Li Tingting, Guan Junhong, Huang Ziji, Hu Xiang, Zheng Xiaofeng
State Key Lab of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing 100871, China Department of Biochemistry and Molecular Biology, School of Life Sciences, Peking University, Beijing 100871, China
Department of Biochemistry and Molecular Biology, School of Life Sciences, Peking University, Beijing 100871, China.
J Cell Sci. 2014 May 15;127(Pt 10):2238-48. doi: 10.1242/jcs.138891. Epub 2014 Mar 14.
NEDD8 is an important regulatory factor in many biological processes. However, the substrates for neddylation, and the relationship between the ubiquitin and NEDD8 pathways remain largely unknown. Here, we show that NEDD8 is covalently conjugated to histone 2A (H2A), and that neddylation of H2A antagonizes its ubiquitylation. NEDD8 suppresses ubiquitylation of H2A, and a decreased level of free NEDD8 promotes H2A ubiquitylation. Furthermore, we found that the E3 ligase RNF168 promotes both H2A ubiquitylation and neddylation. Interestingly, RNF168 is itself a substrate for NEDD8, and neddylation of RNF168 is necessary for its E3 ubiquitin activity. Inhibition of RNF168 neddylation impairs the interaction between RNF168 and its E2 enzyme Ubc13 (also known as UBE2N). Moreover, in response to DNA damage, the level of H2A neddylation decreased with an increase in the ubiquitylation of H2A, which facilitates DNA damage repair. During the later stages of damage repair, H2A neddylation increased gradually, whereas ubiquitylation decreased to basal levels. Mechanistically, NEDD8 negatively regulates the DNA damage repair process through suppression of the ubiquitylation of H2A and γH2AX, which further blocks the recruitment of the damage response protein BRCA1. Our findings elucidate the relationship of H2A ubiquitylation and neddylation, and suggest a novel modulatory approach to DNA damage repair through the neddylation pathway.
NEDD8是许多生物过程中的重要调节因子。然而，NEDD8化修饰的底物以及泛素与NEDD8途径之间的关系在很大程度上仍然未知。在此，我们表明NEDD8与组蛋白2A（H2A）共价结合，并且H2A的NEDD8化修饰拮抗其泛素化修饰。NEDD8抑制H2A的泛素化修饰，而游离NEDD8水平的降低促进H2A的泛素化修饰。此外，我们发现E3连接酶RNF168促进H2A的泛素化修饰和NEDD8化修饰。有趣的是，RNF168自身是NEDD8的底物，并且RNF168的NEDD8化修饰对于其E3泛素活性是必需的。抑制RNF168的NEDD8化修饰会损害RNF168与其E2酶Ubc13（也称为UBE2N）之间的相互作用。此外，在DNA损伤应答中，H2A的NEDD8化修饰水平随着H2A泛素化修饰的增加而降低，这有利于DNA损伤修复。在损伤修复的后期阶段，H2A的NEDD8化修饰逐渐增加，而泛素化修饰降低至基础水平。从机制上讲，NEDD8通过抑制H2A和γH2AX的泛素化修饰来负向调节DNA损伤修复过程，这进一步阻止了损伤应答蛋白BRCA1的募集。我们的研究结果阐明了H2A泛素化修饰与NEDD8化修饰之间的关系，并提出了一种通过NEDD8化修饰途径对DNA损伤修复进行新型调节的方法。
