The exocytosis of lytic granules is impaired in Vti1b- or Vamp8-deficient CTL leading to a reduced cytotoxic activity following antigen-specific activation

Abstract

The exocytosis of cytotoxic proteins stored in lytic granules of activated CTL is a key event during killing of target cells. Membrane fusion events that are mediated by soluble N-ethylmaleimide-sensitive-factor attachment protein receptor (SNARE) proteins are crucial, as demonstrated by patients with familial hemophagocytic lymphohistocytosis type 4 who have mutations in the SNARE protein syntaxin-11 that result in an impaired degranulation of cytotoxic cells. We found an increased mRNA expression of the SNARE protein genes Vti1b and Vamp8 during Ag-specific activation of CTL from TCR-transgenic OT-I mice. Therefore, we investigated the cytolytic activity of CTL from TCR-transgenic Vti1b and Vamp8 knockout mice. At 3 d as well as at 4 d of Ag-specific stimulation, the degranulation of CTL was significantly reduced in Vti1b and Vamp8 knockout mice, as determined by cell surface expression of the degranulation marker CD107a. After 3 d of Ag-specific stimulation, the cytolytic activity of Vti1b- and Vamp8-deficient CTL was reduced to approximately 50% compared with heterozygous controls. However, 4 d after stimulation, the cytotoxic activity of Vti1b- as well as Vamp8-deficient CTL was not impaired anymore. The capacity of Vti1b- and Vamp8-deficient dendritic cells to process Ags and to stimulate the proliferation of CTL was not reduced, arguing against an indirect effect on the activation of CTL. These findings suggest a role of the SNARE proteins vti1b and vesicle-associated membrane protein 8 in the degranulation of CTL. However, a deficiency can apparently be compensated and affects only transiently the cytotoxic activity of CTL during their development to armed effector cells.