Large-scale analysis of lysine SUMOylation by SUMO remnant immunoaffinity profiling
Large-scale analysis of lysine SUMOylation by SUMO remnant immunoaffinity profiling
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DOI：
10.1038/ncomms6409
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发表时间：
2014-11-01
影响因子：
16.6
通讯作者：
Thibault, Pierre
中科院分区：
文献类型：
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作者：
Lamoliatte, Frederic;Caron, Danielle;Thibault, Pierre
小泛素相关修饰因子（SUMO）是一种进化上保守的泛素样蛋白，可调节多种细胞过程，包括细胞周期进程、细胞内运输、蛋白质降解和凋亡。尽管SUMOylation蛋白在不同的生物学途径中具有重要意义，但复杂细胞提取物中受体位点的全球鉴定仍然是一个挑战。在这里，我们产生了一种单克隆抗体，丰富了胰蛋白酶消化后含有SUMO残链的肽。我们鉴定了538种蛋白质上954个sumo3修饰的赖氨酸残基，并通过定量蛋白质组学分析了蛋白酶体抑制后蛋白质sumo3化的动态变化。超过86%的SUMOylation位点以前没有报道过，包括肿瘤抑制因子parafibromin （CDC73）上的5个位点。CDC73在K136位点的修饰在蛋白酶体抑制下影响其在PML核体内的核保留。相比之下，CDC73 K136R突变体在相同条件下易位到细胞质中，进一步证明了我们的方法表征赖氨酸SUMOylation动力学的有效性。
Small ubiquitin-related modifiers (SUMO) are evolutionarily conserved ubiquitin-like proteins that regulate several cellular processes including cell cycle progression, intracellular trafficking, protein degradation and apoptosis. Despite the importance of protein SUMOylation in different biological pathways, the global identification of acceptor sites in complex cell extracts remains a challenge. Here we generate a monoclonal antibody that enriches for peptides containing SUMO remnant chains following tryptic digestion. We identify 954 SUMO3-modified lysine residues on 538 proteins and profile by quantitative proteomics the dynamic changes of protein SUMOylation following proteasome inhibition. More than 86% of these SUMOylation sites have not been reported previously, including 5 sites on the tumour suppressor parafibromin (CDC73). The modification of CDC73 at K136 affects its nuclear retention within PML nuclear bodies on proteasome inhibition. In contrast, a CDC73 K136R mutant translocates to the cytoplasm under the same conditions, further demonstrating the effectiveness of our method to characterize the dynamics of lysine SUMOylation.