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AtAIRP5 RING E3 ubiquitin ligase was recently identified as a positive regulator of the abscisic acid (ABA)-mediated drought stress response by stimulating the degradation of serine carboxypeptidase-like 1. Here, we identified GDSL-type esterase/lipase 22 (AtGELP22) and AtGELP23 as additional interacting partners of AtAIRP5. Yeast two-hybrid, pull-down, co-immunoprecipitation, and ubiquitination analyses verified that AtGELP22 and AtGELP23 are ubiquitinated target proteins of AtAIRP5. AtGELP22 and AtGELP23 were colocalized with AtAIRP5 to punctate-like structures in the cytosolic fraction, in which PYK10 and NAI2, two ER body marker proteins, are localized. T-DNA insertion atgelp22 and atgelp23 single knockout mutant plants showed phenotypes indistinguishable from those of wild-type plants under ABA treatment. In contrast, RNAi-mediated cosuppression of AtGELP22 and AtGELP23 resulted in hypersensitive ABA-mediated stomatal movements and higher tolerance to drought stress than that of the single mutant and wild-type plants. Taken together, our results suggest that the putative GDSL-type esterases/lipases AtGELP22 and AtGELP23 act as redundant negative regulators of the ABA-mediated drought stress response in Arabidopsis., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Ubiquitination is a major mechanism of eukaryotic posttranslational protein turnover that has been implicated in abscisic acid (ABA)-mediated drought stress response. Here, we isolated T-DNA insertion mutant lines in which ABA-insensitive RING protein 5 (AtAIRP5) was suppressed, resulting in hyposensitive ABA-mediated germination compared to wild-type Arabidopsis (Arabidopsis thaliana) plants. A homology search revealed that AtAIRP5 is identical to gibberellin (GA) receptor RING E3 ubiquitin (Ub) ligase (GARU), which downregulates GA signaling by degrading the GA receptor GID1, and thus AtAIRP5 was renamed AtAIRP5/GARU. The atairp5/garu knockout progeny were impaired in ABA-dependent stomatal closure and were markedly more susceptible to drought stress than wild-type plants, indicating a positive role for AtAIRP5/GARU in the ABA-mediated drought stress response. Yeast two-hybrid, pull-down, target ubiquitination, and in vitro and in planta degradation assays identified serine carboxypeptidase-like1 (AtSCPL1), which belongs to the clade 1A AtSCPL family, as a ubiquitinated target protein of AtAIRP5/GARU. atscpl1 single and atairp5/garu-1 atscpl1-2 double mutant plants were more tolerant to drought stress than wild-type plants in an ABA-dependent manner, suggesting that AtSCPL1 is genetically downstream of AtAIRP5/GARU. After drought treatment, the endogenous ABA levels in atscpl1 and atairp5/garu-1 atscpl1-2 mutant leaves were higher than those in wild-type and atairp5/garu leaves. Overall, our results suggest that AtAIRP5/GARU RING E3 Ub ligase functions as a positive regulator of the ABA-mediated drought response by promoting the degradation of AtSCPL1., (© American Society of Plant Biologists 2022. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Drought stress has detrimental effects on plants. Although the abscisic acid (ABA)-mediated drought response is well established, defensive mechanisms to cope with dehydration-induced proteotoxicity have been rarely studied. DRR1 was identified as an Arabidopsis drought-induced gene encoding an ER-localized RING-type E3 Ub ligase. Suppression of DRR1 markedly reduced tolerance to drought and proteotoxic stress without altering ABA-mediated germination and stomatal movement. Proteotoxicity- and dehydration-induced insoluble ubiquitinated protein accumulation was more obvious in DRR1 loss-of-function plants than in wild-type plants. These results suggest that DRR1 is involved in an ABA-independent drought stress response possibly through the mitigation of dehydration-induced proteotoxic stress., (© 2020 Institute of Botany, Chinese Academy of Sciences.)

Ubiquitination is a critical post-translational protein modification that has been implicated in diverse cellular processes, including abiotic stress responses, in plants. In the present study, we identified and characterized a T-DNA insertion mutant in the At5g10650 locus. Compared to wild-type Arabidopsis plants, at5g10650 progeny were hyposensitive to ABA at the germination stage. At5g10650 possessed a single C-terminal C3HC4-type Really Interesting New Gene (RING) motif, which was essential for ABA-mediated germination and E3 ligase activity in vitro. At5g10650 was closely associated with microtubules and microtubule-associated proteins in Arabidopsis and tobacco leaf cells. Localization of At5g10650 to the nucleus was frequently observed. Unexpectedly, At5g10650 was identified as JAV1-ASSOCIATED UBIQUITIN LIGASE1 (JUL1), which was recently reported to participate in the jasmonate signaling pathway. The jul1 knockout plants exhibited impaired ABA-promoted stomatal closure. In addition, stomatal closure could not be induced by hydrogen peroxide and calcium in jul1 plants. jul1 guard cells accumulated wild-type levels of H 2 O 2 after ABA treatment. These findings indicated that JUL1 acts downstream of H 2 O 2 and calcium in the ABA-mediated stomatal closure pathway. Typical radial arrays of microtubules were maintained in jul1 guard cells after exposure to ABA, H 2 O 2 , and calcium, which in turn resulted in ABA-hyposensitive stomatal movements. Finally, jul1 plants were markedly more susceptible to drought stress than wild-type plants. Overall, our results suggest that the Arabidopsis RING E3 ligase JUL1 plays a critical role in ABA-mediated microtubule disorganization, stomatal closure, and tolerance to drought stress., (© 2020 Society for Experimental Biology and John Wiley & Sons Ltd.)

AtUBC32, AtUBC33, and AtUBC34 comprise Arabidopsis group XIV E2 ubiquitin-conjugating enzymes. Yeast two-hybrid, in vitro pull-down, and bimolecular fluorescence complementation assays revealed that group XIV E2s are interacting partners of the U-box-type E3 ligase PUB19, a negative regulator of drought stress response. These three AtUBCs are co-localized with PUB19 to the punctae-like structures, most of which reside on the endoplasmic reticulum membrane of tobacco leaf cells. Suppression of AtUBC32, AtUBC33, and AtUBC34 resulted in increased abscisic acid-mediated stomatal closure and tolerance to drought stress. These results indicate that Arabidopsis group XIV E2s play negative roles in drought stress response., (Copyright © 2018 Elsevier GmbH. All rights reserved.)