Institute for Research in Immunology and Cancer, Université de Montréal, P.O. Box 6128, Station, Centre-ville, Montréal, Québec, Canada H3C 3J7.
Department of Chemistry, Université de Montréal, P.O. Box 6128, Station, Centre-ville, Montréal, Québec, Canada H3C 3J7.
Nat Commun. 2017 Jan 18;8:14109. doi: 10.1038/ncomms14109.
Crosstalk between the SUMO and ubiquitin pathways has recently been reported. However, no approach currently exists to determine the interrelationship between these modifications. Here, we report an optimized immunoaffinity method that permits the study of both protein ubiquitylation and SUMOylation from a single sample. This method enables the unprecedented identification of 10,388 SUMO sites in HEK293 cells. The sequential use of SUMO and ubiquitin remnant immunoaffinity purification facilitates the dynamic profiling of SUMOylated and ubiquitylated proteins in HEK293 cells treated with the proteasome inhibitor MG132. Quantitative proteomic analyses reveals crosstalk between substrates that control protein degradation, and highlights co-regulation of SUMOylation and ubiquitylation levels on deubiquitinase enzymes and the SUMOylation of proteasome subunits. The SUMOylation of the proteasome affects its recruitment to promyelocytic leukemia protein (PML) nuclear bodies, and PML lacking the SUMO interacting motif fails to colocalize with SUMOylated proteasome further demonstrating that this motif is required for PML catabolism.
SUMO 和泛素途径之间的串扰最近有报道。然而，目前还没有方法可以确定这些修饰之间的相互关系。在这里，我们报告了一种优化的免疫亲和方法，允许从单个样品中研究蛋白质的泛素化和 SUMO 化。该方法能够以前所未有的方式鉴定出 HEK293 细胞中的 10388 个 SUMO 位点。SUMO 和泛素残基免疫亲和纯化的顺序使用，促进了在蛋白酶体抑制剂 MG132 处理的 HEK293 细胞中 SUMO 化和泛素化蛋白质的动态分析。定量蛋白质组学分析揭示了控制蛋白质降解的底物之间的串扰，并强调了去泛素化酶上 SUMO 化和泛素化水平的共同调节以及蛋白酶体亚基的 SUMO 化。蛋白酶体的 SUMO 化影响其向早幼粒细胞白血病蛋白（PML）核体的募集，并且缺乏 SUMO 相互作用基序的 PML 不能与 SUMO 化的蛋白酶体共定位，进一步证明该基序是 PML 代谢所必需的。
