Journal Article

In vivo monitoring of plant small GTPase activation using a Förster resonance energy transfer biosensor

Shimamoto, Ko, Wong, Hann Ling, Matsuda, Tomonori, Kosami, Ken-ichi, Tan, Li, Okuda, Jun, Kaneko-Kawano, Takako, Higuchi, Masayuki, Kawasaki, Tsutomu, Akamatsu, Akira, Kawano, Yoji, Inada, Noriko, Wang, Qiong, Nagawa, Shingo

2018

Overview

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.

• Publisher: BioMed Central,Springer Nature B.V,BMC
• Subject: Bioimaging / Biological Techniques / Biomedical and Life Sciences / Biosensors / cell walls / disease resistance / Energy / Energy transfer / FRET sensor / Gene expression / Green fluorescent protein / Guanosine triphosphatases / guanosinetriphosphatase / Kinases / Life Sciences / Methodology / monitoring / Oryza sativa / Plant immunity / plant response / Plant Sciences / pollen tubes / protein-protein interactions / Proteins / protoplasts / rice / root hairs / Sensors / Small GTPase / Thin layer chromatography / transfection