Long Marcus J C, Urul Daniel A, Chawla Shivansh, Lin Hong-Yu, Zhao Yi, Haegele Joseph A, Wang Yiran, Aye Yimon
Department of Chemistry and Chemical Biology, Cornell University , Ithaca, New York 14853, United States.
Department of Biochemistry, Weill Cornell Medicine , New York, New York 10065, United States.
Biochemistry. 2018 Jan 16;57(2):216-220. doi: 10.1021/acs.biochem.7b00642. Epub 2017 Sep 12.
Adduction of an electrophile to privileged sensor proteins and the resulting phenotypically dominant responses are increasingly appreciated as being essential for metazoan health. Functional similarities between the biological electrophiles and electrophilic pharmacophores commonly found in covalent drugs further fortify the translational relevance of these small-molecule signals. Genetically encodable or small-molecule-based fluorescent reporters and redox proteomics have revolutionized the observation and profiling of cellular redox states and electrophile-sensor proteins, respectively. However, precision mapping between specific redox-modified targets and specific responses has only recently begun to be addressed, and systems tractable to both genetic manipulation and on-target redox signaling in vivo remain largely limited. Here we engineer transgenic Caenorhabditis elegans expressing functional HaloTagged fusion proteins and use this system to develop a generalizable light-controlled approach to tagging a prototypical electrophile-sensor protein with native electrophiles in vivo. The method circumvents issues associated with low uptake/distribution and toxicity/promiscuity. Given the validated success of C. elegans in aging studies, this optimized platform offers a new lens with which to scrutinize how on-target electrophile signaling influences redox-dependent life span regulation.
亲电试剂与特殊传感器蛋白的结合以及由此产生的表型显性反应，越来越被认为对后生动物的健康至关重要。生物亲电试剂与共价药物中常见的亲电药效基团之间的功能相似性，进一步加强了这些小分子信号的转化相关性。基于基因编码或小分子的荧光报告基因以及氧化还原蛋白质组学，分别彻底改变了细胞氧化还原状态和亲电试剂传感器蛋白的观察与分析。然而，特定氧化还原修饰靶点与特定反应之间的精确映射直到最近才开始得到解决，并且在体内对基因操作和靶向氧化还原信号传导均易处理的系统仍然非常有限。在这里，我们构建了表达功能性卤代标签融合蛋白的转基因秀丽隐杆线虫，并利用该系统开发了一种通用的光控方法，用于在体内用天然亲电试剂标记典型的亲电试剂传感器蛋白。该方法规避了与低摄取/分布以及毒性/混杂性相关的问题。鉴于秀丽隐杆线虫在衰老研究中已得到验证的成功，这个优化的平台提供了一个新的视角，用以审视靶向亲电试剂信号传导如何影响氧化还原依赖性寿命调节。
