Modular Design of Picroside-II Biosynthesis Deciphered through NGS Transcriptomes and Metabolic Intermediates Analysis in Naturally Variant Chemotypes of a Medicinal Herb, Picrorhiza kurroa

Picroside-II (P-II), an iridoid glycoside, is used as an active ingredient of various commercial herbal formulations available for the treatment of liver ailments. Despite this, the knowledge of P-II biosynthesis remains scarce owing to its negligence in shoots which sets constant barrier for function validation experiments. In this study, we utilized natural variation for P-II content in stolon tissues of different accessions and deciphered its metabolic route by integrating metabolomics of intermediates with differential NGS transcriptomes. Upon navigating through high vs. low P-II content accessions (1.3-2.6%), we have established that P-II is biosynthesized degradation of ferulic acid (FA) to produce vanillic acid (VA) which acts as its immediate biosynthetic precursor. Moreover, the FA treatment at 150 μM concentration provided further confirmation with 2-fold rise in VA content. Interestingly, the cross-talk between different compartments of , i.e., shoots and stolons, resolved spatial complexity of P-II biosynthesis and consequently speculated the burgeoning necessity to bridge gap between VA and P-II production in shoots. This work thus, offers a forward looking strategy to produce both P-I and P-II in shoot cultures, a step toward providing a sustainable production platform for these medicinal compounds via-à-vis relieving pressure from natural habitat of .