Copper is an essential nutrient for sustaining life, and emerging data have expanded the roles of this metal in biology from its canonical functions as a static enzyme cofactor to dynamic functions as a transition metal signal. At the same time, loosely bound, labile copper pools can trigger oxidative stress and damaging events that are detrimental if misregulated. The signal/stress dichotomy of copper motivates the development of new chemical tools to study its spatial and temporal distributions in native biological contexts such as living cells. Here, we report a family of fluorescent copper sensors built upon carbon-, silicon-, and phosphorus-substituted rhodol dyes that enable systematic tuning of excitation/emission colors from orange to near-infrared. These probes can detect changes in labile copper levels in living cells upon copper supplementation and/or depletion. We demonstrate the ability of the carbon–rhodol based congener, Copper Carbo Fluor 1 (CCF1), to identify elevations in labile copper pools in the Atp7a^–/– fibroblast cell model of the genetic copper disorder Menkes disease. Moreover, we showcase the utility of the red-emitting phosphorus-rhodol based dye Copper Phosphorus Fluor 1 (CPF1) in dual-color, dual-analyte imaging experiments with the green-emitting calcium indicator Calcium Green-1 to enable simultaneous detection of fluctuations in copper and calcium pools in living cells. The results provide a starting point for advancing tools to study the contributions of copper to health and disease and for exploiting the rapidly growing palette of heteroatom-substituted xanthene dyes to rationally tune the optical properties of fluorescent indicators for other biologically important analytes.

中文翻译：

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通过 Rhodol 核心的系统杂原子取代来调整荧光铜传感器的调色板

铜是维持生命的必需营养素，新出现的数据扩展了这种金属在生物学中的作用，从其作为静态酶辅因子的典型功能扩展到作为过渡金属信号的动态功能。与此同时，松散结合、不稳定的铜池可能会引发氧化应激和破坏性事件，如果调节不当，这些事件将是有害的。铜的信号/应力二分法促进了新化学工具的开发，以研究其在活细胞等天然生物环境中的空间和时间分布。在这里，我们报告了一系列基于碳、硅和磷取代的对甲酚染料构建的荧光铜传感器，可以系统地调节从橙色到近红外的激发/发射颜色。这些探针可以检测补充和/或耗尽铜后活细胞中不稳定铜水平的变化。我们证明了基于碳-对甲酚的同源物铜碳氟化合物 1 (CCF1) 能够识别遗传性铜紊乱门克斯病的Atp7a –/– 成纤维^{细胞模型中不稳定铜池的升高。}此外，我们还展示了发红光磷对甲酚基染料铜磷荧光 1 (CPF1) 在双色、双分析物成像实验中与发绿光钙指示剂 Calcium Green-1 的实用性，以实现同时检测波动存在于活细胞的铜和钙池中。这些结果为先进工具提供了一个起点，以研究铜对健康和疾病的贡献，并利用快速增长的杂原子取代呫吨染料调色板来合理调整其他重要生物学分析物的荧光指示剂的光学特性。