OsKEAP1 Interacts with OsABI5 and Its Downregulation Increases the Transcription of and the ABA Response Genes in Germinating Rice Seeds.

Kelch-like ECH-associated protein 1 (KEAP1)-nuclear factor E2-related factor 2 (NRF2) is the key antioxidant system in animals. In a previous study, we identified a probable KEAP1 ortholog in rice, OsKEAP1, and demonstrated that the downregulation of could alter the redox system and impair plant growth, as well as increase the susceptibility to abscisic acid (ABA) in seed germination. However, no NRF2 orthologs have been identified in plants and the mechanism underlying the phenotype changes of downregulated mutants is yet unknown. An in silico search showed that OsABI5 is the gene that encodes a protein with the highest amino acid identity score (38.78%) to NRF2 in rice. In this study, we demonstrated that, via yeast two-hybrids analysis and bimolecular fluorescence complementation assays, OsKEAP1 interacted with OsABI5 via its Kelch repeat domain in the nucleus. In germinating seeds, the expression of was significantly downregulated in (39.5% that of the wild-type (WT)) and (64.5% that of WT), while the expression of was significantly increased only in (247.4% that of WT) but not in (104.8% that of WT). ABA (0.5 μM) treatment significantly increased the expression of and in both the mutants and WT, and 4 days post treatment, the transcription level of became significantly greater in (+87.2%) and (+55.0%) than that in the WT. The ABA-responsive genes ( and three genes), which are known to be activated by OsABI5, became more responsive to ABA in both mutants than in the WT. The transcript abundances of genes that regulate OsABI5, e.g., (encodes a kinase that activates OsABI5), , and (both encode proteins binding to OsSnRK2 and are involved in ABA signaling) were not significantly different between the two mutants and the WT. These results demonstrated that OsKEAP1 played a role in the ABA response in rice seed germination via regulating OsABI5, which is the key player in the ABA response. In-depth analyses of the components and their action mode of the KEAP1-NRF2 and ABA signaling pathways suggested that OsKEAP1 likely formed a complex with OsABI5 and OsKEG, and OsABI5 was ubiquitinated by OsKEG and subsequently degraded under physiological conditions; meanwhile, under oxidative stress or with increased an ABA level, OsABI5 was released from the complex, phosphorylated, and transactivated the ABA response genes. Therefore, OsKEAP1-OsABI5 bore some resemblance to KEAP1-NRF2 in terms of its function and working mechanism.