Abstract B190: WASP-dependent actin protrusions mechanically potentiate killing by cytotoxic T-cells

Cytotoxic T Lymphocytes (CTLs) play a central role in immune responses against intracellular pathogens and cancer. Cytotoxic responses are induced by the formation of the immunologic synapse after the recognition of the peptide-major histocompatibility complex by the T-cell receptor. Synapse formation is associated dramatic reorganization of both microtubules and filamentous actin (F-actin) within the T-cell. This promotes the directional secretion of toxic perforin and granzymes into the intercellular space, enhancing both the potency and the specificity of targeT-cell killing. We have previously demonstrated that force exertion by CTLs at the immunologic synapse was strongly correlated with the cytotoxic potential. Indeed, force exertion enhances cytotoxicity by increasing membrane tension on the target cell, which in turn promotes the pore-forming activity of secreted perforin. This correlation between applied force and biochemical responses, which we referr to as mechanopotentiation, raised the prospect that CTLs might use three-dimensional structures at the immunologic synapse to coordinate force exertion and lytic granules secretion. In the present study, we investigated the mechanisms underlying mechanopotentiation in CTLs using a combination of three-dimensional micropatterned stimulatory substrates and high-resolution imaging. Our results revealed that CTLs couple lytic granule release with the formation of highly dynamic F-actin rich protrusions. These protrusions, which are generated by the Wiskott-Aldrich Syndrome protein (WASP) and the Arp2/3 actin nucleation complex, are required for coordinating force exertion with cytolytic secretion allowing efficient killing. Our results provide insight into how cells organize mechanical output and emphasize the importance of studying complex, communicative interfaces in three dimensions. Citation Format: Fella Tamzalit, Mitchell S. Wang, Weiyang Jin, Vitaly Boyko, John M. Heddleston, Charles T. Black, Lance C. Kam, Morgan Huse. WASP-dependent actin protrusions mechanically potentiate killing by cytotoxic T-cells [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr B190.