Mechanical regulation of cytoskeletal dynamics and signaling in cytotoxic T lymphocytes

Cytotoxic T lymphocytes (CTLs) play an integral role in the adaptive immune response. Following engagement of the T cell receptor (TCR) with cognate peptide-MHC on the target cell surface, CTLs release lytic granules containing perforin and granzymes at the CTL-target point of contact, the immune synapse (IS). The cytoskeleton undergoes dramatic reorganization to facilitate synapse formation, targeted granule delivery and force exertion, thus triggering target cell death. CTLs encounter target cells with a range of stiffnesses, from hundreds of pascals to several kilopascals. Prior work has shown that T cell activation, signaling and proliferation depend on substrate stiffness, but the underlying mechanisms remain unknown. We hypothesized that the cytoskeleton plays a central role in mediating this mechanoresponse. We examined the interaction of CTLs with antibody coated elastic hydrogels of tunable rigidity, and found that IS formation, actin accumulation and myosin light chain phosphorylation at the IS are stiffness-dependent. We also found that TCR signaling, centrosome polarization and lytic granule release are tuned by substrate stiffness. These results indicate that synapse formation, early activation and function in CTLs depend on substrate stiffness and suggest that mechanical regulation of the cytoskeleton may be a key factor in modulating this differential response.