T cell receptor (TCR) binding to agonist peptide major histocompatibility complex (pMHC) triggers signaling events that initiate T cell responses. This system is remarkably sensitive, requiring only a few binding events to successfully activate a cellular response. On average, activating pMHC ligands exhibit mean dwell times of at least a few seconds when bound to the TCR. However, a T cell accumulates pMHC-TCR interactions as a stochastic series of discrete, single-molecule binding events whose individual dwell times are broadly distributed. With activation occurring in response to only a handful of such binding events, individual cells are unlikely to experience the average binding time. Here, we mapped the ensemble of pMHC-TCR binding events in space and time while simultaneously monitoring cellular activation. Our findings revealed that T cell activation hinges on rare, long–dwell time binding events that are an order of magnitude longer than the average agonist pMHC-TCR dwell time. Furthermore, we observed that short pMHC-TCR binding events that were spatially correlated and temporally sequential led to cellular activation. These observations indicate that T cell antigen discrimination likely occurs by sensing the tail end of the pMHC-TCR binding dwell time distribution rather than its average properties.

与激动剂肽主要组织相容性复合物（pMHC）结合的T细胞受体（TCR）触发启动T细胞反应的信号事件。该系统非常灵敏，仅需几个结合事件即可成功激活细胞应答。平均而言，激活的pMHC配体与TCR结合时，平均停留时间至少为几秒钟。但是，T细胞积累的pMHC-TCR相互作用是一系列离散的，单分子结合事件的随机序列，其单个驻留时间分布广泛。由于仅响应少数此类结合事件而发生激活，因此单个细胞不太可能经历平均结合时间。在这里，我们在空间和时间上绘制了pMHC-TCR结合事件的集合，同时监测了细胞的活化。我们的发现表明，T细胞活化取决于罕见的长停留时间结合事件，该事件比平均激动剂pMHC-TCR停留时间长一个数量级。此外，我们观察到，短的pMHC-TCR结合事件在空间上相关并且在时间上相继导致细胞激活。这些观察结果表明，通过感知pMHC-TCR结合驻留时间分布的尾端而不是其平均特性，可能会发生T细胞抗原区分。