roles of mechanotransduction and cytoskeletal stiffness remain unclear. Understanding how these biomechanical properties affect CD8+ T cell activation and function could provide new insight into immune regulation. Methods We examined CD8+ T cells from a Protein Tyrosine Phosphatase Non-Receptor Type 21 knockout (Ptpn21⁻/⁻, KO) mouse model, previously reported to exhibit a disorganized actin cytoskeleton in hematopoietic stem cells. Intrinsic biomechanical properties were quantified using atomic force microscopy (AFM) with Hertzian modeling. Functional consequences of altered stiffness were evaluated by flow cytometry, comparing KO and wild-type (WT) mice after in vitro stimulation and in vivo adoptive transfer of CD8+OT-I T cells into B16-OVA tumor-bearing recipients. Results AFM revealed that Ptpn21⁻/⁻ CD8+ T cells were mechanically softer with a reduced elastic modulus. This softening was functionally significant to CD8+ T cell behavior. KO cells showed reduced activation upon CD3/CD28 bead stimulation, while responses to soluble OVA peptide remained comparable, indicating defective mechanotransduction. Calcium-flux assays demonstrated reduced store-operated Ca²+ entry, and AFM showed prolonged relaxation time, together suggesting impaired mechanotransduction during early TCR signaling. In vivo, the percentage of KO CD8+ T cells circulating in the spleen, lymph nodes, and peripheral blood was significantly lower than that of WT cells in a competitive adoptive transfer model, and the residual KO cells were less activated and proliferative. Conclusion In conclusion, our findings demonstrate that cytoskeletal softening in Ptpn21⁻/⁻ CD8+ T cells impairs mechanotransduction, resulting in weakened T cell activation and reduced proliferation within a tumor-bearing environment. Funding Source HL162725, HL130995, CA282579 Topic Categories Immune Response Regulation: Cellular Mechanisms (IRC)