Joaquim V Marques, Doralyn S Dalisay, Hong Yang, Choonseok Lee, Laurence B Davin, Norman G Lewis

Podophyllum hexandrum and, to a much lesser extent P. peltatum, are sources of podophyllotoxin, extensively used as a chemical scaffold for various anti-cancer drugs. In this study, integrated omics technologies (including advanced mass spectrometry/metabolomics, transcriptome sequencing/gene assemblies, and bioinformatics) gave unequivocal evidence that both plant species possess a hitherto unknown aporphine alkaloid metabolic pathway. Specifically, RNA-seq transcriptome sequencing and bioinformatics guided gene assemblies/analyses in silico suggested presence of transcripts homologous to genes encoding all known steps in aporphine alkaloid biosynthesis. A comprehensive metabolomics analysis, including UPLC-TOF-MS and MALDI-MS imaging in situ, then enabled detection, identification, localization and quantification of the aporphine alkaloids, magnoflorine, corytuberine and muricinine, in the underground and aerial tissues. Interestingly, the purported presence of alkaloids in Podophyllum species has been enigmatic since the 19th century, remaining unresolved until now. The evolutionary and phylogenetic ramifications of this discovery are discussed.