Investigation into the role of DNA methylation in cyanogenesis in sorghum (Sorghum bicolor L. Moench)

Sorghum bicolor produces the cyanogenic glucoside dhurrin, a secondary metabolite integral to plant defence and stress responses. Dhurrin production is both developmentally and environmentally regulated in S. bicolor, with high levels of variation within and between lines. Such phenotypic variation may result from polymorphic differences or epigenetic modifications in genes associated with cyanogenesis. In this study the chemical 5-Azacytidine was used to assess S. bicolor's response to genome-wide demethylation, which had not previously been investigated in the context of cyanogenic glucoside regulation. Morphological changes, the expression levels of key genes involved in dhurrin synthesis and turnover, and the cyanogenic potential (HCNp) of leaf tissues were analysed. Treatment resulted in alterations in dhurrin synthesis, gene expression, and dhurrin levels, suggesting that DNA methylation is involved in the regulation of HCNp in the initial stages of S. bicolor development. Previously identified EMS mutants from the adult cyanide deficient class (acdc) have been found to exhibit altered dhurrin concentrations during development. This study shows that acdc mutants possess a CT change in the promoter of CYP79A1, a key gene in dhurrin synthesis, and that this mutation is stably inherited and associated with the acdc phenotype. To further investigate the role of epigenesis in dhurrin production, we determine the methylation status of the 250bp region surrounding the CT mutation site in wild-type and mutant plants over two stages of development.