Nano-clustering of ligands on surrogate antigen presenting cells modulates T cell membrane adhesion and organization

Abstract

We investigate the adhesion and molecular organization of the plasma membrane of T lymphocytes interacting with a surrogate antigen presenting cell comprising glass supported ordered arrays of antibody (α-CD3) nano-dots dispersed in a non-adhesive matrix of polyethylene glycol (PEG). The local membrane adhesion and topography, as well as the distribution of the T cell receptors (TCRs) and the kinase ZAP-70, are influenced by dot-geometry, whereas the cell spreading area is determined by the overall average density of the ligands rather than specific characteristics of the dots. TCR clusters are recruited preferentially to the nano-dots and the TCR cluster size distribution has a weak dot-size dependence. On the patterns, the clusters are larger, more numerous, and more enriched in TCRs, as compared to the homogeneously distributed ligands at comparable concentrations. These observations support the idea that non-ligated TCRs residing in the non-adhered parts of the proximal membrane are able to diffuse and enrich the existing clusters at the ligand dots. However, long distance transport is impaired and cluster centralization in the form of a central supramolecular cluster (cSMAC) is not observed. Time-lapse imaging of early cell-surface contacts indicates that the ZAP-70 microclusters are directly recruited to the site of the antibody dots and this process is concomitant with membrane adhesion. These results together point to a complex interplay of adhesion, molecular organization and activation in response to spatially modulated stimulation.

Biphasic mechanosensitivity of T cell receptor-mediated spreading of lymphocytes

“…Here we explore mechanosensing in T cells via the TCR-CD3 complex, in presence or absence of ICAM-1-a ligand for the T cell integrin LFA-1. The ligand of choice is anti-CD3 which provides sufficient adhesion via the TCR-CD3 complex alone (17,20)-something not possible with the weaker pMHC/TCR bond, at the same time eliciting the same signaling pathways as pMHC ligation. We follow the spreading of Jurkat T cells on functionalized surfaces, spreading being a marker of future proliferation for T lymphocytes (21).…”

Nanobody-antigen catch-bond reveals NK cell mechanosensitivity

“…The T-cell and NK cell synapses exhibit strong ressemblance including the role of integrins (41), actin organisation and depletion for cytotoxic vesicle release (42), actin retrograde flow (6). Based on literature and our own experience with T-cells (32,43), we hypothesize that the NK cell synapse is also exercing and sensing force. Our cellular experiments show that NK cells engage an immune synapse on anti-CD16 coated surfaces, for sufficiently high densities of antibodies.…”

Interferometric Scattering (iSCAT) Microscopy and Related Techniques

“…In the decade that followed right up to the present day we see a broad array of research efforts, drawn from numerous communities, exploring interferometric detection of single nano-objects such as viruses, DNA, microtubules, exosomes, and proteins [75][76][77][78][79][80][81][82][83][84][85][86][87][88][89][90][91]. Interestingly, interferometric microscopies are also flourishing in the general context of label-free imaging of cells and membranes even if nanoparticles are not at the center of attention [75,76,90,[92][93][94][95][96][97][98][99][100][101][102][103][104][105][106]. The underlying physics of these methods remains the same although a plethora of acronyms such as interference reflectance imaging sensing (IRIS) [77], rotating coherent scattering (ROCS) [98], interference plasmonic imaging (iPM) [88], coherent bright-field imaging (COBRI) [107], stroboscopic interference scattering imaging (stroboSCAT) [108], interferometric scattering mass spectrometry (iSCAMS) [109] are on the rise.…”