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首页> 外文期刊>Proteome science >Profiling post-translational modifications of histones in human monocyte-derived macrophages
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Profiling post-translational modifications of histones in human monocyte-derived macrophages

机译：在人类单核细胞衍生的巨噬细胞中分析组蛋白的翻译后修饰

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Background Histones and their post-translational modifications impact cellular function by acting as key regulators in the maintenance and remodeling of chromatin, thus affecting transcription regulation either positively (activation) or negatively (repression). In this study we describe a comprehensive, bottom-up proteomics approach to profiling post-translational modifications (acetylation, mono-, di- and tri-methylation, phosphorylation, biotinylation, ubiquitination, citrullination and ADP-ribosylation) in human macrophages, which are primary cells of the innate immune system. As our knowledge expands, it becomes more evident that macrophages are a heterogeneous population with potentially subtle differences in their responses to various stimuli driven by highly complex epigenetic regulatory mechanisms. Methods To profile post-translational modifications (PTMs) of histones in macrophages we used two platforms of liquid chromatography and mass spectrometry. One platform was based on Sciex5600 TripleTof and the second one was based on VelosPro Orbitrap Elite ETD mass spectrometers. Results We provide side-by-side comparison of profiling using two mass spectrometric platforms, ion trap and qTOF, coupled with the application of collisional induced and electron transfer dissociation. We show for the first time methylation of a His residue in macrophages and demonstrate differences in histone PTMs between those currently reported for macrophage cell lines and what we identified in primary cells. We have found a relatively low level of histone PTMs in differentiated but resting human primary monocyte derived macrophages. Conclusions This study is the first comprehensive profiling of histone PTMs in primary human MDM. Our study implies that epigenetic regulatory mechanisms operative in transformed cell lines and primary cells are overlapping to a limited extent. Our mass spectrometric approach provides groundwork for the investigation of how histone PTMs contribute to epigenetic regulation in primary human macrophages.
机译：背景组蛋白及其翻译后修饰通过充当染色质维持和重塑的关键调节剂来影响细胞功能，从而积极（激活）或消极（抑制）影响转录调控。在这项研究中，我们描述了一种全面的，自下而上的蛋白质组学方法，用于分析人类巨噬细胞中的翻译后修饰（乙酰化，单，二和三甲基化，磷酸化，生物素化，泛素化，瓜氨酸化和ADP-核糖基化）。先天免疫系统的原代细胞。随着我们知识的扩展，越来越明显的是，巨噬细胞是异质种群，它们对由高度复杂的表观遗传调控机制驱动的各种刺激的反应可能存在细微差异。方法为了描述巨噬细胞中组蛋白的翻译后修饰（PTM），我们使用了液相色谱和质谱的两个平台。一个平台基于Sciex5600 TripleTof，第二个平台基于VelosPro Orbitrap Elite ETD质谱仪。结果我们提供了使用两个质谱平台（离子阱和qTOF）以及碰撞诱导和电子转移解离的应用，对分析进行并排比较的结果。我们首次展示了巨噬细胞中His残基的甲基化，并证明了目前报道的巨噬细胞系与原代细胞中组蛋白PTM之间的差异。我们发现在分化但静止的人类原代单核细胞衍生的巨噬细胞中，组蛋白PTM的水平相对较低。结论本研究是人类原发性MDM中首次对组蛋白PTM进行综合分析。我们的研究表明，在转化细胞系和原代细胞中起作用的表观遗传调控机制在一定程度上重叠。我们的质谱方法为研究组蛋白PTM如何促进人类原代巨噬细胞的表观遗传调控提供了基础。

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