Production of wax esters in plant seed oils by oleosomal cotargeting of biosynthetic enzymes

Mareike Heilmann,Tim Iven,Katharina Ahmann,Ellen Hornung,Sten Stymne,Ivo Feussner

doi:10.1194/jlr.m029512

Mareike Heilmann, Tim Iven + Show 4 more

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https://doi.org/10.1194/jlr.m029512

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Abstract

Wax esters are neutral lipids exhibiting desirable properties for lubrication. Natural sources have traditionally been whales. Additionally some plants produce wax esters in their seed oil. Currently there is no biological source available for long chain length monounsaturated wax esters that are most suited for industrial applications. This study aimed to identify enzymatic requirements enabling their production in oilseed plants. Wax esters are generated by the action of fatty acyl-CoA reductase (FAR), generating fatty alcohols and wax synthases (WS) that esterify fatty alcohols and acyl-CoAs to wax esters. Based on their substrate preference, a FAR and a WS from Mus musculus were selected for this study (MmFAR1 and MmWS). MmWS resides in the endoplasmic reticulum (ER), whereas MmFAR1 associates with peroxisomes. The elimination of a targeting signal and the fusion to an oil body protein yielded variants of MmFAR1 and MmWS that were cotargeted and enabled wax ester production when coexpressed in yeast or Arabidopsis. In the fae1 fad2 double mutant, rich in oleate, the cotargeted variants of MmFAR1 and MmWS enabled formation of wax esters containing >65% oleyl-oleate. The data suggest that cotargeting of unusual biosynthetic enzymes can result in functional interplay of heterologous partners in transgenic plants.

Highlights

Wax esters are neutral lipids exhibiting desirable properties for lubrication
To test whether this localization pattern would hold true for plant cells, MmFAR1 was transiently expressed in onion epidermal cells as a fusion to mCherry, and the fluorescence distribution was monitored (Fig. 1A)
The mCherry fluorescence decorated punctate structures that colocalized with EYFPtagged peroxisomal malate dehydrogenase, a peroxisomal marker protein [13](Fig. 1A), confirming peroxisomal localization of mCherry:MmFAR1 in onion cells

Summary

Introduction

Wax esters are neutral lipids exhibiting desirable properties for lubrication. Natural sources have traditionally been whales. Some plants produce wax esters in their seed oil. Wax esters are generated by the action of fatty acyl-CoA reductase (FAR), generating fatty alcohols and wax synthases (WS) that esterify fatty alcohols and acyl-CoAs to wax esters Based on their substrate preference, a FAR and a WS from Mus musculus were selected for this study (MmFAR1 and MmWS). The elimination of a targeting signal and the fusion to an oil body protein yielded variants of MmFAR1 and MmWS that were cotargeted and enabled wax ester production when coexpressed in yeast or Arabidopsis. In the fae fad double mutant, rich in oleate, the cotargeted variants of MmFAR1 and MmWS enabled formation of wax esters containing >65% oleyl-oleate. Production of wax esters in plant seed oils by oleosomal cotargeting of biosynthetic enzymes.

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