Differentially Isotope-Labeled Nucleosomes To Study Asymmetric Histone Modification Crosstalk by Time-Resolved NMR Spectroscopy.
Differentially Isotope-Labeled Nucleosomes To Study Asymmetric Histone Modification Crosstalk by Time-Resolved NMR Spectroscopy.
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DOI：
10.1002/anie.201601938
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发表时间：
2016-07-11
期刊：
影响因子：
--
通讯作者：
Schwarzer D
中科院分区：
文献类型：
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作者：
Liokatis S;Klingberg R;Tan S;Schwarzer D
组蛋白的翻译后修饰（PTM）调节染色质的结构和功能。由于核小体含有四种核心组蛋白中的每一种的两个拷贝，因此难以分析在相同核小体背景下在个体“姐妹”组蛋白上建立不同的PTM，即不对称组蛋白PTM。在这里，我们产生了差异同位素标记的核小体研究不对称组蛋白修饰串扰的时间分辨NMR光谱。具体而言，我们描绘的机制洞察核小体组蛋白H3修饰反应的顺式和反式，因此在个别H3拷贝，或它们之间。我们使用Gcn5乙酰转移酶介导的H3S10phK14ac串扰机制验证了我们的方法。此外，对甲基化底物的磷酸化测定鉴定了Haspin激酶在某些条件下能够产生用两种不同类型的PTM不对称修饰的核小体。
Post-translational modifications (PTMs) of histones regulate chromatin structure and function. Because nucleosomes contain two copies each of the four core histones, the establishment of different PTMs on individual ‘sister’ histones in the same nucleosomal context, i.e. asymmetric histone PTMs, are difficult to analyze. Here, we generated differentially isotope-labeled nucleosomes to study asymmetric histone modification crosstalk by time-resolved NMR spectroscopy. Specifically, we delineate mechanistic insights into nucleosomal histone H3 modification reactions in cis and in trans, hence within individual H3 copies, or between them. We validated our approach using the H3S10phK14ac crosstalk mechanism, mediated by Gcn5 acetyltransferase. Moreover, phosphorylation assays on methylated substrates identified Haspin kinase able under certain conditions to produce nucleosomes decorated asymmetrically with two distinct types of PTMs.