Department of Biochemistry and the Siebens-Drake Medical Research Institute, Schulich School of Medicine and Dentistry, University of Western Ontario, London, ON N6A 5C1, Canada.
Mol Cell. 2013 Jun 6;50(5):723-35. doi: 10.1016/j.molcel.2013.04.025. Epub 2013 May 23.
Lysine methylation occurs on both histone and nonhistone proteins. However, our knowledge on the prevalence and function of nonhistone protein methylation is poor. We describe an approach that combines peptide array, bioinformatics, and mass spectrometry to systematically identify lysine methylation sites and map methyllysine-driven protein-protein interactions. Using this approach, we identified a high-confidence and high-resolution interactome of the heterochromatin protein 1β (HP1β) and uncovered, simultaneously, numerous methyllysine sites on nonhistone proteins. We found that HP1β binds to DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and regulates its localization to double-strand breaks (DSBs) during DNA damage response (DDR). Mutation of the methylation sites in DNA-PKcs or depletion of HP1β in cells caused defects in DDR. Furthermore, we showed that the methylation of DNA-PKcs and many other proteins in the HP1β interactome undergoes large changes in response to DNA damage, indicating that Lys methylation is a highly dynamic posttranslational modification.
赖氨酸甲基化发生在组蛋白和非组蛋白蛋白质上。然而，我们对非组蛋白蛋白质甲基化的普遍性和功能知之甚少。我们描述了一种结合肽阵列、生物信息学和质谱的方法，系统地鉴定赖氨酸甲基化位点，并绘制甲基化赖氨酸驱动的蛋白质-蛋白质相互作用图谱。使用这种方法，我们鉴定了异染色质蛋白 1β (HP1β) 的高可信度和高分辨率的互作组，并同时揭示了非组蛋白蛋白质上的许多甲基化赖氨酸位点。我们发现 HP1β 与 DNA 依赖性蛋白激酶催化亚基 (DNA-PKcs) 结合，并在 DNA 损伤反应 (DDR) 期间调节其定位到双链断裂 (DSBs)。DNA-PKcs 中甲基化位点的突变或细胞中 HP1β 的缺失导致 DDR 缺陷。此外，我们表明，HP1β 互作组中 DNA-PKcs 和许多其他蛋白质的甲基化在 DNA 损伤后发生了很大变化，表明 Lys 甲基化是一种高度动态的翻译后修饰。
