Horse Chestnut Tree Genome Reveals the evolutionary mechanism of Aescin and Aesculin biosynthesis

Abstract

Abstract Medicinal trees provide a main resource for diverse medicinal compounds. However, the biosynthesis of tree metabolites and their pathway evolution has gained limited understanding. Horse chestnut (Aesculus chinensis) is an important medicinal tree and its seeds are rich in aescins, barrigenol-type triterpenoid saponins (BAT), and aesculin, a coumarin glycoside, which are effective in the therapy of chronic venous insufficiency and asthenopia (eye strain). To understand the biosynthesis of these compounds, herein, we assembled a 470.04-Mb high-quality horse chestnut genome and characterized an Aesculus-specific whole-genome duplication (WGD) event. Spatial metabolome imaging, co-expression, and biosynthetic gene cluster analyses indicated that the Aesculus-specific WGD event led to the formation of two gene clusters (BGCs) including oxidosqualene cyclase, cytochrome P450 monooxygenase, cellulose synthase-derived glycotransferases, and BADH acyltransferases. Further biochemical investigation revealed the roles of AcOCS6, AcCYP716A278, AcCYP716A275, AcCSL1, and AcBAHD3 genes distributed between these two BGCs in catalyzing the formation of aescins. To understand the evolution of BAT pathways, the collinearity analysis showed the collinear BGC segments could be traced back to early-diverging angiosperms, then the essential gene-encoding enzymes necessary for the BAT biosynthesis were recruited before the split of Aesculus, Acer, and Xanthoceras. Meanwhile, we identified three UDP-glucosyltransferases and demonstrated their involvement in the biosynthesis of aesculin via a de novo synthesis. Taken together, these findings provide important information in understanding the evolution of gene clusters associated with medicinal tree metabolites.