Catalytic mechanism and kinase interactions of ABA-signaling PP2C phosphatases

Abscisic acid (ABA) is an essential hormone that controls plant growth, development and responses to abiotic stresses. ABA signaling is mediated by type 2C protein phosphatases (PP2Cs), including HAB1 and ABI2, which inhibit stress-activated SnRK2 kinases and whose activity is regulated by ABA and ABA receptors. Based on biochemical data and our previously determined crystal structures of ABI2 and the SnRK2.6–HAB1 complex, we present the catalytic mechanism of PP2C and provide new insight into PP2C–SnRK2 interactions and possible roles of other SnRK2 kinases in ABA signaling.     

Brassinosteroid signaling positively regulates abscisic acid biosynthesis in response to chilling stress in tomato

Brassinosteroids (BRs) and abscisic acid (ABA) are essential regulators of plant growth and stress tolerance. Although the antagonistic interaction of BRs and ABA is proposed to ensure the balance between growth and defense in model plants, the crosstalk between BRs and ABA in response to chilling in tomato (Solanum lycopersicum), a warm-climate horticultural crop, is unclear. Here, we determined that overexpression of the BR biosynthesis gene DWARF (DWF) or the key BR signaling gene BRASSINAZOLE-RESISTANT1 (BZR1) increases ABA levels in response to chilling stress via positively regulating the expression of the ABA biosynthesis gene 9-CIS-EPOXYCAROTENOID DIOXYGENASE1 (NCED1). BR-induced chilling tolerance was mostly dependent on ABA biosynthesis. Chilling stress or high BR levels decreased the abundance of BRASSINOSTEROID-INSENSITIVE2 (BIN2), a negative regulator of BR signaling. Moreover, we observed that chilling stress increases BR levels and results in the accumulation of BZR1. BIN2 negatively regulated both the accumulation of BZR1 protein and chilling tolerance by suppressing ABA biosynthesis. Our results demonstrate that BR signaling positively regulates chilling tolerance via ABA biosynthesis in tomato. The study has implications in production of warm-climate crops in horticulture.     

The abi1-1 mutation blocks ABA signaling downstream of cADPR action

Arabidopsis thaliana abscisic acid insensitive 1-1 (abi1-1) is a dominant mutant that is insensitive to the inhibition of germination and growth by the plant hormone, abscisic acid (ABA). The mutation severely decreases the catalytic activity of the ABI1 type 2C protein phosphatase (PP2C). However, the site of action of the abi1-1/ABI1 in the ABA signal transduction pathway has not yet been determined. Using single cell assays, we showed that microinjecting mutant abi1-1 protein inhibited the activation of RD29A-GUS and KIN2-GUS in response to ABA, cyclic ADP-ribose (cADPR), and Ca2+. The inhibitory effect of the mutant protein, however, was reversed by co-microinjection of an excess amount of the ABI1 protein. In transgenic Arabidopsis plants, overexpression of abi1-1 rendered the plants insensitive to ABA during germination, whereas overexpression of ABI1 did not have any apparent effect. Moreover, transgenic plants overexpressing abi1-1 were blocked in the induction of ABA-responsive genes; however, overexpression of ABI1 did not affect gene expression. Taken together, our results demonstrate that abi1-1 is likely to be a dominant negative mutation and ABI1 likely acts downstream of cADPR in the ABA-signaling pathway. Our results on ABI1 overexpression in Arabidopsis are not compatible with a negative regulatory role of this phosphatase in ABA responses.     

