Characterization of defensive cadinenes and a novel sesquiterpene synthase responsible for their biosynthesis from the invasive Eupatorium adenophorum

Eupatorium adenophorum is a malignant invasive plant possessing extraordinary defense potency, but its chemical weaponry and formation mechanism have been rarely investigated. We identified six cadinene sesquiterpenes including two volatiles (amorpha-4,7(11)-diene and (-)-amorph-4-en-7-ol) and four non-volatiles (9-oxo-10,11-dehydroageraphorone, muurol-4-en-3,8-dione, 9-oxo-ageraphorone and 9β-hydroxy-ageraphorone) as the major constitutive and inducible chemicals of E. adenophorum. All cadinenes showed potent antifeedant activity against a generalist insect Spodoptera exigua, indicating their significant defensive roles. We cloned and functionally characterized a sesquiterpene synthase from E. adenophorum (EaTPS1) catalyzing the conversion of farnesyl diphosphate to amorpha-4,7(11)-diene and (-)-amorph-4-en-7-ol that were purified from engineered Escherichia coli and identified by extensive NMR spectroscopy. EaTPS1 was highly expressed in the above-ground organs, which was congruent with the dominant distribution of cadinenes, suggesting that EaTPS1 is likely involved in cadinene biosynthesis. Mechanical wounding and methyl jasmonate negatively regulated EaTPS1 expression but caused the release of amorpha-4,7(11)-diene and (-)-amorph-4-en-7-ol. Nicotiana benthamiana transiently expressing EaTPS1 also produced amorpha-4,7(11)-diene and (-)-amorph-4-en-7-ol, and showed enhanced defense function. The findings uncovered the chemical defense role and formation mechanism of E. adenophorum that probably contribute to invasive success of this plant, and also provided a tool for manipulating the biosynthesis of biologically active cadinene natural products.

Keywords: Eupatorium adenophorum; cadinenes; defensive chemicals; sesquiterpene synthase; sesquiterpenes.