LAUSR

Antigen-specific T-cell responses are triggered via interaction of T cell receptors (TCR) with pathogenor tumorderived peptides presented on major histocompatibility complexes (pMHC) by antigen-presenting cells (APCs). The mechanisms of TCR signaling are fundamentally important to our understanding of adaptive immunity and TCR-based therapies. This can be exemplified by the fact that studies of TCR have led to clinical development of chimeric antigen receptor T (CAR-T) cells (Srivastava and Riddell, 2015). TCR is a multiprotein complex consisting of variable TCR receptor α and β chains (TCR α/β) associated with the dimeric signaling modules CD3γ/ε, δ/ε, and ζ/ζ. Over the past two decades, many models have been proposed on the arrangement of the receptor subunits, stoichiometry of the TCR-CD3 complex and the mechanisms of TCR triggering (Rudolph et al., 2006). Our knowledge of TCR signaling, however, is far from being complete partly due to the lack of structural information of a complete TCR-CD3 complex. In one recent remarkable study published in Nature (Dong et al. 2019), an atomic-resolution view of a full TCR-CD3 complex has been revealed. The structure significantly advanced our understanding of the mechanism of TCR-CD3 assembly and offered unprecedented insight into TCR triggering. The authors reconstituted a human TCR-CD3 complex using an elegant screening system. Combination of glutaraldehyde-based cross-liking and cryo-electron microscopy (cryo-EM) allowed them to obtain a structure of the complex at 3.7 Å, revealing for the first time the molecular architecture of an intact TCR-CD3 complex at an atomicresolution. The structure showed a 1:1:1:1 stoichiometry and relative subunit positioning of TCR-CD3. The TCR α/β constant domains (TCR Cα/Cβ) and the extracellular domains (ECDs) of CD3γ/ε and CD3δ/ε’ form a trimer-like structure adjacent to the plasma membrane (PM), whereas the TCR α/ β variable domains (TCR Vα/Vβ) are positioned distal to the PM (Fig. 1A). Despite the contacts made in the ECDs, assembly of the TCR-CD3 complex is mainly mediated by its transmembrane (TM) domains and connecting peptide (CP) regions between ECDs and TMs. The two TM helices of TCR α/β are surrounded by the six TM helices of the CD3 subunits via extensive hydrophobic and ionic interactions, forming an α-helical barrel-like structure that has a major role in assembling the TCR-CD3 complex. Formation of the barrel-like structure agrees with the data showing a compact assembly of the TM domains of TCR-CD3 (Krshnan et al., 2016). Interactions involving the CP regions further fortify assembly of the complex. By contrast, the intracellular tails of the CD3 subunits are unstructured, consistent with previous NMR study (Xu et al., 2008). Several models hypothesize that pMHC or antibody binding to TCR α/β allosterically induces conformational changes in the CD3 subunits, thus exposing their intracellular signaling tails for phosphorylation by the Lck kinase and initiating signaling (Schamel et al., 2019). Unexpectedly, however, structural comparison indicated that pMHC binding induces no substantial conformational changes in the TCRCD3 complex (Fig. 1B). Similar results were also obtained from structural studies of the ECDs of TCR α/β (Baker et al., 2000; Yin et al. 2012). Furthermore, the OKT3 antibody and pMHC are differently positioned for interaction with the TCRCD3 complex (Fig. 1B and 1C), though they activate the same TCR pathways. As noted by the authors, the possibility still remains that ligand-induced oligomerization or clustering for TCR triggering. The ice cream-like structure, however, seems incompatible with TM domain-mediated oligomerization of the TCR-CD3 complex, although TM-mediated dimerization of two tilted (with respect to the PM) TCR-CD3 molecules is possible. It might be that oligomerization mediated by the ECDs further triggers conformational changes that are transmitted into the intracellular signaling domains of CD3. But TCR-CD3 oligomerization appears dispensable for TCR triggering, because monomeric agonist pMHCs anchored to a surface are sufficient to induce TCR activation (Ma et al., 2008). Numerous studies support conformational changes in the ECDs, TMs and the intracellular tails of TCR-CD3 during activation (Schamel et al., 2019). Then how does the cryoEM structure fit with these data? As noted by the authors, the