Increased benzoxazinoid (Bx) levels in wheat seedlings via jasmonic acid treatment and etiolation and their effects on Bx genes including Bx6

Wheat accumulates benzoxazinoid (Bx) as a defensive compound. While Bx occurs at high concentrations, particularly in the early growth stages, its mechanism of regulation remains unclear. In the present study, we first examined the effects of several plant hormones on Bx concentrations in wheat seedlings. Among the compounds tested, jasmonate (JA) elevated the concentrations of DIMBOA-Glc (2-β-D-glucoside of 2,4-dihydroxy-7-methoy-1,4-benzoxazin-3-one), the primary Bx species in intact wheat seedlings, without a significant increase in HDMBOA-Glc (4-O-methyl-DIMBOA-Glc), which is known to be upregulated by stresses. In addition, growing the plants in the dark increased DIMBOA-Glc levels. Quantification of the Bx-biosynthetic genes showed that TaBx8 (UDP-Glc:Bx glucosyltrasferase) was influenced by neither JA nor etiolation, indicating that TaBx8 is under the regulation mechanism distinct from the mechanisms influencing the others. In addition, none of the other gene expression patterns exhibited considerable correlation with DIMBOA-Glc accumulation. Since there was no correlation between transcript levels of the genes involved in Bx biosynthesis and Bx accumulation, other factors may control the levels of Bx in wheat. In the course of gene analyses, we isolated TaBx6, one of the last two genes that had not been identified in wheat in the DIMBOA-Glc biosynthetic pathway. All the four TaBx6 genes cloned in the present study were expressed in Escherichia coli and characterized their activity.