Molecular anatomy of antigen-specific CD8+ T cell engagement and synapse formation in vivo

Abstract

Antigen-specific CD8+ T cells are required for the clearance of most viral infections and several cancers. However, it is not clear in vivo whether CD8+ T cells can engage multiple targets simultaneously, engagement results in the formation of an immunologic synapse or molecules involved in CD8 function are redistributed to the synapse. We used here high-resolution microscopy to visualize interactions between virus-specific effectors and target cells in vivo. Using either in situ tetramer staining or green fluorescent protein–labeled virus-specific T cells, we have shown that a single CD8+ T cell can engage two or three targets, a synapse occurs at the site of engagement and molecules involved in attachment (lymphocyte function–associated antigen 1), signaling (Lck) and lytic activity (perforin) are differentially positioned on the T cell. In addition, we have established an in vivo approach for assessing the intricacies of antigen-specific T cell activation, migration, engagement, memory and other defining elements of adaptive immunity.

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Figure 1: Tetramer staining of virus-specific CD8+ T cells in the spleens and CNS of GP33 TCR–Tg mice.
Figure 2: Visualization of antiviral immunity in the CNS of B6 mice during the induction of lethal meningitis.
Figure 3: Expansion and migration of GFP+ Db-GP(33–41)-specific T cells after an i.c. infection.
Figure 4: Cellular reorganization of Db-GP(33–41)-specific T cells juxtaposed to LCMV-infected targets.
Figure 5: Patterns of interfacial LFA-1, Lck and perforin staining on Db-GP(33–41)-specific T cells in the CNS.
Figure 6: LFA-1 and Lck 3D localization on Db-GP(33–41)-specific T cells in the CNS.

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Acknowledgements

Supported by NIH grant AI09484, training grant AG00080 (to D. B. M.) and Juvenile Diabetes Research Foundation Award 3-2000-510 (to U. C.)

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Correspondence to Dorian B. McGavern.

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Supplementary information

Web Movie 1.

A 3D rotation of a GFP+ Db-GP(33-41)-specific T cell (green) in juxtaposition with three nucleated (blue) LCMV-infected targets (red) in the CNS. (MOV 6527 kb)

Web Movie 2.

A 3D rotation illustrating the distribution of LFA-1 (red) on the same GFP+ Db-GP(33-41)-specific T cell (green) shown in Web Movie 1. Nuclei are shown in blue. Note the polarization of LFA-1 toward the virus-infected targets and the LFA-1 coated process that the CTL has extended. (MOV 6150 kb)

Web Movie 3.

A 3D rotation illustrating the distribution of LFA-1 (red) on GFP+ Db-GP(33-41)-specific T cell (green) that has trafficked to the CNS but is not engaged with a LCMV-infected target. Note the homogenous clusters of LFA-1 around the cell membrane. Nuclei are shown in blue. (MOV 2824 kb)

Web Movie 4.

A 3D rotation of a GFP+ Db-GP(33-41)-specific T cell (green) in juxtaposition with a single LCMV-infected target (blue) in the CNS. Note the interfacial aggregation of Lck (red) on the CTL. (MOV 3029 kb)

Web Movie 5.

A 3D rotation showing the distribution of Lck (red) on a GFP+ Db-GP(33-41)-specific T cell (green) not engaged with an LCMV-infected target. Note the homogenous distribution of Lck on the CTL membrane. Nuclei are shown in blue. (MOV 2817 kb)

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McGavern, D., Christen, U. & Oldstone, M. Molecular anatomy of antigen-specific CD8+ T cell engagement and synapse formation in vivo. Nat Immunol 3, 918–925 (2002). https://doi.org/10.1038/ni843

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