Tissue- and cell-specific transcriptomics of pyrrolizidine alkaloidbiosynthesis

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The system of pyrrolizidine alkaloids (PAs) is a model for our studies about the evolution of pathways in plant secondary metabolism. PAs are toxic to vertebrates and insects and are believed to be selected as part of the chemical defence against herbivores. For the first pathway-specific enzyme, homospermidine synthase (HSS), we were able to show that it originated by duplication of a gene involved in primary metabolism at least five times independently during angiosperm evolution. This observation suggests that also the complete pathway of PAs has several independent origins. In the present project, we intend to identify three further enzymes of PA biosynthesis to study their evolution and compare that data with the extensive knowledge about HSS evolution. Using our knowledge about the tissue- and cell-specific expression of HSS we successfully generated transcriptome databases of various tissues of several PA-producing species. By comparing the sequence data from different tissues with bioinformatic tools, we were able to denominate sequences that encode enzymes postulated to be involved in PA biosynthesis. We will use this second phase of the project to prove the functional role of these candidate sequences in PA biosynthesis by comparing their biochemical properties with paralogous sequences that are also available in the transcriptome databases. This data in addition to phylogenetic analyses should allow conclusions about the evolution of these enzymes of PA biosynthesis.