NEDD8 is an important regulatory factor in many biological processes. However, the substrates of neddylation and the relation between ubiquitin and NEDD8 pathways are remained largely unknown. Here, we showed that NEDD8 is covalently conjugated to H2A, and neddylation of H2A antagonizes its ubiquitination. NEDD8 suppresses H2A ubiquitination and decrease of the free NEDD8 level promotes H2A ubiquitination. We further found that E3 ligase RNF168 promotes both H2A ubiquitination and neddylation. Interestingly, RNF168 is a substrate of NEDD8 and neddylation of RNF168 is necessary for its E3 ubiquitin activity. Inhibition of RNF168 neddylation impairs the interaction between RNF168 and its E2 Ubc13. Moreover, in response to DNA damage, the level of H2A neddylation decreased with the increase of H2A ubiquitination, which facilitates DNA damage repair. And at the late stage of damage repair, H2A neddylation increased gradually while ubiquitination decreased to the basal levels. Mechanistically, NEDD8 negatively regulates DNA damage repair process by suppressing the ubiquitination of H2A and γH2AX, which further blocks the recruitment of damage-response protein BRCA1. Our findings elucidate the relation of H2A ubiquitination and neddylation, and suggest a novel modulate approach of DNA damage repair through neddylation pathway.