Light-based tuning of ligand half-life supports kinetic proofreading model of T cell signaling.

Doug K Tischer,Orion David Weiner

doi:10.7554/elife.42498

Abstract

T cells are thought to discriminate self from foreign peptides by converting small differences in ligand binding half-life into large changes in cell signaling. Such a kinetic proofreading model has been difficult to test directly, as existing methods of altering ligand binding half-life also change other potentially important biophysical parameters, most notably the mechanical stability of the receptor-ligand interaction. Here we develop an optogenetic approach to specifically tune the binding half-life of a chimeric antigen receptor without changing other binding parameters and provide direct evidence of kinetic proofreading in T cell signaling. This half-life discrimination is executed in the proximal signaling pathway, downstream of ZAP70 recruitment and upstream of diacylglycerol accumulation. Our methods represent a general tool for temporal and spatial control of T cell signaling and extend the reach of optogenetics to probe pathways where the individual molecular kinetics, rather than the ensemble average, gates downstream signaling.

Highlights

The T cell response begins when the T cell antigen receptor (TCR) binds its cognate peptide-major histocompatibility complex on an antigen-presenting cell
T cells likely use kinetic proofreading in combination with other mechanisms, such as co-receptor involvement (Irvine et al, 2002), mechanical forces (Kim et al, 2009; Feng et al, 2017; Liu et al, 2014), or signal amplification downstream of DAG (Das et al, 2009), to fully discriminate self from non-self
What biochemical steps underlie the observed kinetic proofreading? We addressed this question by measuring one of the earliest signaling events: the recruitment of ZAP70 to the chimeric antigen receptor (CAR)

Summary

Introduction

The T cell response begins when the T cell antigen receptor (TCR) binds its cognate peptide-major histocompatibility complex (pMHC) on an antigen-presenting cell. A major model of T cell ligand discrimination is kinetic proofreading (McKeithan, 1995), which predicts that small differences in ligand binding half-life can be amplified into large differences in signaling Such a model is attractive because it allows a few receptors bound to long-lived ligands to Tischer and Weiner. By directly controlling ligand binding half-life with light and holding all other binding parameters constant, we show that longer binding lifetimes are a key parameter for potent T cell signaling. This discrimination occurs in the proximal signaling pathway, downstream of ZAP70 recruitment and upstream of DAG accumulation. This work aids our understanding of how T cell discriminate ligands and expands optogenetics as a tool for controlling the timing of single molecular interactions

Results

Discussion

Materials and methods

Funding Funder Genentech Foundation