The arabidopsis RING E3 ubiquitin ligase AtAIRP2 plays combinatory roles with AtAIRP1 in abscisic acid-mediated drought stress responses

The ubiquitin (Ub)-26S proteasome pathway is implicated in various cellular processes in higher plants. AtAIRP1, a C3H2C3-type RING (for Really Interesting New Gene) E3 Ub ligase, is a positive regulator in the Arabidopsis (Arabidopsis thaliana)abscisic acid (ABA)-dependent drought response. Here, the AtAIRP2 (for Arabidopsis ABA-insensitive RING protein 2) genewas identified and characterized. AtAIRP2 encodes a cytosolic C3HC4-type RING E3 Ub ligase whose expression wasmarkedly induced by ABA and dehydration stress. Thus, AtAIRP2 belongs to a different RING subclass than AtAIRP1 with alimited sequence identity. AtAIRP2-overexpressing transgenic (35S:AtAIRP2-sGFP) and atairp2 loss-of-function mutant plantsexhibited hypersensitive and hyposensitive phenotypes, respectively, to ABA in terms of seed germination, root growth, andstomatal movement. 35S:AtAIRP2-sGFP plants were highly tolerant to severe drought stress, and atairp2 alleles were moresusceptible to water stress than were wild-type plants. Higher levels of drought-induced hydrogen peroxide production weredetected in 35S:AtAIRP2-sGFP as compared with atairp2 plants. ABA-inducible drought-related genes were up-regulated in35S:AtAIRP2-sGFP and down-regulated in atairp2 progeny. The positive effects of AtAIRP2 on ABA-induced stress genes weredependent on SNF1-related protein kinases, key components of the ABA signaling pathway. Therefore, AtAIRP2 is involved inpositive regulation of ABA-dependent drought stress responses. To address the functional relationship between AtAIRP1 andAtAIRP2, FLAG-AtAIRP1 and AtAIRP2-sGFP genes were ectopically expressed in atairp2-2 and atairp1 plants, respectively.Constitutive expression of FLAG-AtAIRP1 and AtAIRP2-sGFP in atairp2-2 and atairp1 plants, respectively, reciprocally rescuedthe loss-of-function ABA-insensitive phenotypes during germination. Additionally, atairp1/35S:AtAIRP2-sGFP and atairp2-2/35S:FLAG-AtAIRP1 complementation lines were more tolerant to dehydration stress relative to atairp1 and atairp2-2 singleknockout plants. Overall, these results suggest that AtAIRP2 plays combinatory roles with AtAIRP1 in Arabidopsis ABAmediateddrought stress responses.