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Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes
BACKGROUND: Sesquiterpene lactones are characteristic metabolites of Asteraceae (or Compositae) which often display potent bioactivities and are sequestered in specialized organs such as laticifers, resin ducts, and trichomes. For characterization of sunflower sesquiterpene synthases we employed a s...
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Formato: | Texto |
Lenguaje: | English |
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BioMed Central
2009
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2715020/ https://www.ncbi.nlm.nih.gov/pubmed/19580670 http://dx.doi.org/10.1186/1471-2229-9-86 |
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author | Göpfert, Jens C MacNevin, Gillian Ro, Dae-Kyun Spring, Otmar |
author_facet | Göpfert, Jens C MacNevin, Gillian Ro, Dae-Kyun Spring, Otmar |
author_sort | Göpfert, Jens C |
collection | PubMed |
description | BACKGROUND: Sesquiterpene lactones are characteristic metabolites of Asteraceae (or Compositae) which often display potent bioactivities and are sequestered in specialized organs such as laticifers, resin ducts, and trichomes. For characterization of sunflower sesquiterpene synthases we employed a simple method to isolate pure trichomes from anther appendages which facilitated the identification of these genes and investigation of their enzymatic functions and expression patterns during trichome development. RESULTS: Glandular trichomes of sunflower (Helianthus annuus L.) were isolated, and their RNA was extracted to investigate the initial steps of sesquiterpene lactone biosynthesis. Reverse transcription-PCR experiments led to the identification of three sesquiterpene synthases. By combination of in vitro and in vivo characterization of sesquiterpene synthase gene products in Escherichia coli and Saccharomyces cerevisiae, respectively, two enzymes were identified as germacrene A synthases, the key enzymes of sesquiterpene lactone biosynthesis. Due to the very low in vitro activity, the third enzyme was expressed in vivo in yeast as a thioredoxin-fusion protein for functional characterization. In in vivo assays, it was identified as a multiproduct enzyme with the volatile sesquiterpene hydrocarbon δ-cadinene as one of the two main products with α-muuorlene, β-caryophyllene, α-humulene and α-copaene as minor products. The second main compound remained unidentified. For expression studies, glandular trichomes from the anther appendages of sunflower florets were isolated in particular developmental stages from the pre- to the post-secretory phase. All three sesquiterpene synthases were solely upregulated during the biosynthetically active stages of the trichomes. Expression in different aerial plant parts coincided with occurrence and maturity of trichomes. Young roots with root hairs showed expression of the sesquiterpene synthase genes as well. CONCLUSION: This study functionally identified sesquiterpene synthase genes predominantly expressed in sunflower trichomes. Evidence for the transcriptional regulation of sesquiterpene synthase genes in trichome cells suggest a potential use for these specialized cells for the identification of further genes involved in the biosynthesis, transport, and regulation of sesquiterpene lactones. |
format | Text |
id | pubmed-2715020 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2009 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-27150202009-07-24 Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes Göpfert, Jens C MacNevin, Gillian Ro, Dae-Kyun Spring, Otmar BMC Plant Biol Research Article BACKGROUND: Sesquiterpene lactones are characteristic metabolites of Asteraceae (or Compositae) which often display potent bioactivities and are sequestered in specialized organs such as laticifers, resin ducts, and trichomes. For characterization of sunflower sesquiterpene synthases we employed a simple method to isolate pure trichomes from anther appendages which facilitated the identification of these genes and investigation of their enzymatic functions and expression patterns during trichome development. RESULTS: Glandular trichomes of sunflower (Helianthus annuus L.) were isolated, and their RNA was extracted to investigate the initial steps of sesquiterpene lactone biosynthesis. Reverse transcription-PCR experiments led to the identification of three sesquiterpene synthases. By combination of in vitro and in vivo characterization of sesquiterpene synthase gene products in Escherichia coli and Saccharomyces cerevisiae, respectively, two enzymes were identified as germacrene A synthases, the key enzymes of sesquiterpene lactone biosynthesis. Due to the very low in vitro activity, the third enzyme was expressed in vivo in yeast as a thioredoxin-fusion protein for functional characterization. In in vivo assays, it was identified as a multiproduct enzyme with the volatile sesquiterpene hydrocarbon δ-cadinene as one of the two main products with α-muuorlene, β-caryophyllene, α-humulene and α-copaene as minor products. The second main compound remained unidentified. For expression studies, glandular trichomes from the anther appendages of sunflower florets were isolated in particular developmental stages from the pre- to the post-secretory phase. All three sesquiterpene synthases were solely upregulated during the biosynthetically active stages of the trichomes. Expression in different aerial plant parts coincided with occurrence and maturity of trichomes. Young roots with root hairs showed expression of the sesquiterpene synthase genes as well. CONCLUSION: This study functionally identified sesquiterpene synthase genes predominantly expressed in sunflower trichomes. Evidence for the transcriptional regulation of sesquiterpene synthase genes in trichome cells suggest a potential use for these specialized cells for the identification of further genes involved in the biosynthesis, transport, and regulation of sesquiterpene lactones. BioMed Central 2009-07-06 /pmc/articles/PMC2715020/ /pubmed/19580670 http://dx.doi.org/10.1186/1471-2229-9-86 Text en Copyright © 2009 Göpfert et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( (http://creativecommons.org/licenses/by/2.0) ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Article Göpfert, Jens C MacNevin, Gillian Ro, Dae-Kyun Spring, Otmar Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes |
title | Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes |
title_full | Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes |
title_fullStr | Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes |
title_full_unstemmed | Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes |
title_short | Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes |
title_sort | identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2715020/ https://www.ncbi.nlm.nih.gov/pubmed/19580670 http://dx.doi.org/10.1186/1471-2229-9-86 |
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