The regulatory function of isoprenyl diphosphate synthases in terpene biosynthesis
Terpenes are essential secondary metabolites they are important defensive compounds of plants and insects. The biosynthesis of terpenes starts with the formation isopentenyl diphosphate (IDP) and dimethylallyl diphosphate (DMADP). In the next step of biosynthesis, isoprenyl diphosphate synthases (IDS) condense DMADP with a varying number of IDP molecules to produce longer prenyl diphosphates, like geranyl- (GDP), farnesyl- (FDP) and geranylgeranyl diphosphate (GGDP), which are substrates for terpene synthases that produce the various terpene skeletons. IDS have a critical role in terpene biosynthesis since they are located at a central point where the pathway branches towards products of different sizes. The published findings of the present dissertation begin with the description of a new LC-MS/MS based method for identification of the product specificity of IDS and the quantification of total IDS activity. This was the methodical basis for investigations on the regulatory function of IDS1 after overexpression in Picea glauca. Here it was shown that overexpression is only functional in needles. Overexpression increases the total IDS-activity. Despite the increases in terpene precursors the defensive oleoresin was unaltered. Instead large amounts of geranylgeranyl fatty acid esters were formed that were shown to have defensive properties against the needle feeding insect Lymantria monacha. Also insect employ terpenes to defend themselve. One example here is the horseradish leaf beetle Phaedon cochleariae, which has pairs of dorsal glands, that excrete the terpene chrysomelidial in case of an attack. Here IDS1 was identified to produce either GDP or FDP, depending on the metal ion cofactor. With an RNAi approach it was revealed that IDS1 is indeed a key enzyme in chrysomelidial biosynthesis. RNAi mediated transcript suppression, reduced the total IDS enzyme activity, the amount of intermediates in the larvae and the volume of the defensive secretions.