Abstract
T cells can recognize a few molecules of cognate antigen amongst vastly outnumbering non-cognate ligands. The T cell receptor (TCR) differentiates antigens based on antigen-TCR binding dwell time through a kinetic proofreading process. Historically, this has been modeled as the ligated receptor undergoing a series of reactions before producing a signal. In such a sequential mechanism, the number of steps is a key determinant of discrimination fidelity. Here, we consider two features of the molecular mechanism of TCR activation that diverge from a sequential process and suggest that an alternative kinetic proofreading mechanism may be at play. First, activation processes of multiple ITAM domains of the TCR represent parallel reaction sequences taking place on a single TCR molecule. Second, the states of the parallel proofreading reactions are integrated to produce a binary output from each TCR in the form of a discrete LAT condensation event, which may or may not occur. We examine a revised kinetic proofreading scheme based on parallel reactions followed by an integration step (multi-thread scheme) and compare its performance with the sequential scheme in a stochastic setting. A distinct difference in a multi-thread scheme is that multiplicity of the parallel reaction threads provides an additional means to increase discrimination fidelity. This relieves the need for fine-tuned kinetics among chemically distinct reaction steps, which is a major hurdle for physical implementation of a sequential mechanism. Lastly, we reinterpret previously reported experimental observations and find that various proofreading behaviors are well described as proofreading through parallel reaction threads. SIGNIFICANCE STATEMENT: The kinetic proofreading mechanism by which T cells discriminate antigen has attracted interest from both physical and immunological perspectives. The proofreading process has generally been modeled as a multi-step sequence of reactions, and many experimental observations have been interpreted based on this presumed mechanism. However, these models do not capture key molecular features of the TCR signaling mechanism, including ITAM multiplicity and LAT condensation. Here, we find distinct consequences of these molecular features by examining kinetic models and reinterpreting published experimental data. ITAM multiplicity offers multiple parallel reaction threads, re-integrated by the LAT condensation step, which readily improves discrimination fidelity up to the observed levels. Multi-thread reactions are suggested to play a central role in amplifying TCR kinetic proofreading performance.