Dataset Information

Cytotoxic Granule Exocytosis From Human Cytotoxic T Lymphocytes Is Mediated by VAMP7.

ABSTRACT: Cytotoxic T lymphocytes kill infected or malignant cells through the directed release of cytotoxic substances at the site of target cell contact, the immunological synapse. While genetic association studies of genes predisposing to early-onset life-threatening hemophagocytic lymphohistiocytosis has identified components of the plasma membrane fusion machinery, the identity of the vesicular components remain enigmatic. Here, we identify VAMP7 as an essential component of the vesicular fusion machinery of primary, human T cells. VAMP7 co-localizes with granule markers throughout all stages of T cell maturation and simultaneously fuses with granule markers at the IS. Knock-down of VAMP7 expression significantly decreased the killing efficiency of T cells, without diminishing early T cell receptor signaling. VAMP7 exerts its function in a SNARE complex with Syntaxin11 and SNAP-23 on the plasma membrane. The identification of the minimal fusion machinery in T cells provides a starting point for the development of potential drugs in immunotherapy.

SUBMITTER: Chitirala P

PROVIDER: S-EPMC6692471 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Cytotoxic Granule Exocytosis From Human Cytotoxic T Lymphocytes Is Mediated by VAMP7.

Chitirala Praneeth P Ravichandran Keerthana K Galgano Donatella D Sleiman Marwa M Krause Elmar E Bryceson Yenan T YT Rettig Jens J

Frontiers in immunology 20190807

Cytotoxic T lymphocytes kill infected or malignant cells through the directed release of cytotoxic substances at the site of target cell contact, the immunological synapse. While genetic association studies of genes predisposing to early-onset life-threatening hemophagocytic lymphohistiocytosis has identified components of the plasma membrane fusion machinery, the identity of the vesicular components remain enigmatic. Here, we identify VAMP7 as an essential component of the vesicular fusion mach ...[more]

PMID: 31447853

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Project description:Granules of cytotoxic T lymphocytes (CTL) are derived from the lysosomal compartment. Synaptotagmin7 (Syt7) appears to be the calcium sensor triggering fusion of lysosomes in fibroblasts. Syt7 has been proposed to control cytotoxic granule (CG) fusion in lymphocytes and mice lacking Syt7 have reduced ability to clear infections. However, fusion of CG persists in the absence of Syt7. To clarify the role of Syt7 in CTL function, we have examined the fusion of cytotoxic granules of CD8+ T-lymphocytes from Syt7 knock-out mice. We have recorded granule fusion in living CTL, using total internal reflection microscopy. Since Syt7 is considered a high affinity calcium-sensor specialized for fusion under low calcium conditions, we have compared cytotoxic granule fusion under low and high calcium conditions in the same CTL. There was no difference in latencies or numbers of fusion events per CTL under low-calcium conditions, indicating that Syt7 is not required for cytotoxic granule fusion. A deficit of fusion in Syt7 KO CTL was seen when a high-calcium solution was introduced. Expressing wild type Syt7 in Syt7 KO lymphocytes reversed this deficit, confirming its Syt7-dependence. Mutations of Syt7 which disrupt calcium binding to its C2A domain reduced the efficacy of this rescue. We counted the cytotoxic granules present at the plasma membrane to determine if the lack of fusion events in the Syt7 KO CTL was due to a lack of granules. In low calcium there were no differences in fusion events per CTL, and granule numbers were similar. In high calcium, granule number was similar though wild type CTL exhibited significantly more fusion than Syt7 KO CTL. The modest differences in granule counts do not account for the lack of fusion in high calcium in Syt7 KO CTL. In Syt7 KO CTL expressing wild type Syt7, delivery of cytotoxic granules to the plasma membrane was comparable to that of wild type CTL. Syt7 KO CTL expressing Syt7 with deficient calcium binding in the C2A domain had significantly less fusion and fewer CG at the plasma membrane. These results indicate that Syt7 is involved in trafficking of CG to the plasma membrane.

Dataset Information

Cytotoxic Granule Exocytosis From Human Cytotoxic T Lymphocytes Is Mediated by VAMP7.

Publications

Cytotoxic Granule Exocytosis From Human Cytotoxic T Lymphocytes Is Mediated by VAMP7.

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