Rho GTPase Activity Analysis in Plant Cells

Rho-family small GTPases are conserved molecular switches of signaling networks in eukaryotic cells, and regulate many cellular responses, such as cytoskeletal reorganization, gene expression, and polarized vesicular trafficking. To understand the functions of Rho GTPase, it is important to investigate how the activity of Rho GTPase is regulated. Plant Rho-like GTPases (ROPs) are known to be regulated by hormones and environmental cues. A rapid activation of ROPs by a stimulus implies a direct signaling role for the ROP GTPases. Here, I describe an ROP activity assay that allows the measurement of ROP GTPase activity that occurs within seconds upon treatment with a stimulus. This method has been successfully used to investigate auxin activation of ROPs in plant cells and will be generally useful for measuring changes in ROP activity with high time resolution (Xu et al., Cell 143:99–110, 2010).