Xie Xin, Kang Houxiang, Liu Wende, Wang Guo-Liang
†State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
‡Department of Plant Pathology, College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China.
J Proteome Res. 2015 May 1;14(5):2017-25. doi: 10.1021/pr5009724. Epub 2015 Apr 6.
Protein ubiquitination is a major post-translational modification that regulates development, apoptosis, responses to environmental cues, and other processes in eukaryotes. Although several ubiquitinated proteins have been identified in rice, large-scale profiling of the rice ubiquitome has not been reported because of limitations in the current analytical methods. Here, we report the first rice ubiquitome, determined by combining highly sensitive immune affinity purification and high resolution LC-MS/MS. We identified 861 di-Gly-Lys-containing peptides in 464 proteins in rice leaf cells. Bioinformatic analyses of the ubiquitome identified a variety of cellular functions and diverse subcellular localizations for the ubiquitinated proteins, and also revealed seven putative ubiquitination motifs in rice. Proteins related to binding and catalytic activity were predicted to be the preferential targets of lysine ubiquitination. A protein interaction network and KEGG analysis indicated that a wide range of signaling and metabolic pathways are modulated by protein ubiquitination in rice. Our results demonstrate the usefulness of the significantly improved method for assaying proteome-wide ubiquitination in plants. The identification of the 464 ubiquitinated proteins in rice leaves provides a foundation for the analysis of the physiological roles of these ubiquitination-related proteins.
蛋白质泛素化是一种主要的翻译后修饰，可调节真核生物的发育、细胞凋亡、对环境信号的反应及其他过程。尽管在水稻中已鉴定出几种泛素化蛋白，但由于当前分析方法的局限性，尚未有关于水稻泛素组大规模分析的报道。在此，我们报告了首个通过结合高灵敏度免疫亲和纯化和高分辨率液相色谱-串联质谱法测定的水稻泛素组。我们在水稻叶细胞的464种蛋白质中鉴定出861个含双甘氨酸-赖氨酸的肽段。对泛素组的生物信息学分析确定了泛素化蛋白的多种细胞功能和不同的亚细胞定位，还揭示了水稻中的七个假定泛素化基序。预测与结合和催化活性相关的蛋白质是赖氨酸泛素化的优先靶标。蛋白质相互作用网络和KEGG分析表明，水稻中的多种信号传导和代谢途径受蛋白质泛素化调控。我们的结果证明了显著改进的植物全蛋白质组泛素化检测方法的实用性。水稻叶片中464种泛素化蛋白的鉴定为分析这些泛素化相关蛋白的生理作用奠定了基础。
