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Unravelling the Function of a Bacterial Effector from a Non-cultivable Plant Pathogen Using a Yeast Two-hybrid Screen

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Functional complementation of yeast mutants to study plant signalling pathways.

Norbert Mehlmer¹, Elisabeth Scheikl-Pourkhalil, Markus Teige

¹Department of Biochemistry, Max F. Perutz Laboratories, University of Vienna, Vienna, Austria.

Methods in Molecular Biology (Clifton, N.J.)

|December 17, 2008

Summary

Researchers identified plant protein kinases involved in abiotic stress adaptation using yeast mutants. This method leverages the osmo-sensitive Hog1 pathway for efficient gene function elucidation in Arabidopsis thaliana.

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A Modified Yeast-one Hybrid System for Heteromeric Protein Complex-DNA Interaction Studies

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Area of Science:

Plant Molecular Biology
Yeast Genetics
Abiotic Stress Response

Background:

The growing number of sequenced genomes necessitates efficient methods for gene function discovery.
Functional complementation in yeast mutants has proven effective for isolating plant genes involved in various pathways.
The Hog1 mitogen-activated protein (MAP) kinase pathway in yeast is crucial for osmotic stress adaptation.

Purpose of the Study:

To describe a method for isolating plant protein kinases involved in abiotic stress adaptation.
To utilize the well-characterized Hog1 pathway in yeast as a model system.
To identify genes in Arabidopsis thaliana that confer tolerance to osmotic stress.

Main Methods:

Employing functional complementation of osmo-sensitive yeast mutants.
Utilizing yeast mutants with deletions in components of the Hog1 pathway (e.g., Pbs2, Hog1).
Selecting for yeast transformants exhibiting osmotolerance on high osmolarity media (e.g., 0.4 M NaCl).

Main Results:

Successfully isolated plant protein kinases that complement yeast osmotic stress sensitivity.
Demonstrated the feasibility of identifying components of signaling pathways sequentially.
Identified novel plant genes contributing to abiotic stress adaptation.

Conclusions:

Functional complementation in yeast is a powerful tool for discovering plant genes involved in abiotic stress.
The Hog1 pathway provides a robust system for screening and isolating stress-related plant kinases.
This approach facilitates the elucidation of plant signaling pathways in response to environmental challenges.