Identification and characterization of NEDD8-modified proteins from plants

NEDD8是真核细胞中的泛素相关蛋白修饰物。类泛素化调节cullin-RING型E3泛素连接酶在动物、植物和真菌中的组装和活性。除cullins外，已知的nedd8修饰蛋白数量非常有限，但迄今为止还没有从植物中发现此类蛋白的报道。利用拟南芥NEDD8 （RUB1）的标记版本，我们从植物模式物种拟南芥中纯化了许多推测的NEDD8偶联物。在这里，我们建议在体内确认这些蛋白质的类化修饰，绘制类化修饰位点，并检查类化修饰与蛋白质功能的相关性。我们还建议建立一个更精细的NEDD8偶联物纯化方案，以更全面地了解植物nedd8yome，并研究缺乏NEDD8偶联和去偶联的突变体中NEDD8修饰蛋白的差异积累。

NEDD8 is a ubiquitin-related protein modifier in eukaryotic cells. Neddylation regulates the assembly and activity of cullin-RING type E3 ubiquitin ligases in animals, plants, and fungi. Besides cullins, only a very limited number of NEDD8-modified proteins are known, but no such proteins have as yet been reported from plants. Using a tagged version of Arabidopsis NEDD8 (RUB1), we have purified a number of putative NEDD8-conjugates from the plant model species Arabidopsis thaliana. Here, we propose to confirm the neddylation of these proteins in vivo, to map the neddylation sites, and to examine the relevance of neddylation for protein function. We also propose to establish a more refined purification scheme for NEDD8 conjugates to get a more complete overview of the plant NEDD8ylome and to examine the differential accumulation of NEDD8-modified proteins in mutants deficient in NEDD8 conjugation and deconjugation.