Semmaphorin 3 A causes immune suppression by inducing cytoskeletal paralysis in tumour-specific CD8 T cells.

Semaphorin-3A (SEMA3A) functions as a chemorepulsive signal during development and can affect T cells by altering their filamentous actin (F-actin) cytoskeleton. The exact extent of these effects on tumour-specific T cells are not completely understood. Here we demonstrate that Neuropilin-1 (NRP1) and Plexin-A1 and Plexin-A4 are upregulated on stimulated CD8 T cells, allowing tumour-derived SEMA3A to inhibit T cell migration and assembly of the immunological synapse.

Dynamics and spatial organization of Kv1.3 at the immunological synapse of human CD4+ T cells.

Formation of the immunological synapse (IS) is a key event during initiation of an adaptive immune response to a specific antigen. During this process, a T cell and an antigen presenting cell form a stable contact that allows the T cell to integrate both internal and external stimuli in order to decide whether to activate. The threshold for T cell activation depends on the strength and frequency of the calcium (Ca) signaling induced by antigen recognition, and it must be tightly regulated to avoid undesired harm to healthy cells.

Comparison of different methods for isolating CD8 T lymphocyte-derived extracellular vesicles and supramolecular attack particles.

CD8 T lymphocytes play vital roles in killing infected or deranged host cells, recruiting innate immune cells, and regulating other aspects of immune responses. Like any other cell, CD8 T cells also produce extracellular particles. These include extracellular vesicles (EVs) and non-vesicular extracellular particles (NVEPs).

Clathrin mediates both internalization and vesicular release of triggered T cell receptor at the immunological synapse.

Ligation of T cell receptor (TCR) to peptide-MHC (pMHC) complexes initiates signaling leading to T cell activation and TCR ubiquitination. Ubiquitinated TCR is then either internalized by the T cell or released toward the antigen-presenting cell (APC) in extracellular vesicles. How these distinct fates are orchestrated is unknown.

An intercellular transfer of telomeres rescues T cells from senescence and promotes long-term immunological memory.

The common view is that T lymphocytes activate telomerase to delay senescence. Here we show that some T cells (primarily naïve and central memory cells) elongated telomeres by acquiring telomere vesicles from antigen-presenting cells (APCs) independently of telomerase action. Upon contact with these T cells, APCs degraded shelterin to donate telomeres, which were cleaved by the telomere trimming factor TZAP, and then transferred in extracellular vesicles at the immunological synapse.

T-cell trans-synaptic vesicles are distinct and carry greater effector content than constitutive extracellular vesicles.

The immunological synapse is a molecular hub that facilitates the delivery of three activation signals, namely antigen, costimulation/corepression and cytokines, from antigen-presenting cells (APC) to T cells. T cells release a fourth class of signaling entities, trans-synaptic vesicles (tSV), to mediate bidirectional communication. Here we present bead-supported lipid bilayers (BSLB) as versatile synthetic APCs to capture, characterize and advance the understanding of tSV biogenesis.

Characterization of mechanisms positioning costimulatory complexes in immune synapses.

Small immunoglobulin superfamily (sIGSF) adhesion complexes form a corolla of microdomains around an integrin ring and secretory core during immunological synapse (IS) formation. The corolla recruits and retains major costimulatory/checkpoint complexes, such as CD28, making forces that govern corolla formation of particular interest. Here, we investigated the mechanisms underlying molecular reorganization of CD2, an adhesion and costimulatory molecule of the sIGSF family during IS formation.

Increasing LFA-1 Expression Enhances Immune Synapse Architecture and T Cell Receptor Signaling in Jurkat E6.1 Cells.

The Jurkat E6.1 clone has been extensively used as a powerful tool for the genetic and biochemical dissection of the TCR signaling pathway. More recently, these cells have been exploited in imaging studies to identify key players in immunological synapse (IS) assembly in superantigen-specific conjugates and to track the dynamics of signaling molecules on glass surfaces coated with activating anti-CD3 antibodies.

