T cell activation relies on the recognition, by the T cell receptor (TCR), of an antigenic peptide displayed at the surface of an antigen-presenting cell in association with major histocompatibility complex molecules (pMHC). This encounter triggers the formation of a dynamic contact between the two cells, the immunological synapse, an area where TCR engagement initiates a cascade of phosphorylation and dephosphorylation events affecting proteins and lipids. It is this cascade that ultimately shapes the lymphocyte's response. The TCR is a multimeric receptor composed of the clonotypic α and β chains, which are responsible for antigen binding, and the CD3 chains (γ, ε, δ and ζ), which carry out signal transduction. Its signaling must be tightly regulated, as it governs the establishment of both tolerance and the immune response.
This review starts from an often-overlooked observation: although pMHC recognition does indeed occur at the plasma membrane, several key signaling players — the kinase Lck, the CD3ζ chain and the adaptor protein LAT — are not confined to this membrane. They also reside, sometimes in greater abundance, in intracellular membrane compartments of both endocytic and exocytic nature. Upon TCR binding to pMHC, these intracellular pools of signaling molecules are rapidly polarized toward the immunological synapse, and the strict regulation of their trafficking contributes to lymphocyte activation. The authors summarize the current state of knowledge regarding the nature of these endosomes, their origin and the mechanisms that govern their mobility toward the synapse.
The analysis draws in particular on approaches developed by the teams to track LAT and CD3ζ. An "average cell" method, applied to conjugates of Jurkat T cells and Raji B cells, made it possible to visualize the early polarization of an intracellular pool of LAT toward the synapse followed by its progressive enrichment. Surface-labeled chimeric LAT constructs revealed its internalization into endosomes recruited to the synapse, while a SNAP-tag capture assay revealed retrograde transport of LAT toward the Golgi apparatus. In addition, the use of a fluorescence resonance energy transfer reporter (FRET-FLIM) showed that intracellular pools of CD3ζ interact with the kinase ZAP70, and membrane fractionation documented the distribution of the phosphorylated form of CD3ζ following activation.
Based on these observations, the authors put forward several hypotheses regarding the functional role(s) that these intracellular pools might play in lymphocyte activation, and discuss the tools that could test them. They emphasize, however, that how these signaling-molecule-bearing endosomes actually regulate T cell activation remains unknown.