Functional Nanoscale Organization of Signaling Molecules Downstream of the T Cell Antigen Receptor

SummaryReceptor-regulated cellular signaling often is mediated by formation of transient, heterogeneous protein complexes of undefined structure. We used single and two-color photoactivated localization microscopy to study complexes downstream of the T cell antigen receptor (TCR) in single-molecule detail at the plasma membrane of intact T cells. The kinase ZAP-70 distributed completely with the TCRζ chain and both partially mixed with the adaptor LAT in activated cells, thus showing localized activation of LAT by TCR-coupled ZAP-70. In resting and activated cells, LAT primarily resided in nanoscale clusters as small as dimers whose formation depended on protein-protein and protein-lipid interactions. Surprisingly, the adaptor SLP-76 localized to the periphery of LAT clusters. This nanoscale structure depended on polymerized actin and its disruption affected TCR-dependent cell function. These results extend our understanding of the mechanism of T cell activation and the formation and organization of TCR-mediated signaling complexes, findings also relevant to other receptor systems.

► Two-color PALM reveals nanoscale organization of T cell receptor signaling complexes ► Activation occurs in LAT nanoclusters formed by protein and lipid interactions ► LAT nanoclusters get activated by mixing at discrete sites with TCR and ZAP-70 ► LAT nanoclusters demonstrate functional structure with SLP-76 at their periphery