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

Abstract

ABSTRACT 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. 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 rice 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. Our protocol should be widely adaptable for monitoring activation for other plant small GTPases and for other FRET sensors in various plant cells.

Plant phenotyping relevance

植物細胞内のGTPase活性という生理状態を、FRETバイオセンサーで可視化・測定するプロトコルを中心的に開発・記述しているため、植物フェノタイピング手法に該当する。

Code and data availability

This is a methods/protocol preprint describing a Raichu-OsRac1 FRET biosensor in rice protoplasts. The supplied blocks contain no public phenotype/trait datasets, no deposited plant images or sensor data, and no author analysis code, scripts, workflows, or trained models with availability statements or public URLs. The

No evidence-backed public reproduction asset is currently recorded.