Tyrosine kinase A (TrkA) is a membrane receptor which, upon ligand binding, activates several pathways including MAPK/ERK signaling, implicated in a spectrum of human pathologies; thus, TrkA is an emerging therapeutic target in treatment of neuronal diseases and cancer. However, mechanistic insights into TrKA signaling are lacking due to lack of site-dependent phosphorylation control. Here we engineer two light-sensitive tyrosine analogues, namely p-azido-L-phenylalanine (AzF) and the caged-tyrosine (ONB), through amber codon suppression to optically manipulate the phosphorylation state of individual intracellular tyrosines in TrkA. We identify TrkA-AzF and ONB mutants, which can activate the ERK pathway in the absence of NGF ligand binding through light control. Our results not only reveal how TrkA site-dependent phosphorylation controls the defined signaling process, but also extend the genetic code expansion technology to enable regulation of receptor-type kinase activation by optical control at the precision of a single phosphorylation site. It paves the way for comprehensive analysis of kinase-associated pathways as well as screening of compounds intervening in a site-directed phosphorylation pathway for targeted therapy.

酪氨酸激酶A（TrkA）是一种膜受体，在配体结合后会激活多种途径，包括MAPK / ERK信号传导，涉及多种人类病理学。因此，TrkA是治疗神经元疾病和癌症的新兴治疗靶标。但是，由于缺乏位点依赖性的磷酸化控制，因此缺乏对TrKA信号传导的机械性见解。在这里，我们设计了两个光敏酪氨酸类似物，即p-叠氮基-L-苯丙氨酸（AzF）和笼状酪氨酸（ONB），通过琥珀色密码子抑制来光学操纵TrkA中单个细胞内酪氨酸的磷酸化状态。我们确定了TrkA-AzF和ONB突变体，它们可以在没有NGF配体通过光控制结合的情况下激活ERK途径。我们的研究结果不仅揭示了TrkA位点依赖性磷酸化如何控制所定义的信号传导过程，而且扩展了遗传密码扩展技术，从而能够通过光学控制在单个磷酸化位点的精度上调节受体型激酶的活化。它为全面分析激酶相关途径，以及筛选干预靶向治疗的定点磷酸化途径的化合物铺平了道路。