A Trichome-Specific, Plastid-Localized Tanacetum cinerariifolium Nudix Protein Hydrolyzes the Natural Pyrethrin Pesticide Biosynthetic Intermediate trans-Chrysanthemyl Diphosphate.

Wei Li,Robert L Last,Daniel B Lybrand,Haiyang Xu,Fei Zhou,Eran Pichersky

doi:10.3389/fpls.2020.00482

Abstract

Tanacetum cinerariifolium flowers synthesize six pyrethrins that function as effective insecticides. trans-Chrysanthemol is an early intermediate in the synthesis of the monoterpene moiety of pyrethrins. Previously, the pyrethrum enzyme chrysanthemyl diphosphate synthase (TcCDS) was shown to catalyze the formation of the prenyl diphosphate compound chrysanthemyl diphosphate (CPP) by condensing two molecules of dimethylallyl diphosphate (DMAPP). Later work also showed that with a low concentration of DMAPP, TcCDS can also remove the diphosphate group to give chrysanthemol. The removal of the phosphate groups from other prenyl diphosphates, such as DMAPP, isopentenyl diphosphate (IPP) and geranyl diphosphate (GPP), was previously shown to occur in two steps. In those cases, the first phosphate group is removed by a member of the Nudix hydrolase protein family, and the second by other unidentified phosphatases. These previously characterized Nudix proteins involved in the hydrolysis of prenyl diphosphates were shown to be cytosolic. Here we report that a plastidic Nudix protein from pyrethrum, designated TcNudix1, has high specificity for CPP and can hydrolyze it to chrysanthemol monophosphate (CMP). TcNudix1 is expressed specifically in the trichomes of the ovaries, where chrysanthemol is produced. TcNudix1 expression patterns and pathway reconstitution experiments presented here implicate the TcNudix1 protein in the biosynthesis of chrysanthemol.

Highlights

Prenyl diphosphates are intermediates in the synthesis of terpenoid compounds (Pichersky and Raguso, 2018; Karunanithi and Zerbe, 2019)
The two previously identified Nudix proteins involved in terpenoid metabolism – a geranyl diphosphate (GPP) monohydrolase in rose and an isopentenyl diphosphate (IPP) mono-phosphate hydrolase in Arabidopsis – were shown to be cytosolically localized
Chrysanthemyl diphosphate is an intermediate in the synthesis of chrysanthemic and pyrethric acids, which are combined with jasmone-derived alcohols in the biosynthesis of the natural pyrethrin insecticides (Rivera et al, 2001)

Summary

INTRODUCTION

Prenyl diphosphates are intermediates in the synthesis of terpenoid compounds (Pichersky and Raguso, 2018; Karunanithi and Zerbe, 2019). The prenyl moiety of each prenyl diphosphate consists of either a single five-carbon isoprene unit or a linear polymer of this isoprene building block (Figure 1). These prenyl diphosphates serve as terpene synthase substrates, which remove. TcNudix Hydrolase in Pyrethrins Synthesis by a cytosolic monophosphatase, which is a member of the Nudix (nucleoside diphosphate linked to other moieties X) hydrolase family (Mildvan et al, 2005), followed by the hydrolysis of the second phosphate in a reaction catalyzed by another phosphatase (Magnard et al, 2015). We report that a plastidic Nudix protein in pyrethrum, designated TcNudix, has high specificity to CPP and is capable of hydrolyzing it to chrysanthemol monophosphate (CMP). TcNudix expression patterns and pathway reconstitution experiments presented here provide evidence for the involvement of the TcNudix protein in chrysanthemol biosynthesis

RESULTS

DISCUSSION

Findings

MATERIALS AND METHODS