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TCRα transmembrane domain coordinates antigen triggering by regulating bilayer immersion and CD3 associations

ABSTRACT: Initial molecular details of cellular activation following αβT-cell receptor (TCR) ligation by pMHC remain unexplored. We determined the NMR structure of the TCRα subunit transmembrane (TM) segment revealing a bipartite helix whose segmentation fostersdynamic movement. Positively charged TM residues Arg251 and Lys256 project from opposite faces of the helix, with Lys256 controlling immersion depth. Their modification causes step-wise reduction in associations with T-cell surface CD3ζζ and CD3εγ/CD3εδ, respectively, leading to an activated transcriptome. Optical tweezers reveal that Arg251 and Lys256 mutations alter αβTCR-pMHC bond lifetimes, while mutations within interacting TCRα connecting peptide and CD3δ CxxC motif juxtamembrane elements selectively attenuate signal transduction. Our findings suggest that mechanical forces applied during pMHC ligation initiate T-cell activation by altering the disposition of those basic sidechains to rearrange TCR complex membrane topology and weaken TCRαβ and CD3 associations. This αβTCR dissociative mechanism impacts future immunotherapy and synthetic receptor design on CAR-T cells.

ORGANISM(S): Mus musculus

PROVIDER: GSE106760 | GEO | 2017/12/01

SECONDARY ACCESSION(S): PRJNA417864

REPOSITORIES: GEO

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Project description:Developmental checkpoints in stem/progenitor cells are critical to the determination, commitment and differentiation into distinct lineages. Cancer cells often retain expression of lineage-specific checkpoint proteins, but their potential impact in cancer remains elusive. T lymphocytes mature in the thymus following a highly orchestrated developmental process that entails the successive rearrangements and expression of T-cell receptor (TCR) genes. Low affinity recognition of self-peptide/MHC complexes (self-pMHC) presented by thymic epithelial cells by the TCR of CD4+CD8+ (DP) cortical thymocytes transduces positive selection signals that ultimately shape the developing T cell repertoire. DP thymocytes not receiving these signals die by lack of stimulation whereas those that recognize self-pMHC with high affinity undergo TCR-mediated apoptosis and negative selection. In T-cell acute lymphoblastic leukaemia (T-ALL), leukaemic transformation of maturating thymocytes results from the acquisition of multiple genetic and epigenetic alterations in oncogenes and tumour suppressor genes, that disrupt the normal regulatory circuits and drive clonal expansion of differentiation-arrested lymphoblasts. We show here that TCR triggering by negatively-selecting self-pMHC prevented T-ALL development and leukaemia maintenance in mice. Induction of TCR signalling by high affinity self-pMHC or treatment with monoclonal antibodies to the CD3 signalling chain (anti-CD3) caused massive leukaemic cell death and a gene expression program resembling that of thymocyte negative selection. Importantly, anti-CD3 treatment hampered leukaemogenesis in mice transplanted with either mouse or patient-derived T-ALLs. These data provide a rationale for targeted therapy based on anti-CD3 treatment of T-ALL patients and demonstrate that endogenous developmental checkpoint proteins are amenable to therapeutic intervention in cancer cells. Gene expression data from four culture conditions was performed for the following cells: ALL-SIL-TCRÎ±/Î²-GFP co-cultured on OP9-DL1 for 48 h, ALL-SIL-TCRÎ±/Î²-GFP without co-culture, ALL-SIL cells transduced with TLX shRNA (sh-TLX), and ALL-SIL transduced with sh-control vectors. All conditions are performed in two replicates.

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TCRα transmembrane domain coordinates antigen triggering by regulating bilayer immersion and CD3 associations

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