Proteomic analysis of rice defense response induced by probenazole

Here, we report the first proteomic analysis of rice defense response induced by probenazole (PBZ), an agricultural chemical that has been widely used to protect rice plants from rice blast and the bacterial blight pathogen. Two-dimensional gel electrophoresis (2-DE) was utilized to identify a total of 40 protein spots including 9 protein spots that are up-regulated by PBZ and 31 abundant protein spots. A total of 11 unique proteins from these 9 spots were identified by LC-MS/MS, and the majority of them were classified and/or possessed orthologs in defense-related functions. Five protein spots with only one protein species identified in each spot appear to be PBZ-regulated proteins. They are a putative glutathione S-transferase GSTU17, a putative phenylalanine ammonia-lyase (PAL, XP_466843), a putative caffeic acid 3-O-methyltransferase (COMT), a putative NADH-ubiquinone oxidoreductase, and a putative glucose-1-phosphate adenyltransferase. However, the other six protein species identified from the remaining four protein spots could not be conclusively described as PBZ-regulated proteins due to either the co-migration of two protein species in one spot or the presence of one protein species in two spots. Through real-time reverse transcription polymerase chain reaction (RT-PCR), it was determined that PAL (XP_466843) is likely regulated at the protein level, whereas GSTU17 and COMT were regulated at the mRNA level after PBZ application. Interestingly, the mRNA transcripts of two PAL paralogs were found to be up-regulated by PBZ. We propose that PAL, COMT, and GSTU17 are likely to confer PBZ-induced disease resistance via such functions as biosynthesis and transport of flavonoid-type phytoalexin and/or lignin biogenesis.