| Cytotoxic lymphocytes (CTLs) play an important role in the immune response by eliminating virally infected or tumor cells through the Ca2+-mediated exocytosis of granzyme and perforin molecules, stored in secretory lysosomes. Previous work from our laboratory showed that the ubiquitously-expressed, high affinity calcium-binding synaptotagmin, Syt VII, modulates the exocytosis of lysosomes in several cell types. Here we examined the role of Syt VII in the regulation of 'secretory lysosome' exocytosis in CTLs, a low [Ca2+]-dependent process. Using CD8+ CTLs generated from Syt VII wild type (WT) and knockout (KO) mice, we investigated its cellular localization and function in secretory lysosomes. Syt VII is expressed in CTLs and co-localizes with granzyme A and Lamp-1 on the secretory lysosome by immunofluorescence. No morphological differences were observed between the CTL lytic granules from either Syt VII WT or KO mice, and similar levels of granzyme A and perform were found to be expressed in CTLs from both genotypes. However, cytotoxicity assays revealed that Syt VII KO CTLs have an impaired ability to kill target cells, in both allogeneic and peptidespecific assays. Although their granule-mediated cytotoxicity is reduced, we found normal polarization of secretory lysosomes towards target contact sites and the normal formation of an immunological synapse in Syt VII KO CTLs. The cytokine secretion and proliferation response after anti-CD3 antigen stimulation of Syt VII KO CTLs was also comparable to controls. Interestingly, in vivo cytotoxicity and tumor growth assays revealed an enhanced response against syngeneic cells, confirming an autoimmunity response in Syt VII KO mice. Taken together, our findings reveal that secretory lysosomal exocytosis in CTLs is regulated by Synaptotagmin VII. |
