In vivo monitoring of plant small GTPase activation using a Förster resonance energy transfer biosensor
Hann Ling Wong,
Akira Akamatsu,
Qiong Wang,
Masayuki Higuchi,
Tomonori Matsuda,
Jun Okuda,
Ken-ichi Kosami,
Noriko Inada,
Tsutomu Kawasaki,
Takako Kaneko-Kawano,
Shingo Nagawa,
Li Tan,
Yoji Kawano,
Ko Shimamoto
Affiliations
Hann Ling Wong
Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology
Akira Akamatsu
Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology
Qiong Wang
Signal Transduction and Immunity Group, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Center for Plant Stress Biology, Chinese Academy of Sciences
Masayuki Higuchi
Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology
Tomonori Matsuda
Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology
Jun Okuda
Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology
Ken-ichi Kosami
Signal Transduction and Immunity Group, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Center for Plant Stress Biology, Chinese Academy of Sciences
Noriko Inada
College of Life, Environment, and Advanced, Osaka Prefecture University Sciences
Tsutomu Kawasaki
Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology
Takako Kaneko-Kawano
College of Pharmaceutical Sciences, Ritsumeikan University
Shingo Nagawa
Core Facility of Cell Biology, Shanghai Center for Plant Stress Biology
Li Tan
Core Facility of Cell Biology, Shanghai Center for Plant Stress Biology
Yoji Kawano
Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology
Ko Shimamoto
Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology
Abstract Background Small GTPases act as molecular switches that regulate various plant responses such as disease resistance, pollen tube growth, root hair development, cell wall patterning and hormone responses. Thus, to monitor their activation status within plant cells is believed to be the key step in understanding their roles. Results We have established a plant version of a Förster resonance energy transfer (FRET) probe called Ras and interacting protein chimeric unit (Raichu) that can successfully monitor activation of the rice small GTPase OsRac1 during various defence responses in cells. Here, we describe the protocol for visualizing spatiotemporal activity of plant Rac/ROP GTPase in living plant cells, transfection of rice protoplasts with Raichu-OsRac1 and acquisition of FRET images. Conclusions Our protocol should be adaptable for monitoring activation for other plant small GTPases and protein–protein interactions for other FRET sensors in various plant cells.