Microclusters as T Cell Signaling Hubs: Structure, Kinetics, and Regulation.

Lakshmi Balagopalan,Kumarkrishna Raychaudhuri,Lawrence E Samelson

doi:10.3389/fcell.2020.608530

Abstract

When T cell receptors (TCRs) engage with stimulatory ligands, one of the first microscopically visible events is the formation of microclusters at the site of T cell activation. Since the discovery of these structures almost 20 years ago, they have been studied extensively in live cells using confocal and total internal reflection fluorescence (TIRF) microscopy. However, due to limits in image resolution and acquisition speed, the spatial relationships of signaling components within microclusters, the kinetics of their assembly and disassembly, and the role of vesicular trafficking in microcluster formation and maintenance were not finely characterized. In this review, we will summarize how new microscopy techniques have revealed novel insights into the assembly of these structures. The sub-diffraction organization of microclusters as well as the finely dissected kinetics of recruitment and disassociation of molecules from microclusters will be discussed. The role of cell surface molecules in microcluster formation and the kinetics of molecular recruitment via intracellular vesicular trafficking to microclusters is described. Finally, the role of post-translational modifications such as ubiquitination in the downregulation of cell surface signaling molecules is also discussed. These results will be related to the role of these structures and processes in T cell activation.

Highlights

The central event in the initiation of the adaptive immune response to foreign antigen is the interaction of the T cell antigen receptor (TCR) with an antigenic peptide presented by a protein encoded by the major histocompatibility complex
Live-cell total internal reflection fluorescence (TIRF)-SIM and TIRF microscopy approaches showed that, following TCR engagement, ZAP-70 was first recruited to TCR microclusters, followed by simultaneous recruitment of signaling and adaptor domain proteins (LAT, SLP-76, GRB2, ADAP, VAV1, NCK, and PLCγ)
We have shown previously that the endocytosis of microclusters containing LAT and SLP-76 is regulated by c-Cbl mediated ubiquitination, and inhibition of c-Cbl function increases microcluster lifetime (Barr et al, 2006; Balagopalan et al, 2007)

Summary

Introduction

The central event in the initiation of the adaptive immune response to foreign antigen is the interaction of the T cell antigen receptor (TCR) with an antigenic peptide presented by a protein encoded by the major histocompatibility complex (pMHC). Advanced imaging techniques such as lattice light sheet microscopy (LLSM; see Box 1 in Supplementary Material) have enabled the visualization of microclusters at the initiation of T cell contact, confirming the role of these structures as signaling units that drive T cell activation (Ritter et al, 2015).

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Conclusion