The exocytosis of lytic granules is impaired in Vti1b- or Vamp8-deficient CTL leading to a reduced cytotoxic activity following antigen-specific activation.
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.
Dressel
Ralf
R
Department of Cellular and Molecular Immunology, University Medical Center, University of Göttingen, Göttingen, Germany. rdresse@gwdg.de
Elsner
Leslie
L
Novota
Peter
P
Kanwar
Namita
N
Fischer von Mollard
Gabriele
G
D000818:Animals; D000941:Antigens / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology / Q000378:metabolism / Q000502:physiology; D015550:Cell Degranulation / Q000276:immunology; D045744:Cell Line, Tumor; D003594:Cytoplasmic Granules / Q000276:immunology / Q000378:metabolism; D003602:Cytotoxicity, Immunologic / Q000235:genetics / Q000276:immunology; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D005089:Exocytosis / Q000276:immunology*; D005260:Female; D005434:Flow Cytometry; D015870:Gene Expression; D006801:Humans; D008213:Lymphocyte Activation / Q000276:immunology*; D051907:Lysosomal Membrane Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D018345:Mice, Knockout; D008822:Mice, Transgenic; D050766:Qb-SNARE Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D050683:R-SNARE Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology* / Q000378:metabolism; D052118:Lysosomal-Associated Membrane Protein 1
D000941:Antigens; D051907:Lysosomal Membrane Proteins; D050766:Qb-SNARE Proteins; D050683:R-SNARE Proteins; C052255:Lamp1 protein, mouse; D052118:Lysosomal-Associated Membrane Protein 1; C491831:Vamp8 protein, mouse; C491415:Vti1b protein, mouse