Cytotoxic T lymphocytes form an antigen-independent ring junction
Kristina Somersalo, … , Yuri Sykulev, Michael L. Dustin
Kristina Somersalo, … , Yuri Sykulev, Michael L. Dustin
Published January 1, 2004
Citation Information: J Clin Invest. 2004;113(1):49-57. https://doi.org/10.1172/JCI19337.
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Cytotoxic T lymphocytes form an antigen-independent ring junction

Abstract

Immunological synapses are organized cell-cell junctions between T lymphocytes and APCs composed of an adhesion ring, the peripheral supramolecular activation cluster (pSMAC), and a central T cell receptor cluster, the central supramolecular activation cluster (cSMAC). In CD8+ cytotoxic T lymphocytes, the immunological synapse is thought to facilitate specific killing by confining cytotoxic agents to the synaptic cleft. We have investigated the interaction of human CTLs and helper T cells with supported planar bilayers containing ICAM-1. This artificial substrate provides identical ligands to CD4+ and CD8+ T cells, allowing a quantitative comparison. We found that cytotoxic T lymphocytes form a ring junction similar to a pSMAC in response to high surface densities of ICAM-1 in the planar bilayer. MICA, a ligand for NKG2D, facilitated the ring junction formation at lower surface densities of ICAM-1. ICAM-1 and MICA are upregulated in tissues by inflammation- and stress-associated signaling, respectively. Activated CD8+ T cells formed fivefold more ring junctions than did activated CD4+ T cells. The ring junction contained lymphocyte function associated antigen-1 and talin, but did not trigger polarization and granule translocation to the interface. This result has specific implications for the mechanism of effective CTL hunting for antigen in tissues. Abnormalities in this process may alter CTL reactivity.

Authors

Kristina Somersalo, Nadja Anikeeva, Tasha N. Sims, V. Kaye Thomas, Roland K. Strong, Thomas Spies, Tatiana Lebedeva, Yuri Sykulev, Michael L. Dustin

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Figure 4

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Confocal images of TCR, CD8, talin, and PKC-θ distribution of CTL clones...
Confocal images of TCR, CD8, talin, and PKC-θ distribution of CTL clones. CTL clone CER43 was layered on bilayer containing 200 molecules/μm2 Cy3–ICAM-1–GPI with or without unstained agonist MHCp complexes (a–i) or containing 200 molecules/μm2 Cy3–ICAM-1–GPI only (j, l–s). In k, the CD8-deficient CTL clone 115ix was layered on a bilayer containing 200 molecules/μm2 Cy3-ICAM-1–GPI only. After 30 minutes of incubation, cells were fixed, permeabilized, and stained with the indicated antibodies. Fluorescence was imaged with a laser-scanning confocal microscope. The green signal in j and k is increased to reveal the presence of levels of CD3 staining above the baseline density. (k) CD3 can accumulate in the center of the ring junction without CD8. (b and c) Examples of CD8 accumulations (green) that are filled (b) or have central holes (c). (l and m) Examples of different levels of CD8 in ring junctions, which are not as enriched as in IS. Pairs d/e, n/o, and p/q are the same contacts with talin (green) or ICAM-1 (red) fluorescence; n/o is a ring junction, while p/q is a motile junction, which also shows talin redistribution. Pairs f/g, h/i, and r/s are the same contacts stained for PKC−θ (green) and ICAM-1 (red); f/g shows the classical central localization of PKC-θ compared with h/i, which shows the more frequent localization of PKC-θ to the microaggregates in the pSMAC; r/s shows that PKC-θ was not detected in the ring junctions. The images were processed with a median filter (radius 4 pixels [pixel = .2 μm]). Scale bar: 5 μm.