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1.
J Agric Food Chem ; 72(22): 12445-12458, 2024 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-38771652

RESUMO

Global water deficit is a severe abiotic stress threatening the yielding and quality of crops. Abscisic acid (ABA) is a phytohormone that mediates drought tolerance. Protein kinases and phosphatases function as molecular switches in eukaryotes. Protein phosphatases type 2C (PP2Cs) are a major family that play essential roles in ABA signaling and stress responses. However, the role and underlying mechanism of PP2C in rapeseed (Brassica napus L.) mediating drought response has not been reported yet. Here, we characterized a PP2C family member, BnaPP2C37, and its expression level was highly induced by ABA and dehydration treatments. It negatively regulates drought tolerance in rapeseed. We further identified that BnaPP2C37 interacted with multiple PYR/PYL receptors and a drought regulator BnaCPK5 (calcium-dependent protein kinase 5) through yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays. Specifically, BnaPYL1 and BnaPYL9 repress BnaPP2C37 phosphatase activity. Moreover, the pull-down assay and phosphatase assays show BnaPP2C37 interacts with BnaCPK5 to dephosphorylate BnaCPK5 and its downstream BnaABF3. Furthermore, a dual-luciferase assay revealed BnaPP2C37 transcript level was enhanced by BnaABF3 and BnaABF4, forming a negative feedback regulation to ABA response. In summary, we identified that BnaPP2C37 functions negatively in drought tolerance of rapeseed, and its phosphatase activity is repressed by BnaPYL1/9 whereas its transcriptional level is upregulated by BnaABF3/4.


Assuntos
Ácido Abscísico , Brassica napus , Secas , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas , Ácido Abscísico/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Plantas/genética , Brassica napus/genética , Brassica napus/metabolismo , Proteína Fosfatase 2C/metabolismo , Proteína Fosfatase 2C/genética , Estresse Fisiológico , Reguladores de Crescimento de Plantas/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Fosfoproteínas Fosfatases/genética , Resistência à Seca
2.
J Biol Chem ; 299(4): 103060, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-36841482

RESUMO

The mitogen-activated protein kinase kinase kinase 18 (MAPKKK18) has been reported to play a role in abiotic stress priming in long-term abscisic acid (ABA) response including drought tolerance and leaf senescence. However, the upstream transcriptional regulators of MAPKKK18 remain to be determined. Here, we report ABA-responsive element binding factors (ABFs) as upstream transcription factors of MAPKKK18 expression. Mutants of abf2, abf3, abf4, and abf2abf3abf4 dramatically reduced the transcription of MAPKKK18. Our electrophoresis mobility shift assay and dual-luciferase reporter assay demonstrated that ABF2, ABF3, and ABF4 bound to ABA-responsive element cis-elements within the promoter of MAPKKK18 to transactivate its expression. Furthermore, enrichments of the promoter region of MAPKKK18 by ABF2, ABF3, and ABF4 were confirmed by in vivo chromatin immunoprecipitation coupled with quantitative PCR. In addition, we found that mutants of mapkkk18 exhibited obvious delayed leaf senescence. Moreover, a genetic study showed that overexpression of ABF2, ABF3, and ABF4 in the background of mapkkk18 mostly phenocopied the stay-green phenotype of mapkkk18 and, expression levels of five target genes of ABFs, that is, NYE1, NYE2, NYC1, PAO, and SAG29, were attenuated as a result of MAPKKK18 mutation. These findings demonstrate that ABF2, ABF3, and ABF4 act as transcription regulators of MAPKKK18 and also suggest that, at least in part, ABA acts in priming leaf senescence via ABF-induced expression of MAPKKK18.


Assuntos
Ácido Abscísico , Proteínas de Arabidopsis , Arabidopsis , Regulação da Expressão Gênica de Plantas , Folhas de Planta , Senescência Vegetal , Elementos Reguladores de Transcrição , Ácido Abscísico/metabolismo , Ácido Abscísico/farmacologia , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Senescência Vegetal/genética , Senescência Vegetal/fisiologia , Plantas Geneticamente Modificadas/metabolismo , Fatores de Transcrição/metabolismo , Folhas de Planta/genética , Folhas de Planta/fisiologia
3.
Plant Sci ; 315: 111125, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-35067297

RESUMO

Drought is an environmental stress that causes severe crop loss. Drought stress can induce abscisic acid (ABA) accumulation and cytoplasmic calcium oscillation. Calcium-dependent protein kinases (CPKs) constitute a group of Ser/Thr protein kinases decoding calcium signals. However, the function and molecular mechanisms of most CPKs in oilseed rape (Brassica napus) remain unknown. Here, we report the functional characterization of BnaCPK5 in drought stress tolerance. BnaCPK5 belongs to Group I of the CPK family and was localized at the plasma membrane and nuclei. Overexpression of BnaCPK5 enhanced drought stress tolerance compared with the control. A screening of interacting proteins identified that BnaCPK5 interacted strongly with two ABA-Responsive Element Binding Factors (ABF/AREBs), BnaABF3 and BnaABF4. BnaCPK5 was shown to phosphorylate both BnaABF3 and BnaABF4 in a kinase assay. Further, it was found that the phosphorylation of BnaABF3 and BnaABF4 by BnaCPK5 increased their transcriptional activities against the famous drought stress marker gene, Responsive to Dehydration (RD) 29B and protein stability. Taken together, these data demonstrate that BnaCPK5 acts as a positive regulator of drought tolerance by, at least in part, phosphorylating two core ABA-signaling components to modulate Late-Embryogenesis Abundant (LEA)-like RD29B expression.


Assuntos
Ácido Abscísico/metabolismo , Adaptação Fisiológica/genética , Brassica napus/genética , Brassica napus/metabolismo , Cálcio/metabolismo , Secas , Proteínas Quinases/metabolismo , Produtos Agrícolas/genética , Produtos Agrícolas/metabolismo , Fatores de Ligação G-Box , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Fosforilação/genética , Fosforilação/fisiologia , Proteínas Quinases/genética
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