Substrate promiscuity of acyltransferases contributes to the diversity of hydroxycinnamic acid derivatives in purple coneflower

High pliability and promiscuity are observed widely exist in plant specialized metabolism, especially the hydroxycinnamic acid metabolism. Here, we identified an addition BAHD acyltransferase (EpHMT) that catalyzes phaselic acid biosynthesis and found that the substrate promiscuities of identified BAHD and SCPL acyltransferases are responsible for the diversity of hydroxycinnamic acid derivatives in purple coneflower.Hydroxycinnamic acid derivatives (HADs) such as chicoric acid are the main active ingredients of purple coneflower (Echinacea purpurea (L.) Moench). Recently, the biosynthesis of chicoric acid catalyzed by both BAHD and serine carboxypeptidase-like (SCPL) acyltransferases has been elucidated. However, the diversity of HADs in purple coneflower and their biosynthesis pathways remain unclear. Here, through an alignment of extract ion chromatograms of potential fragments, tentative HADs, such as phaselic acid and chicoric acid analogs (deoxy- and methyl-), were found in purple coneflower. Phylogenetic analyses based on the full-length transcriptome were conducted to explore additional BAHD and SCPL acyltransferases that might be involved in HAD biosynthesis. One BAHD, conserved among Echinacea species, showed weak hydroxycinnamoyl-CoA:tartaric acid hydroxycinnamoyl transferase (HTT) activity. Further results indicated that this BAHD was hydroxycinnamoyl-CoA:malic acid hydroxycinnamoyl transferase (HMT), which catalyzes the biosynthesis of phaselic acid. Although no potential isozyme was found for chicoric acid synthase (CAS), CAS itself showed acyl acceptor promiscuity and catalyzed the biosynthesis of deoxychicoric acid and methylchicoric acid. Overall, we identified additional HADs and depicted their biosynthesis network in purple coneflower. The substrate promiscuity of identified acyltransferases diversifies HADs, indicating the complexity of plant secondary metabolism.