Biochemical and partial molecular characterization of bitter and sweet forms of Lupinus angustifolius, an experimental model for study of molecular regulation of quinolizidine alkaloid biosynthesis.
The bitter and sweet forms of a plant species differing with alkaloid contents may provide a model system for investigation of alkaloid biosynthesis at a molecular level. The pattern and concentration of quinolizidine alkaloids were determined by capillary GC-MS in bitter and sweet plants of Lupinus angustifolius. Bitter plant contained lupanine, 13alpha-hydroxylupanine, angustifoline, alpha-isolupanine, tetrahydrorhombifoline, and ester-derivatives of 13alpha-hydroxylupanine. In contrast, no alkaloid was detected in sweet plant. The enzymatic activity of acyltransferase for formation of 13alpha-tigloyloxylupanine was similar or even higher in the cell-free extracts of sweet plant than that in bitter plant. These results suggest that the biosynthetic step(s) of ring closure forming the initial cyclic alkaloid, lupanine, from cadaverine is presumably blocked in sweet plant, and that the later steps for modification of the cyclized alkaloids are not altered. We hypothesized that the gene(s) encoding enzyme(s) for ring-closure step might be repressed in sweet plant, and that the expression might take place only in bitter plant. To isolate the genes specifically expressed in bitter plant, cDNA-amplified fragment length polymorphism (cDNA-AFLP) analysis was carried out. However, no bitter-specific gene was isolated, suggesting that alkaloid biosynthesis in sweet plant may be down-regulated at a post-transcriptional level.