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The T cell receptor (TCR) initiates the elimination of pathogens and tumors by T cells. To avoid damage to the host, the receptor must be capable of discriminating between wild-type and mutated self and nonself peptide ligands presented by host cells. Exactly how the TCR does this is unknown. In resting T cells, the TCR is largely unphosphorylated due to the dominance of phosphatases over the kinases expressed at the cell surface. However, when agonist peptides are presented to the TCR by major histocompatibility complex proteins expressed by antigen-presenting cells (APCs), very fast receptor triggering, i.e., TCR phosphorylation, occurs. Recent work suggests that this depends on the local exclusion of the phosphatases from regions of contact of the T cells with the APCs. Here, we developed and tested a quantitative treatment of receptor triggering reliant only on TCR dwell time in phosphatase-depleted cell contacts constrained in area by cell topography. Using the model and experimentally derived parameters, we found that ligand discrimination likely depends crucially on individual contacts being ∼200 nm in radius, matching the dimensions of the surface protrusions used by T cells to interrogate their targets. The model not only correctly predicted the relative signaling potencies of known agonists and nonagonists but also achieved this in the absence of kinetic proofreading. Our work provides a simple, quantitative, and predictive molecular framework for understanding why TCR triggering is so selective and fast and reveals that, for some receptors, cell topography likely influences signaling outcomes., Competing Interests: The authors declare no conflict of interest., (Copyright © 2019 the Author(s). Published by PNAS.)

The αβ T-cell coreceptor CD4 enhances immune responses more than 1 million-fold in some assays, and yet the affinity of CD4 for its ligand, peptide-major histocompatibility class II (pMHC II) on antigen-presenting cells, is so weak that it was previously unquantifiable. Here, we report that a soluble form of CD4 failed to bind detectably to pMHC II in surface plasmon resonance-based assays, establishing a new upper limit for the solution affinity at 2.5 mM. However, when presented multivalently on magnetic beads, soluble CD4 bound pMHC II-expressing B cells, confirming that it is active and allowing mapping of the native coreceptor binding site on pMHC II. Whereas binding was undetectable in solution, the affinity of the CD4/pMHC II interaction could be measured in 2D using CD4- and adhesion molecule-functionalized, supported lipid bilayers, yielding a 2D Kd of ∼5,000 molecules/μm(2) This value is two to three orders of magnitude higher than previously measured 2D Kd values for interacting leukocyte surface proteins. Calculations indicated, however, that CD4/pMHC II binding would increase rates of T-cell receptor (TCR) complex phosphorylation by threefold via the recruitment of Lck, with only a small, 2-20% increase in the effective affinity of the TCR for pMHC II. The affinity of CD4/pMHC II therefore seems to be set at a value that increases T-cell sensitivity by enhancing phosphorylation, without compromising ligand discrimination.

We have previously shown that lipoprotein(a) [Lp(a)], an atherogenic lipoprotein that contains apolipoprotein(a), which shares partial structural homology to plasminogen, binds to a plasmin-modified fibrin surface, and we have postulated that this interaction may be atherogenic. Moderate elevations in blood homocysteine, a relatively common condition, predispose to premature atherosclerosis. The reasons for this are not established. We now report that homocysteine, at concentrations as low as 8 microM, significantly increases the affinity of Lp(a) for fibrin. Homocysteine induces a 20-fold increase in the affinity between Lp(a) and plasmin-treated fibrin and a 4-fold increase with unmodified fibrin. Lp(a) binding is inhibited by epsilon-aminocaproic acid, indicating lysine binding site specificity. Homocysteine does not enhance the binding of Lp(a) to other surface-bound proteins. Cysteine, glutathione, and N-acetylcysteine also increase the affinity between Lp(a) and fibrin. Homocysteine does not affect the binding of low density lipoprotein or plasminogen to fibrin, nor does it alter the gel-filtration elution pattern of Lp(a). Immunoblot analysis documents the fact that homocysteine partially reduces Lp(a). These results suggest that homocysteine alters the intact Lp(a) particle so as to increase the reactivity of the plasminogen-like apolipoprotein(a) portion of the molecule. The observation that sulfhydryl amino acids increase Lp(a) binding to fibrin suggests a biochemical relationship between sulfhydryl compound metabolism, thrombosis, and atherogenesis.