Ren Ruotong, Deng Liping, Xue Yanhong, Suzuki Keiichiro, Zhang Weiqi, Yu Yang, Wu Jun, Sun Liang, Gong Xiaojun, Luan Huiqin, Yang Fan, Ju Zhenyu, Ren Xiaoqing, Wang Si, Tang Hong, Geng Lingling, Zhang Weizhou, Li Jian, Qiao Jie, Xu Tao, Qu Jing, Liu Guang-Hui
National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
University of Chinese Academy of Sciences, Beijing 100049, China.
Cell Res. 2017 Apr;27(4):483-504. doi: 10.1038/cr.2017.18. Epub 2017 Jan 31.
Visualization of specific genomic loci in live cells is a prerequisite for the investigation of dynamic changes in chromatin architecture during diverse biological processes, such as cellular aging. However, current precision genomic imaging methods are hampered by the lack of fluorescent probes with high specificity and signal-to-noise contrast. We find that conventional transcription activator-like effectors (TALEs) tend to form protein aggregates, thereby compromising their performance in imaging applications. Through screening, we found that fusing thioredoxin with TALEs prevented aggregate formation, unlocking the full power of TALE-based genomic imaging. Using thioredoxin-fused TALEs (TTALEs), we achieved high-quality imaging at various genomic loci and observed aging-associated (epi) genomic alterations at telomeres and centromeres in human and mouse premature aging models. Importantly, we identified attrition of ribosomal DNA repeats as a molecular marker for human aging. Our study establishes a simple and robust imaging method for precisely monitoring chromatin dynamics in vitro and in vivo.
在活细胞中可视化特定基因组位点是研究不同生物过程（如细胞衰老）中染色质结构动态变化的先决条件。然而，目前的精确基因组成像方法受到缺乏具有高特异性和信噪比对比度的荧光探针的阻碍。我们发现传统的转录激活样效应因子（TALEs）倾向于形成蛋白质聚集体，从而损害其在成像应用中的性能。通过筛选，我们发现将硫氧还蛋白与TALEs融合可防止聚集体形成，从而释放基于TALE的基因组成像的全部潜力。使用硫氧还蛋白融合的TALEs（TTALEs），我们在各种基因组位点实现了高质量成像，并在人类和小鼠早衰模型中观察到端粒和着丝粒处与衰老相关的（表观）基因组改变。重要的是，我们确定核糖体DNA重复序列的损耗是人类衰老的分子标志物。我们的研究建立了一种简单而强大的成像方法，用于精确监测体外和体内的染色质动态。
