Initiation of T cell antigen receptor (TCR) signaling involves phosphorylation of CD3 cytoplasmic tails by the tyrosine kinase Lck. How Lck is recruited to the TCR to initiate signaling is not well known. We report a previously unknown binding motif in the CD3ε cytoplasmic tail that interacts in a noncanonical mode with the Lck SH3 domain: the receptor kinase (RK) motif. The RK motif is accessible only upon TCR ligation, demonstrating how ligand binding leads to Lck recruitment. Binding of the Lck SH3 domain to the exposed RK motif resulted in local augmentation of Lck activity, CD3 phosphorylation, T cell activation and thymocyte development. Introducing the RK motif into a well-characterized 41BB-based chimeric antigen receptor enhanced its antitumor function in vitro and in vivo. Our findings underscore how a better understanding of the functioning of the TCR might promote rational improvement of chimeric antigen receptor design for the treatment of cancer.

T细胞抗原受体（TCR）信号的启动涉及酪氨酸激酶Lck对CD3细胞质尾部的磷酸化作用。Lck如何被招募到TCR来启动信号传输尚不清楚。我们报告了以前未知的结合模体在CD3ε细胞质尾巴中与Lck SH3域以非经典模式相互作用：受体激酶（RK）模体。RK基序只有在TCR连接时才能使用，这说明配体结合如何导致Lck募集。Lck SH3结构域与暴露的RK基序的结合导致Lck活性，CD3磷酸化，T细胞活化和胸腺细胞发育的局部增强。将RK基序引入已充分表征的基于41BB的嵌合抗原受体可增强其在体内和体外的抗肿瘤功能。