A dynamic CD2-rich compartment at the outer edge of the immunological synapse boosts and integrates signals.

The CD2-CD58 recognition system promotes adhesion and signaling and counters exhaustion in human T cells. We found that CD2 localized to the outer edge of the mature immunological synapse, with cellular or artificial APC, in a pattern we refer to as a 'CD2 corolla'. The corolla captured engaged CD28, ICOS, CD226 and SLAM-F1 co-stimulators.

Composition and structure of synaptic ectosomes exporting antigen receptor linked to functional CD40 ligand from helper T cells.

Planar supported lipid bilayers (PSLB) presenting T cell receptor (TCR) ligands and ICAM-1 induce budding of extracellular microvesicles enriched in functional TCR, defined here as synaptic ectosomes (SE), from helper T cells. SE bind peptide-MHC directly exporting TCR into the synaptic cleft, but incorporation of other effectors is unknown. Here, we utilized bead supported lipid bilayers (BSLB) to capture SE from single immunological synapses (IS), determined SE composition by immunofluorescence flow cytometry and enriched SE for proteomic analysis by particle sorting.

T-derived dopamine accelerates productive synapses in germinal centres.

Protective high-affinity antibody responses depend on competitive selection of B cells carrying somatically mutated B-cell receptors by follicular helper T (T) cells in germinal centres. The rapid T-B-cell interactions that occur during this process are reminiscent of neural synaptic transmission pathways. Here we show that a proportion of human T cells contain dense-core granules marked by chromogranin B, which are normally found in neuronal presynaptic terminals storing catecholamines such as dopamine.

Membrane nanoclusters of FcγRI segregate from inhibitory SIRPα upon activation of human macrophages.

Signal integration between activating Fc receptors and inhibitory signal regulatory protein α (SIRPα) controls macrophage phagocytosis. Here, using dual-color direct stochastic optical reconstruction microscopy, we report that Fcγ receptor I (FcγRI), FcγRII, and SIRPα are not homogeneously distributed at macrophage surfaces but are organized in discrete nanoclusters, with a mean radius of 71 ± 11 nm, 60 ± 6 nm, and 48 ± 3 nm, respectively. Nanoclusters of FcγRI, but not FcγRII, are constitutively associated with nanoclusters of SIRPα, within 62 ± 5 nm, mediated by the actin cytoskeleton.

Comprehensive Analysis of Immunological Synapse Phenotypes Using Supported Lipid Bilayers.

Supported lipid bilayers (SLB) formed on glass substrates have been a useful tool for study of immune cell signaling since the early 1980s. The mobility of lipid-anchored proteins in the system, first described for antibodies binding to synthetic phospholipid head groups, allows for the measurement of two-dimensional binding reactions and signaling processes in a single imaging plane over time or for fixed samples. The fragility of SLB and the challenges of building and validating individual substrates limit most experimenters to ~10 samples per day, perhaps increasing this few-fold when examining fixed samples.

Inward-facing conformation of the zinc transporter YiiP revealed by cryoelectron microscopy.

YiiP is a dimeric Zn(2+)/H(+) antiporter from Escherichia coli belonging to the cation diffusion facilitator family. We used cryoelectron microscopy to determine a 13-Å resolution structure of a YiiP homolog from Shewanella oneidensis within a lipid bilayer in the absence of Zn(2+). Starting from the X-ray structure in the presence of Zn(2+), we used molecular dynamics flexible fitting to build a model consistent with our map.

The physical state of lipid substrates provides transacylation specificity for tafazzin.

Cardiolipin is a mitochondrial phospholipid with a characteristic acyl chain composition that depends on the function of tafazzin, a phospholipid-lysophospholipid transacylase, although the enzyme itself lacks acyl specificity. We incubated isolated tafazzin with various mixtures of phospholipids and lysophospholipids, characterized the lipid phase by (31)P-NMR and measured newly formed molecular species by MS. Substantial transacylation was observed only in nonbilayer lipid aggregates, and the substrate specificity was highly sensitive to the lipid phase.