Molecular design of the γδ T cell receptor ectodomain encodes biologically fit ligand recognition in the absence of mechanosensing

High acuity αβT cell receptor (TCR) recognition of peptides bound to MHC molecules (pMHC) requires mechanosensing, a process whereby piconewton (pN) bioforces exert physical load on αβTCR-pMHC bonds to dynamically alter their lifetimes and foster digital sensitivity cellular signaling. While mechanotransduction is operative for both αβTCRs and preTCRs within the αβT-lineage, its role in γδT cells is unknown. Here we show that the human DP10.7 γδTCR specific for the sulfoglycolipid sulfatide bound to CD1d only sustains significant load and undergoes force-induced structural transitions when the binding interface-distal γδ constant domain (C) module is replaced with that of αβ. The chimeric γδ-αβTCR also signals more robustly than the wild-type γδTCR as revealed by RNA-seq analysis of TCR-transduced Rag2-/- thymocytes, consistent with structural, single molecule and molecular dynamics studies reflective of γδTCRs as mediating recognition via a more canonical immunoglobulin-like receptor interaction. Absence of robust, force-related catch bonds as well as γδTCR structural transitions implies that γδT cells do not use mechanosensing for ligand recognition. This distinction is consonant with the fact that their innate-type ligands, including markers of cellular stress, are expressed at high copy number relative to sparse pMHC ligands of αβT cells arrayed on activating target cells. We posit that mechanosensing emerged over ~200 million years of vertebrate evolution to fulfill indispensable adaptive immune recognition requirements for pMHC in the αβT cell lineage that are unnecessary for the γδ T cell lineage mechanism of non-pMHC ligand detection. Comparison of a native γδTCR with a chimeric γδTCR where the constant domains have been replaced with those of an αβTCR, focusing on signaling properties during the DN3 and DN4 stages of thymocyte development. 2 receptors (γδTCR, γδ/αβTCR) x 2 stages (DN3, DN4) x 2 signaling environments (control, +sulfatide/CD1d agonist) x 3 replicates (except for 2 samples with 2 replicates) = 22 libraries