Large-Scale Identification of Protein Crotonylation Reveals Its Role in Multiple Cellular Functions

Lysine
crotonylation on histones is a recently identified post-translational
modification that has been demonstrated to associate with active promoters
and to directly stimulate transcription. Given that crotonyl-CoA is
essential for the acyl transfer reaction and it is a metabolic intermediate
widely localized within the cell, we postulate that lysine crotonylation
on nonhistone proteins could also widely exist. Using specific antibody
enrichment followed by high-resolution mass spectrometry analysis,
we identified hundreds of crotonylated proteins and lysine residues.
Bioinformatics analysis reveals that crotonylated proteins are particularly
enriched for nuclear proteins involved in RNA processing, nucleic
acid metabolism, chromosome organization, and gene expression. Furthermore,
we demonstrate that crotonylation regulates HDAC1 activity, expels
HP1α from heterochromatin, and inhibits cell cycle progression
through S-phase. Our data thus indicate that lysine crotonylation
could occur in a large number of proteins and could have important
regulatory roles in multiple nuclei-related cellular processes.

组蛋白赖氨酸巴豆酰化是近年来发现的一种翻译后修饰，现已证实其与活性启动子相关联，并可直接促进转录。鉴于巴豆酰辅酶A（crotonyl-CoA）是酰基转移反应的必需底物，且其作为广泛分布于细胞内的代谢中间产物，我们推测非组蛋白上的赖氨酸巴豆酰化修饰也可广泛存在。我们通过特异性抗体富集结合高分辨率质谱分析，鉴定出了数百个发生巴豆酰化修饰的蛋白质及赖氨酸位点。生物信息学分析显示，巴豆酰化修饰蛋白显著富集于参与RNA加工、核酸代谢、染色体组织及基因表达的核蛋白中。此外，我们证实巴豆酰化可调控组蛋白去乙酰化酶1（HDAC1）的活性，将异染色质蛋白1α（HP1α）从异染色质中逐出，并通过阻滞S期抑制细胞周期进程。综上，我们的研究数据表明，赖氨酸巴豆酰化修饰可发生于大量蛋白质中，并在多种核相关细胞过程中发挥重要的调控作用。