Mechanical regulation of cytoskeletal organization, signaling and force generation in cytotoxic T lymphocytes

Cytotoxic T lymphocytes (CTLs) are an integral part of the adaptive immune response. Upon activation by engaging antigen on the surface of target cells, they form an Immune Synapse at the contact site at which they deliver lytic granules leading to target cell death. CTLs encounter target cells with a wide range of stiffnesses. How CTLs adapt and respond to these different stiffnesses and optimize their function is not well known. We hypothesized that the cytoskeleton plays a central role in mediating this mechanoresponse. We performed live-cell confocal imaging of mouse CTLs activated on ligand-coated hydrogels of varying stiffness to mimic target cells. We observed that early signaling activation markers such as calcium influx as well as cell shape, synapse morphology and actin organization are modulated by stiffness of the activating surface. We further found that centrosome polarization and lytic granule delivery are also tuned to the rigidity of hydrogels. Traction force microscopy revealed that CTLs exert higher forces on stiffer substrates during activation. Our results suggest that the cytoskeleton may be a key factor in modulating CTL response to stiffness. Our work highlights the importance of mechanical cues in immune function during CTL-target cell interaction and indicates that CTLs are tuned to respond to the biophysical properties of their targets.