Mapping post-translational modifications of the histone variant macroH2A1 using tandem mass spectrometry
Mapping post-translational modifications of the histone variant macroH2A1 using tandem mass spectrometry
复制标题
DOI：
10.1074/mcp.m500285-mcp200
复制
发表时间：
2006-01-01
影响因子：
7
通讯作者：
Burlingame, AL
中科院分区：
文献类型：
--
作者：
Chu, FX;Nusinow, DA;Burlingame, AL
翻译后的组蛋白修饰调节染色质模板化过程，从而影响细胞的增殖、生长和发育。尽管对核心组蛋白的翻译后修饰已经进行了多年的深入研究，但对变异型组蛋白的修饰却知之甚少。我们使用串联质谱仪来确定组蛋白H_2A变异体宏H_2A_1.2上的共价修饰。MacroH2A1.2可以是一元化的；然而，一元化的位置还没有文献记载。在这项研究中，我们使用绿色荧光蛋白标记的宏H_2A1.2来确定Lys(115)是泛素化的位点。此外，我们发现该变异体在赖氨酸残基Lys(17)、Lys(122)和Lys(238)的epsilon氨基上发生甲基化，并在Thr(128)上发生磷酸化。质谱分析还发现，内源性蛋白质中也存在其中的三种修饰。这些结果首次直接证明了宏H_2A 1.2上有多个翻译后修饰，这表明，像典型的H_2A一样，这种变异体H_2A受到共价修饰的组合调控。
Post-translational histone modifications modulate chromatin-templated processes and therefore affect cellular proliferation, growth, and development. Although posttranslational modifications on the core histones have been under intense investigation for several years, the modifications on variant histones are poorly understood. We used tandem mass spectrometry to identify covalent modifications on a histone H2A variant, macroH2A1.2. MacroH2A1.2 can be monoubiquitinated; however, the site of monoubiquitination has not been documented. In this study we used green fluorescent protein-tagged macroH2A1.2 to determine that Lys(115) is a site of ubiquitination. In addition, we found that this variant H2A is methylated on the epsilon amino group of lysine residues Lys(17), Lys(122), and Lys(238) and phosphorylated on Thr(128). Three of these modifications were also found to be present in the endogenous protein by mass spectrometric analysis. These results provide the first direct evidence that multiple post-translational modifications are imposed on macroH2A1.2, suggesting that, like canonical H2A, this variant H2A is subject to regulation by combinatorial use of covalent modifications.