During the last funding interval as a merit award, this grant focused on the T cell receptor (TCR) complex composed of an ?? heterodimer and non-covalently associated CD3 signaling components. We continued crystallographic analysis with TCRs, providing the first structure of an ?? heterodimer in complex with a peptide/MHC (pMHC) class II ligand. In conjunction with a crystallographic structure of CD4 D1D2 binding to the same MHC class II molecule, a hydrophobic concavity formed by residues from membrane proximal ?2 and ?2 MHC class II domains was revealed. We excluded a direct TCRab-CD4 interaction, instead revealing how TCR?? and CD4 signaling is coordinated in a """"""""V-shape"""""""" around the antigenic peptide/pMHC class II complex. Solution structures of CD3?? and CD3?? ectodomain complexes were determined by NMR, uncovering for each dimer a unique side-to-side hydrophobic interface between their two Ig-like domains with parallel pairing of respective C-terminal ?-strands, and suggesting how rigidified CD3 elements participate in TCR-based signal transduction. To now characterize the structural basis for early signal transduction events via this TCR?? complex, four aims are proposed involving the CD3?? heterodimer and associated TCR ? chain. First, NMR-based methods will be used to determine the precise binding site on CD3?? of activating and non-activating anti-CD3? mAbs, given equivalent binding affinities. Second, CD3??-associated TCR? chain recognition function during thymic development and T cell activation accounting for V? repertoire bias will be assessed. Third, the structure and function of the transmembrane (TM) segments of CD3? and ? will be determined and their interaction with that of TCR ? ascertained. Fourth, electron paramagnetic resonance (EPR) methods will be exploited to define the orientation and disposition of TM elements and how pMHC or anti-CD3? mAbs affect TM depth, orientation and structural conformation. Distance measurements between the ? chain and CD3? and CD3? ectodomains will also be examined before and after pMHC or anti- CD3 mAb ligation. That orthological rather than vertical force via CD3 components activates T cells suggests a dynamic mechanosensor TCR model;pMHC pulls on the TCR from the opposing antigen-presenting cell surface such that the ?? heterodimer then presses on the CD3 ectodomains to initiate signaling.
The TCR is responsible for mediating antigen-specific T cell stimulation. As such, insights from this proposal will be relevant for design of inhibitory and activating anti-TCR monoclonal antibodies and other molecules to treat human autoimmune and immunodeficiency states, respectively.
|Mallis, Robert J; Arthanari, Haribabu; Lang, Matthew J et al. (2018) NMR-directed design of pre-TCR? and pMHC molecules implies a distinct geometry for pre-TCR relative to ??TCR recognition of pMHC. J Biol Chem 293:754-766|
|Mallis, Robert J; Reinherz, Ellis L; Wagner, Gerhard et al. (2016) Backbone resonance assignment of N15, N30 and D10 T cell receptor ? subunits. Biomol NMR Assign 10:35-9|
|Reinherz, Ellis L; Wang, Jia-huai (2015) Codification of bidentate pMHC interaction with TCR and its co-receptor. Trends Immunol 36:300-6|
|Mallis, Robert J; Bai, Ke; Arthanari, Haribabu et al. (2015) Pre-TCR ligand binding impacts thymocyte development before ??TCR expression. Proc Natl Acad Sci U S A 112:8373-8|
|Choi, Young I; Duke-Cohan, Jonathan S; Chen, Wei et al. (2014) Dynamic control of ?1 integrin adhesion by the plexinD1-sema3E axis. Proc Natl Acad Sci U S A 111:379-84|
|Choi, Young I; Duke-Cohan, Jonathan S; Tan, Jing et al. (2013) Plxnd1 expression in thymocytes regulates their intrathymic migration while that in thymic endothelium impacts medullary topology. Front Immunol 4:392|
|Touma, Maki; Keskin, Derin B; Shiroki, Fumiko et al. (2011) Impaired B cell development and function in the absence of IkappaBNS. J Immunol 187:3942-52|
|Kim, Sun Taek; Touma, Maki; Takeuchi, Koh et al. (2010) Distinctive CD3 heterodimeric ectodomain topologies maximize antigen-triggered activation of alpha beta T cell receptors. J Immunol 185:2951-9|
|Vernier, Gregory; Wang, Jie; Jennings, Laura D et al. (2009) Solubilization and characterization of the anthrax toxin pore in detergent micelles. Protein Sci 18:1882-95|
|Kim, Sun Taek; Takeuchi, Koh; Sun, Zhen-Yu J et al. (2009) The alphabeta T cell receptor is an anisotropic mechanosensor. J Biol Chem 284:31028-37|
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