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Traffic. 2019 Feb;20(2):168-180. doi: 10.1111/tra.12625. Epub 2019 Jan 8.

The use of quantitative imaging to investigate regulators of membrane trafficking in Arabidopsis stomatal closure.

Author information

1
The Sainsbury Laboratory, Norwich Research Park, Norwich, UK.
2
School of Biological Sciences, Life Sciences Building, University of Bristol, Bristol, UK.
3
The Earlham Institute, Norwich Research Park, Norwich, UK.
4
John Innes Centre, Norwich Research Park, Norwich, UK.
5
Unit of Plant Molecular Cell Biology, Institute for Biology I, RWTH Aachen University, Aachen, Germany.

Abstract

Expansion of gene families facilitates robustness and evolvability of biological processes but impedes functional genetic dissection of signalling pathways. To address this, quantitative analysis of single cell responses can help characterize the redundancy within gene families. We developed high-throughput quantitative imaging of stomatal closure, a response of plant guard cells, and performed a reverse genetic screen in a group of Arabidopsis mutants to five stimuli. Focussing on the intersection between guard cell signalling and the endomembrane system, we identified eight clusters based on the mutant stomatal responses. Mutants generally affected in stomatal closure were mostly in genes encoding SNARE and SCAMP membrane regulators. By contrast, mutants in RAB5 GTPase genes played specific roles in stomatal closure to microbial but not drought stress. Together with timed quantitative imaging of endosomes revealing sequential patterns in FLS2 trafficking, our imaging pipeline can resolve non-redundant functions of the RAB5 GTPase gene family. Finally, we provide a valuable image-based tool to dissect guard cell responses and outline a genetic framework of stomatal closure.

KEYWORDS:

ABA; ARA6; ARA7; FLS2; ROS; calcium; chitin; endosome; flg22; immunity

PMID:
30447039
DOI:
10.1111/tra.12625
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