CD4⁺T-cell synapses involve multiple distinct stages

Abstract

One very striking feature of T-cell recognition is the formation of an immunological synapse between a T cell and a cell that it is recognizing. Formation of this complex structure correlates with cytotoxicity in the case of killer (largely CD8 + ) T-cell activity, or robust cytokine release and proliferation in the case of the much longer lived synapses formed by helper (CD4 + ) T cells. Here we have used electron microscopy and 3D tomography to characterize the synapses of antigen-specific CD4 + T cells recognizing B cells and dendritic cells at different time points. We show that there are at least four distinct stages in synapse formation, proceeding over several hours, including an initial stage involving invasive T-cell pseudopodia that penetrate deeply into the antigen-presenting cell, almost to the nuclear envelope. This must involve considerable force and may serve to widen the search for potential ligands on the surface of the cell being recognized. We also show that centrioles and the Golgi complex are always located immediately beneath the synapse and that centrioles are significantly shifted toward the late contact zone with either B lymphocytes or bone marrow-derived dendritic cells such as antigenpresenting cells, and that there are dynamic, stage-dependent changes in the organization of microtubules beneath the synapse. These data reinforce and extend previous data on cytotoxic T cells that one of the principal functions of the immunological synapse is to facilitate cytokine secretion into the synaptic cleft, as well as provide important insights into the overall dynamics of this phenomenon. microtubule organizing center | centrosome A prominent feature of many T-lymphocyte interactions with other cells on which it recognizes a particular antigen is the formation of an immunological synapse (IS). The formation of this structure correlates with robust signaling, lineage commitment, and fate determination of T cells (1-6) and the directed secretion of cytokines and/or cytotoxic molecules. There is a wholesale reorganization of the T cell's cytoskeleton, surface molecules, and organelles, and the loss of polarity regulators or guidance cues that negatively affect T-cell activation (7,8). Whereas a great deal has been learned about the dynamics of cell-surface and signaling molecules within this interface (9, 10), much less is known about changes within the cell, especially in the long-lived helper T cell, namely in antigen-presenting cell (APC) interactions, which can last for 6 or more h and need continuous T-cell receptor (TCR) engagement (11). Here electron microscopic studies are particularly valuable for their very high resolution, especially with the power of 3D tomography and highpressure freezing, which preserves cell structure more faithfully. Although a number of high-resolution electron microscopy studies of T cell-APC interactions have been reported (3,(12)(13)(14)(15), these involve studies of cytotoxic T or natural killer cells and their targets for only the first few minutes of CD4 + ...

An initial and rapid step of lytic granule secretion precedes microtubule organizing center polarization at the cytotoxic T lymphocyte/target cell synapse

“…3 and Fig. S4), we observed parallel polarization of MTOC and lytic granules at the IS in agreement with previous results obtained using electron microscopy on fixed CTL/target cell conjugates (11,21).…”

Mechanical Criterion for the Rupture of a Cell Membrane under Compression

“…Membrane rupture may occur during cell-cell interactions (52). The pushing forces generated by these filipodia/ protrusions were not measured in these studies.…”

T cell activation requires force generation

“…T cells plunge their invadopodia more than 1 µm deep into APCs (Ueda et al, 2011). Oscillatory movements of the actin cytoskeleton in the lamellipodium were observed for T cells interacting with pMHC on planar lipid bilayers and showed speeds of ∼0.04 µm/s for the cytoskeletal extensions, with a periodicity of ∼2 min (Sims et al, 2007).…”