电离辐射诱导组蛋白乙酰化引起的染色质结构解聚研究

Histone Acetylation Induced by Ionizing Radiation Causes Chromatin Decondensation

真核生物的DNA分子经高度压缩以染色质形式存在于细胞核中,染色质动态结构在DNA复制、基因转录和DNA修复等过程中起着重要的调控作用。核内染色质结构的原位高分辨解析和其结构变化定量表征一直受困于显微成像观测分辨率的限制。通过点击化学荧光标记EdU和STORM单分子定位显微成像,实验得到了细胞核内超分辨率染色质结构图像。基于提出的单分子团簇分析和最近邻距离算法分析发现,X射线辐照和TSA处理后的细胞核内核小体团簇数量显著增多,核小体团簇所占细胞核内的面积比相对于对照组增加,且团簇内平均EdU分子数降低。同时,重离子辐照活细胞在线成像实验获得的XRCC1招募动力学速率常数表明乙酰化处理使得DNA损伤密度降低。这些结果表明电离辐射和乙酰化处理均导致了染色质结构的松散化。STORM超分辨成像方法和分析算法及其获得的核小体团簇分布规律为染色质结构的松散提供了直接的定量表征数据支持。

Eukaryotic DNA molecule is highly compressed and exist in the form of chromatin in the nucleus.The dynamic structure of chromatin plays an important role in the process of DNA replication,gene transcription and DNA repair.In situ high-resolution analysis of chromatin structure and quantitative characterization of its structural changes have been limited by the optical resolution of microscopic imaging.The super-resolution chromatin structure image in the nucleus was obtained by click-reaction labeled EdU and the STORM single-molecule-localization microscopy.Based on the proposed single molecule cluster analysis and nearest neighbor distance algorithm,we found that the number of nuclear nucleosome clusters increased significantly after X-ray irradiation and TSA treatment,the area ratio of nucleosome clusters in the cell nucleus increased compared with the control group,and the average number of EdU molecules in the cluster decreased.At the same time,the rate constant of XRCC1 recruitment kinetics obtained from the on-line imaging experiment of heavy ion irradiated living cells showed that acetylation reduced the density of DNA damage.These results showed that both ionizing radiation and acetylation led to the decondensation of chromatin structure.The imaging method,analysis algorithm and the distribution of nucleosome clusters provide direct quantitative characterization data support for the decondensation of chromatin structure in cell nucleus.