Direct Activation of ATM by Resveratrol under Oxidizing Conditions

Resveratrol has been widely reported to reduce cancer progression in model systems and to selectively induce cell death in transformed cell lines. Many enzymes have been reported to respond to resveratrol in mammalian cells, including the Ataxia-Telangiectasia Mutated (ATM) protein kinase that acts in DNA damage recognition, signaling, and repair. Here we investigate the responses of ATM to resveratrol exposure in normal and transformed human cell lines and find that ATM autophosphorylation and substrate phosphorylation is stimulated by resveratrol in a manner that is promoted by reactive oxygen species (ROS). We observe direct stimulatory effects of resveratrol on purified ATM in vitro and find that the catalytic efficiency of the kinase on a model substrate is increased by resveratrol. In the purified system we also observe a requirement for oxidation, as the effect of resveratrol on ATM signaling is substantially reduced by agents that prevent disulfide bond formation in ATM. These results demonstrate that resveratrol effects on ATM are direct, and suggest a mechanism by which the oxidizing environment of transformed cells promotes ATM activity and blocks cell proliferation.

白藜芦醇（Resveratrol）已被广泛报道可在模型系统中延缓癌症进展，并选择性诱导转化细胞系发生细胞死亡。已有研究表明，哺乳动物细胞内多种酶可响应白藜芦醇，其中包括参与DNA损伤识别、信号转导与修复的共济失调毛细血管扩张症突变（Ataxia-Telangiectasia Mutated, ATM）蛋白激酶。本研究针对正常及转化人类细胞系中ATM对白藜芦醇暴露的响应展开探究，发现白藜芦醇可通过活性氧（Reactive Oxygen Species, ROS）介导的方式，刺激ATM的自磷酸化及其底物磷酸化过程。我们在体外实验中观察到白藜芦醇对纯化ATM的直接激活作用，且发现该物质可提升激酶对模型底物的催化效率。在纯化蛋白体系中，我们还观察到氧化修饰的必要性：当使用可阻断ATM分子内二硫键形成的试剂时，白藜芦醇对ATM信号通路的调控作用会显著减弱。上述实验结果证实，白藜芦醇对ATM的调控作用是直接的，并提示转化细胞内的氧化环境可通过该机制促进ATM活性，进而抑制细胞增殖。