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Abscisic Acid Increases Hydrogen Peroxide in Multiple Subcellular Compartments to Drive Stomatal Closure

View ORCID ProfileAnthony E. Postiglione, View ORCID ProfileGloria K. Muday
doi: https://doi.org/10.1101/2022.01.11.475946
Anthony E. Postiglione
1Department of Biology and the Center for Molecular Signaling, Wake Forest University, Winston Salem, NC, USA
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Gloria K. Muday
1Department of Biology and the Center for Molecular Signaling, Wake Forest University, Winston Salem, NC, USA
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  • For correspondence: muday@wfu.edu
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Abstract

Stomatal closure regulates transpiration and gas exchange in response to environmental cues. Drought upregulates ABA signaling, which elevates levels of reactive oxygen species (ROS). However, the subcellular location and identity of these ROS has received limited study. We found that in guard cells, ABA increased fluorescence of the general redox sensor, dichlorofluorescein (DCF), in distinct subcellular locations including chloroplasts, cytosol, nuclei, and cytosolic puncta. These changes were lost in an ABA-insensitive quintuple receptor mutant and accentuated in an ABA-hypersensitive mutant. ABA induced ROS accumulation in these subcellular compartments was lost in mutants with defects in genes encoding H2O2 elevating respiratory burst oxidase homolog (RBOH) enzymes and guard cells treated with the RBOH inhibitor VAS2870, while exogenous H2O2 treatment is sufficient to close guard cells. The H2O2-selective probe, peroxy orange 1, also showed ABA-dependent increases in chloroplasts and cytosolic puncta. Using the more sensitive genetically-encoded H2O2 reporter roGFP-Orp1, we also detected significant H2O2 increases in the cytosol and nucleus. These cytosolic puncta accumulate ROS after ABA treatment and show colocalization with Mitotracker and with a mitochondrial targeted mt-roGFP2-Orp1, which also reports ABA-increased H2O2 in mitochondria. These results indicate that elevated H2O2 after ABA treatment in these subcellular compartments is necessary and sufficient to drive stomatal closure.

Competing Interest Statement

The authors have declared no competing interest.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted January 12, 2022.
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Abscisic Acid Increases Hydrogen Peroxide in Multiple Subcellular Compartments to Drive Stomatal Closure
Anthony E. Postiglione, Gloria K. Muday
bioRxiv 2022.01.11.475946; doi: https://doi.org/10.1101/2022.01.11.475946
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Abscisic Acid Increases Hydrogen Peroxide in Multiple Subcellular Compartments to Drive Stomatal Closure
Anthony E. Postiglione, Gloria K. Muday
bioRxiv 2022.01.11.475946; doi: https://doi.org/10.1101/2022.01.11.475946

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