title: Ethylene mediates brassinosteroid-induced stomatal closure via Gα protein-activated hydrogen peroxide and nitric oxide production in Arabidopsis.
abstract: Brassinosteroids (BRs) are essential for plant growth and development; however, whether and how they promote stomatal closure is not fully clear. In this study, we report that 24-epibrassinolide (EBR), a bioactive BR, induces stomatal closure in Arabidopsis (Arabidopsis thaliana) by triggering a signal transduction pathway including ethylene synthesis, the activation of Gα protein, and hydrogen peroxide (H(2)O(2)) and nitric oxide (NO) production. EBR initiated a marked rise in ethylene, H(2)O(2) and NO levels, necessary for stomatal closure in the wild type. These effects were abolished in mutant bri1-301, and EBR failed to close the stomata of gpa1 mutants. Next, we found that both ethylene and Gα mediate the inductive effects of EBR on H(2)O(2) and NO production. EBR-triggered H(2)O(2) and NO accumulation were canceled in the etr1 and gpa1 mutants, but were strengthened in the eto1-1 mutant and the cGα line (constitutively overexpressing the G protein α-subunit AtGPA1). Exogenously applied H(2)O(2) or sodium nitroprusside (SNP) rescued the defects of etr1-3 and gpa1 or etr1 and gpa1 mutants in EBR-induced stomatal closure, whereas the stomata of eto1-1/AtrbohF and cGα/AtrbohF or eto1-1/nia1-2 and cGα/nia1-2 constructs had an analogous response to H(2)O(2) or SNP as those of AtrbohF or Nia1-2 mutants. Moreover, we provided evidence that Gα plays an important role in the responses of guard cells to ethylene. Gα activator CTX largely restored the lesion of the etr1-3 mutant, but ethylene precursor ACC failed to rescue the defects of gpa1 mutants in EBR-induced stomatal closure. Lastly, we demonstrated that Gα-activated H(2)O(2) production is required for NO synthesis. EBR failed to induce NO synthesis in mutant AtrbohF, but it led to H(2)O(2) production in mutant Nia1-2. Exogenously applied SNP rescued the defect of AtrbohF in EBR-induced stomatal closure, but H(2)O(2) did not reverse the lesion of EBR-induced stomatal closure in Nia1-2. Together, our results strongly suggest a signaling pathway in which EBR induces ethylene synthesis, thereby activating Gα, and then promotes AtrbohF-dependent H(2)O(2) production and subsequent Nia1-catalyzed NO accumulation, and finally closes stomata.
authors.affiliation: School of Life Sciences, Shaanxi Normal University, Xi'an, 710062, China; School of Chemistry and Bioengineering, Hechi University, Yizhou, 546300, China.
authors.forename: Chenyu
authors.initials: C
authors.lastname: Shi
authors.forename: Cheng
authors.initials: C
authors.lastname: Qi
authors.forename: Hongyan
authors.initials: H
authors.lastname: Ren
authors.forename: Aixia
authors.initials: A
authors.lastname: Huang
authors.forename: Shumei
authors.initials: S
authors.lastname: Hei
authors.forename: Xiaoping
authors.initials: X
authors.lastname: She
mesh_terms: D017360:Arabidopsis / Q000187:drug effects* / Q000378:metabolism*; D060406:Brassinosteroids / Q000378:metabolism / Q000494:pharmacology*; D005030:Ethylenes / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D054046:Plant Stomata / Q000187:drug effects* / Q000378:metabolism*; D013260:Steroids, Heterocyclic / Q000494:pharmacology*
chemical_list: D060406:Brassinosteroids; D005030:Ethylenes; D013260:Steroids, Heterocyclic; D009569:Nitric Oxide; C036216:ethylene; D006861:Hydrogen Peroxide; C023623:brassinolide
keywords: Gα protein; brassinosteroid; ethylene; hydrogen peroxide; nitric oxide; stomata