Poster: Plant-pathogen interactions

Abs # 544: Dual role of the small GTPase OsRac1 as an inducer of ROS and a suppressor of ROS scavenger

Presenter:	Wong, Hann Ling , h-wong@bs.naist.ac.jp
Authors	Wong, Hann Ling  (A)   Pinontoan, Reinhard  (A)   Sakamoto, Tsuyoshi  (A)   Kawasaki, Tsutomu  (A)   Shimamoto, Ko  (A)
Affiliations:	(A): Nara Institute of Science and Technology (NAIST)

Reactive oxygen species (ROS) produced by NADPH oxidase are thought to play critical roles in the plant defense against pathogens.In rice, we found that overexpression of the constitutively active form of the small GTPase OsRac1 enhanced resistance to virulent rice fungus and bacterial blight, and enhanced elicitor-induced hydrogen peroxide (H₂O₂) production. Using yeast two-hybrid assays we found that the constitutively active form of OsRac1, but not the dominant negative form, interacted with OsRBOH-B, the rice homolog of the gp91phox subunit of the human neutrophil NADPH oxidase complex. The N-terminus of OsRBOH-B, which contained two Ca²⁺-binding EF-hands domains, was sufficient for the OsRac1-OsRBOH-B interaction. This suggests that unlike in the human NADPH oxidase, plant Rac directly interacts with RBOH, to regulate ROS production. Presently, we are studying the OsRac1-RBOH interaction using a single molecular mutant GFP-tagged fluorescence resonance energy transfer (FRET) system. In addition, we found that the expression of a metallothionein gene OsMT2b was synergically downregulated by OsRac1 and rice blast-derived elicitors. Elicitor-induced H₂O₂ production was reduced in OsMT2b-overexpressing cells. In contrast, homozygous OsMT2b::Tos17-inserted mutant and OsMT2b-RNAi-silenced transgenic cells showed significantly enhanced elicitor-induced H₂O₂ production. In vitro assays using recombinant OsMT2b fusion protein showed that OsMT2b is an ROS scavenger. Taken together, the results suggest that OsRac1 plays a dual role as an inducer of ROS and a suppressor of ROS scavenger.