A multifunctional sesquiterpene synthase integrates with cytochrome P450s to reinforce the terpenoid defense network in maize

TL;DR

This paper identifies a new defense pathway in maize involving a terpene synthase and P450 enzymes that produce antimicrobial compounds.

Contribution

The study reveals a previously unknown sesquiterpenoid defense pathway in maize centered on α-santalenoic acid.

Findings

ZmTPS9 is a multiproduct terpene synthase that produces α-santalene and β-bisabolene.

Three cytochrome P450 enzymes oxidize ZmTPS9 products into antimicrobial metabolites like α-santalenoic acid and zealexins.

The pathway represents a convergent biosynthetic strategy for maize's terpenoid defense network.

Abstract

Terpenoids, the largest and most structurally diverse class of plant natural products, play essential roles in maize defense and ecological interactions. In this study, we identified and functionally characterized a sesquiterpenoid‐based defense pathway in maize centered on α‐santalenoic acid, a pathogen‐inducible sesquiterpenoid antibiotic. Using a combination of metabolite‐based genome‐wide association studies (mGWAS), linkage mapping, and heterologous expression assays, we identified ZmTPS9 as a multiproduct terpene synthase that primarily produces α‐santalene and β‐bisabolene. Sequence analysis and site‐directed mutagenesis revealed that threonine at position 413 is critical for enzyme activity, with its deletion resulting in a complete loss of enzyme activity. The sesquiterpene hydrocarbons produced by ZmTPS9 are further oxidized by three cytochrome P450 monooxygenases, ZmCYP71Z16, ZmCYP71Z18, and ZmCYP71Z19, to yield antimicrobial metabolites including α‐santalenoic acid, zealexin D1 (ZD1), and zealexin D2 (ZD2). Together, these findings demonstrate a convergent biosynthetic strategy in maize, where multiproduct terpene synthases and promiscuous P450s collaboratively generate a flexible and robust terpenoid defense network.

This study uncovers a previously unknown maize sesquiterpenoid defense pathway centered on the antibiotic α‐santalenoic acid. Using integrated genetic and biochemical approaches, we identified ZmTPS9 as a pathogen‐inducible terpene synthase responsible for producing multiple sesquiterpenes, primarily α‐santalene and β‐bisabolene. These olefin precursors are further oxidized by three cytochrome P450 enzymes into bioactive metabolites, including α‐santalenoic acid and zealexins. Our findings highlight a convergent biosynthetic strategy underpinning maize's versatile and robust terpenoid defense network.

Figures