Broccoli Cytochrome P450 Genes Associated with Indole Glucosinolate Biosynthesis

Broccoli (Brassica oleracea L. ssp.. Italica) is a rich source of indolyl glucosinolates (glucobrassicin and neoglucobrassicin), phytochemicals that are hydrolyzed into isothiocyanates with known anti-carcinogenic bioactivity.  Information on glucosinolate metabolism in crucifer plant species is incomplete in its understanding of how glucosinolates accumulate, and how they are transported and undergo catabolic degradation during the plant's life cycle.  The dramatic increase in the amounts of these indolyl glucosinolates in broccoli induced in response to treatments with methyl jasmonate (MeJA) provides a system to investigate the synthesis and flux of glucosinolates and associated changes in gene expression patterns.  Genes involved with the biosynthesis of glucosinolates (CYP79B2, CYP79F1, CYP83A1 and CYP83B1) were isolated from broccoli using primer sets based on Arabidopsis sequences, which averaged 88 % sequence homology with corresponding Arabidopsis transcripts.  Expression levels of these and two other genes (myrosinase and epithiospecifier protein), involved in the hydrolysis of glucosinolates, were analyzed in different broccoli plant tissues (leaf, stem and immature florets) at two different developmental stages by quantitative RT-PCR.  Glucosinolate concentrations were also profiled in the same tissues with or without MeJA treatment to determine correlations between metabolite profiling and gene expression data.  Expression levels of the four broccoli P450 genes differentially responded to MeJA treatment with upregulation of BoCYP79B2 and BoCYP83A1 correlating with increased indolyl glucosinolate concentrations in broccoli tissues.  This suggests that these two genes may play a key role in regulating indoyl glucosinolate accumulation in broccoli tissues.