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.
Data availability
Data generated or analysed during this study are included in the manuscript and supporting files.
Article and author information
Author details
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.
Reviewing Editor
- Dominique C Bergmann, Stanford University, United States
Publication history
- Received: February 25, 2020
- Accepted: May 20, 2020
- Accepted Manuscript published: May 28, 2020 (version 1)
- Version of Record published: June 11, 2020 (version 2)
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.
Metrics
-
- 3,671
- Page views
-
- 775
- Downloads
-
- 26
- Citations
Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, PubMed Central.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Plant Biology
Plant genomes encode hundreds of secreted peptides; however, relatively few have been characterised. We report here an uncharacterised, stress-induced family of plant signalling peptides, which we call CTNIPs. Based on the role of the common co-receptor BRASSINOSTEROID INSENSITIVE 1-ASSOCIATED KINASE 1 (BAK1) in CTNIP-induced responses, we identified in Arabidopsis thaliana the orphan receptor kinase HAESA-LIKE 3 (HSL3) as the CTNIP receptor via a proteomics approach. CTNIP-binding, ligand-triggered complex formation with BAK1, and induced downstream responses all involve HSL3. Notably, the HSL3-CTNIP signalling module is evolutionarily conserved amongst most extant angiosperms. The identification of this novel signalling module will further shed light on the diverse functions played by plant signalling peptides and will provide insights into receptor-ligand co-evolution.
-
- Developmental Biology
- Plant Biology
Positional information is a central concept in developmental biology. In developing organs, positional information can be idealized as a local coordinate system that arises from morphogen gradients controlled by organizers at key locations. This offers a plausible mechanism for the integration of the molecular networks operating in individual cells into the spatially coordinated multicellular responses necessary for the organization of emergent forms. Understanding how positional cues guide morphogenesis requires the quantification of gene expression and growth dynamics in the context of their underlying coordinate systems. Here, we present recent advances in the MorphoGraphX software (Barbier de Reuille et al., 2015) that implement a generalized framework to annotate developing organs with local coordinate systems. These coordinate systems introduce an organ-centric spatial context to microscopy data, allowing gene expression and growth to be quantified and compared in the context of the positional information thought to control them.