Role of jasmonate in the rice (Oryza sativa L.) self-defense mechanism using proteome analysis

Abstract

Exogenously applied jasmonic acid (JA) was used to study changes in protein patterns in rice (Oryza sativa L.) seedling tissues, to classify these changes, and to assign a role for these changes, in order to define the role of JA in the rice self-defense mechanism. High resolution two-dimensional polyacrylamide gel electrophoretic analysis revealed induction of new proteins in both leaf and stem tissues after JA treatment, with the major protein spots further analyzed through N-terminal and internal amino acid sequencing, purification, antibody production, and immunoblot analysis. JA treatment results in necrosis in these tissues, which is accompanied by drastic reductions in ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) subunits, and was confirmed using immunoblotting. Induction of novel proteins was found particularly in the stem tissues, including a new basic 28 kDa Bowman-Birk proteinase inhibitor protein (BBPIN; jasmonate-induced stem protein, JISP 6), and acidic 17 kDa pathogenesis-related class 1 protein (PR-1, JISP 9). This induction of proteins was blocked by a protein synthesis inhibitor cycloheximide (CHX) indicating de novo protein synthesis. Kinetin (KIN), a cytokinin and free radical scavenger reversed RuBisCO decreases, but not induction of proteins. Immunoblot analysis using antibodies generated against these purified proteins revealed a tissue-specific expression pattern and time-dependent induction after JA treatment. Our results indicate that jasmonate affects defense-related gene expression in rice seedlings, as evidenced by de novo synthesis of novel proteins with potential roles in plant defense.