FRET kinase sensor development reveals SnRK2/OST1 activation by ABA but not by MeJA and high CO2 during stomatal closure
Abstract
Sucrose-non-fermenting-1-related protein kinase-2s (SnRK2s) are critical for plant abiotic stress responses, including abscisic acid (ABA) signaling. Here, we develop a genetically encoded reporter for SnRK2 kinase activity. This sensor, named SNACS, shows an increase in the ratio of yellow to cyan fluorescence emission by OST1/SnRK2.6-mediated phosphorylation of a defined serine residue in SNACS. ABA rapidly increases FRET efficiency in N. benthamiana leaf cells and Arabidopsis guard cells. Interestingly, protein kinase inhibition decreases FRET efficiency in guard cells, providing direct experimental evidence that basal SnRK2 activity prevails in guard cells. Moreover, in contrast to ABA, the stomatal closing stimuli, elevated CO2 and MeJA, did not increase SNACS FRET ratios. These findings and gas exchange analyses of quintuple/sextuple ABA receptor mutants show that stomatal CO2 signaling requires basal ABA and SnRK2 signaling, but not SnRK2 activation. A recent model that CO2 signaling is mediated by PYL4/PYL5 ABA-receptors could not be supported here in two independent labs. We report a potent approach for real-time live-cell investigations of stress signaling.
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Data generated or analysed during this study are included in the manuscript and supporting files.
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Funding
National Science Foundation (MCB-1900567)
- Julian I Schroeder
National Institutes of Health (GM060396)
- Julian I Schroeder
China Scholarship Council
- Li Zhang
Japan Society for the Promotion of Science
- Yohei Takahashi
Eesti Teadusagentuur (PUT1133)
- Kollist Hannes
Eesti Teadusagentuur (PRG719)
- Kollist Hannes
Eesti Teadusagentuur (PRG433)
- Kollist Hannes
European Regional Development Fund
- Ebe Merilo
National Science Foundation (MCB‐1137950)
- Po-Kai Hsu
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2020, Zhang et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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