Tracking epigenetic histone modifications in single cells using Fab-based live endogenous modification labeling.
Tracking epigenetic histone modifications in single cells using Fab-based live endogenous modification labeling.
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DOI：
10.1093/nar/gkr343
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发表时间：
2011-08
影响因子：
14.9
通讯作者：
Kimura H
中科院分区：
文献类型：
--
作者：
Hayashi-Takanaka Y;Yamagata K;Wakayama T;Stasevich TJ;Kainuma T;Tsurimoto T;Tachibana M;Shinkai Y;Kurumizaka H;Nozaki N;Kimura H
组蛋白修饰在表观遗传基因调控和基因组完整性中起重要作用。然而，这些修饰在个体细胞中如何动态变化在很大程度上仍然未知。通过使用荧光标记的特异性抗原结合片段（Fab），我们已经开发出一种通用的方法来监测活细胞中内源性组蛋白H3赖氨酸修饰的分布和全球水平，而不干扰细胞生长和胚胎发育。Fab产生不同的核模式，这些模式是其靶修饰的特征。例如，H3K27三甲基化特异性Fab集中在无活性的X染色体上。由于Fab瞬时结合其靶标，结合和游离分子的比例取决于靶标浓度，使我们能够测量全局修饰水平的变化。高亲和力Fab适合于小鼠胚胎成像，因此我们使用它们来监测通过体外受精和体细胞核移植产生的小鼠植入前胚胎中的H3K9和H3K27乙酰化水平。这些数据表明，高水平的H3K27乙酰化对正常胚胎发育很重要。由于基于Fab的活内源性修饰标记（FabLEM）广泛用于可视化任何修饰，因此它应该是未来研究细胞信号传导和诊断的有力工具。
Histone modifications play an important role in epigenetic gene regulation and genome integrity. It remains largely unknown, however, how these modifications dynamically change in individual cells. By using fluorescently labeled specific antigen binding fragments (Fabs), we have developed a general method to monitor the distribution and global level of endogenous histone H3 lysine modifications in living cells without disturbing cell growth and embryo development. Fabs produce distinct nuclear patterns that are characteristic of their target modifications. H3K27 trimethylation-specific Fabs, for example, are concentrated on inactive X chromosomes. As Fabs bind their targets transiently, the ratio of bound and free molecules depends on the target concentration, allowing us to measure changes in global modification levels. High-affinity Fabs are suitable for mouse embryo imaging, so we have used them to monitor H3K9 and H3K27 acetylation levels in mouse preimplantation embryos produced by in vitro fertilization and somatic cell nuclear transfer. The data suggest that a high level of H3K27 acetylation is important for normal embryo development. As Fab-based live endogenous modification labeling (FabLEM) is broadly useful for visualizing any modification, it should be a powerful tool for studying cell signaling and diagnosis in the future.
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DOI：
10.1083/jcb.149.2.271
发表时间：
2000-04-17
期刊：
The Journal of cell biology
影响因子：
--
作者：
Leonhardt H;Rahn HP;Weinzierl P;Sporbert A;Cremer T;Zink D;Cardoso MC
通讯作者：
Cardoso MC
影响因子：
4.8
作者：
Nightingale, Karl P.;Gendreizig, Susanne;Turner, Bryan M.
通讯作者：
Turner, Bryan M.
影响因子：
48
作者：
Rothbauer, Ulrich;Zolghadr, Kourosh;Leonhardt, Heinrich
通讯作者：
Leonhardt, Heinrich
影响因子：
7.8
作者：
Aye-Han, Nwe-Nwe;Ni, Qiang;Zhang, Jin
通讯作者：
Zhang, Jin
DOI：
10.1083/jcb.144.5.813
发表时间：
1999-03-08
期刊：
The Journal of cell biology
影响因子：
--
作者：
Manders EM;Kimura H;Cook PR
通讯作者：
Cook PR