The ABSCISIC ACID-INSENSITIVE 3 (ABI3) gene is expressed during vegetative quiescence processes in Arabidopsis

The ABSCISIC ACID-INSENSITIVE 3 ( ABI3 ) gene of Arabidopsis thaliana (L.) Heynh is known to play an important role during seed maturation and dormancy. Here, we present evidence suggesting an additional role for ABI3 during vegetative quiescence processes. During growth in the dark, ABI3 is expressed in the apex of the seedlings after cell division is arrested. The 2S seed storage protein gene, a target gene of ABI3 in seeds, is also induced in the arrested apex under similar darkness conditions. In addition, β -glucuronidase expression under the control of the ABI3 promoter is abolished by treatments that provoke leaf development in the dark [sucrose and abscisic acid (ABA) biosynthesis inhibitors] and induced by treatments that prevent leaf development (darkness and ABA). Furthermore, ABI3 expression is absent in apices of dark-grown de-etiolated ( det 1 ) and abi3 mutants, both known to develop leaves or leaf primordia in the dark. The fact that the expression of the ABI3 gene is only observed in a fraction of the analysed plants suggests that ABI3 is probably only one of the components of a molecular network underlying quiescence. In addition to the expression of ABI3 in apices of dark-grown seedlings, the ABI3 promoter confers expression in other vegetative organs as well, such as the stipules and the abscission zones of the siliques. In conclusion, apart from its role in seed development, ABI3 might have additional functions.     

ABA is required for Leptosphaeria maculans resistance via ABI1- and ABI4-dependent signaling

Abscisic acid (ABA) is a defense hormone with influence on callose-dependent and -independent resistance against Leptosphaeria maculans acting in the RLMcol pathway. ABA-deficient and -insensitive mutants in Ler-0 background (abal-3 and abil-1) displayed susceptibility to L. maculans, along with a significantly decreased level of callose depositions, whereas abi2-1 and abi3-1 remained resistant, together with the abi5-1 mutant of Ws-0 background. Suppressor mutants of abil-1 confirmed that the L. maculans-susceptible response was due to the dominant negative nature of the abil-1 mutant. Highly induced camalexin levels made ABA mutants in Col-0 background (aba2-1, aba3-1, and abi4-1) appear resistant, but displayed enhanced susceptibility as double mutants with pad3-1, impaired in camalexin biosynthesis. beta-Aminobutyric acid (BABA) pretreatment of Ler-0 contributed to an elevated level of endogenous ABA after L. maculans inoculation. Comparisons between (RLM1co1)pad3 and rlmlLerpad3 showed that ABA and BABA enhancement of callose deposition requires induction from RLM1col. ABII, but not ABI2, was found to be involved in a feedback mechanism that modulates RLM1co, expression. Genetic analysis showed further that this feedback occurs upstream of ABI4 and that components downstream of ABI4 modulate ABIJ activity. ABA and BABA treatments of the L. maculans-susceptible callose synthase mutant pmr4 showed that ABA also induces a callose-independent resistance. Similar treatments enhanced callose depositions and induced resistance to L. maculans in oilseed rape, and BABA-induced resistance was found to be independent of salicylic acid.     

Arabidopsis small ubiquitin‐related modifier protease ASP1 positively regulates abscisic acid signaling during early seedling development

The small ubiquitin‐related modifier (SUMO) modification plays an important role in the regulation of abscisic acid (ABA) signaling, but the function of the SUMO protease, in ABA signaling, remains largely unknown. Here, we show that the SUMO protease, ASP1 positively regulates ABA signaling. Mutations in ASP1 resulted in an ABA‐insensitive phenotype, during early seedling development. Wild‐type ASP1 successfully rescued, whereas an ASP1 mutant (C577S), defective in SUMO protease activity, failed to rescue, the ABA‐insensitive phenotype of asp1‐1. Expression of ABI5 and MYB30 target genes was attenuated in asp1‐1 and our genetic analyses revealed that ASP1 may function upstream of ABI5 and MYB30. Interestingly, ASP1 accumulated upon ABA treatment, and ABA‐induced accumulation of ABI5 (a positive regulator of ABA signaling) was abolished, whereas ABA‐induced accumulation of MYB30 (a negative regulator of ABA signaling) was increased in asp1‐1. These findings support the hypothesis that increased levels of ASP1, upon ABA treatment, tilt the balance between ABI5 and MYB30 towards ABI5‐mediated ABA signaling.