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Increase in nervonic acid content in transformed yeast and transgenic plants by introduction of a Lunaria annua L. 3-ketoacyl-CoA synthase (KCS) gene

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Abstract

Nervonic acid is a Very Long-Chain Monounsaturated Fatty Acid (VLCMFA), 24:1 Δ15 (cis-tetracos-15-enoic acid) found in the seed oils of Lunaria annua, borage, hemp, Acer (Purpleblow maple) and Tropaeolum speciosum (Flame flower). However, of these, only the “money plant” (Lunaria annua L.) has been studied and grown sparingly for future development as a niche crop and the outlook has been disappointing. Therefore, our goal was to isolate and characterize strategic new genes for high nervonic acid production in Brassica oilseed crops. To this end, we have isolated a VLCMFA-utilizing 3-Keto-Acyl-CoA Synthase (KCS; fatty acid elongase; EC 2.3.1.86) gene from Lunaria annua and functionally expressed it in yeast, with the recombinant KCS protein able to catalyze the synthesis of several VLCMFAs, including nervonic acid. Seed-specific expression of the Lunaria KCS in Arabidopsis resulted in a 30-fold increase in nervonic acid proportions in seed oils, compared to the very low quantities found in the wild-type. Similar transgenic experiments using B. carinata as the host resulted in a 7–10 fold increase in seed oil nervonic acid proportions. KCS enzyme activity assays indicated that upon using 14C-22:1-CoA as substrate, the KCS activity from developing seeds of transgenic B. carinata was 20–30-fold higher than the low erucoyl-elongation activity exhibited by wild type control plants. There was a very good correlation between the Lun KCS transcript intensity and the resultant 22:1-CoA KCS activity in developing seed. The highest nervonic acid level in transgenic B. carinata expressing the Lunaria KCS reached 30%, compared to 2.8% in wild type plant. In addition, the erucic acid proportions in these transgenic lines were considerably lower than that found in native Lunaria oil. These results show the functional utility of the Lunaria KCS in engineering new sources of high nervonate/reduced erucic oils in the Brassicaceae.

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Abbreviations

20:1:

cis-Eicosa-11-enoic acid; 20:1Δ11

22:1:

Erucic acid; (cis-docosa-13-enoic acid); 22:1Δ13

24:1:

Nervonic acid; (cis-tetracos-15-enoic acid); 24:1Δ15

FAE:

Fatty acid elongase

FAME:

Fatty acid methyl ester

GC:

Gas chromatography

HEA:

High erucic acid

HPLC:

High performance liquid chromatography

KCS:

3-Keto-acyl-CoA synthase

Lun:

Lunaria

PCR:

Polymerase chain reaction

VLC(M)FA:

Very long-chain (mono-unsaturated) fatty acid

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Acknowledgements

The authors thank B. Panchuk, D. Schwab and Dr. L. Pelcher for DNA sequencing and primer synthesis and Dr. J. Balsevich and Dr. S. R. Abrams for critical reviews of this manuscript. This work was partially supported by a Grant No. 20040417 from the Saskatchewan Agriculture Development Fund, and by the NRC Crops for Enhanced Human Health Program. This National Research Council of Canada Publication No. 50120.

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Author notes

  1. Yiming Guo

    Present address: Department of Biochemistry, University of Missouri, 117 Schweitzer Hall, Columbia, MO, 65211, USA

  2. Elzbieta Mietkiewska

    Present address: Department of Agricultural, Food and Nutritional Science, University of Alberta, 410 Ag/For Building, Edmonton, AB, Canada, T6G 2P5

  3. Vesna Katavic

    Present address: Department of Botany, University of British Columbia, 3529-6270 University Blvd., Vancouver, BC, Canada, V6T 1Z4

  4. Jennifer M. Brost

    Present address: Targeted Growth Canada, 110 Gymnasium Place, Saskatoon, SK, Canada, S7N 0W9

Authors and Affiliations

  1. National Research Council of Canada, Plant Biotechnology Institute, 110 Gymnasium Place, Saskatoon, SK, Canada, S7N 0W9

    Yiming Guo, Elzbieta Mietkiewska, Tammy Francis, Vesna Katavic, Jennifer M. Brost, Michael Giblin, Dennis L. Barton & David C. Taylor

  2. Plant Breeding and Acclimatization Institute, Mlochow Research Center, 05-831, Mlochow, Poland

    Elzbieta Mietkiewska

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  1. Yiming Guo
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Correspondence to David C. Taylor.

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Guo, Y., Mietkiewska, E., Francis, T. et al. Increase in nervonic acid content in transformed yeast and transgenic plants by introduction of a Lunaria annua L. 3-ketoacyl-CoA synthase (KCS) gene. Plant Mol Biol 69, 565–575 (2009). https://doi.org/10.1007/s11103-008-9439-9

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