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Cellular and Subcellular Compartmentation of the 2C-Methyl-D-Erythritol 4-Phosphate Pathway in the Madagascar Periwinkle
The Madagascar periwinkle (Catharanthus roseus) synthesizes the highly valuable monoterpene indole alkaloids (MIAs) through a long metabolic route initiated by the 2C-methyl-D-erythritol 4-phosphate (MEP) pathway. In leaves, a complex compartmentation of the MIA biosynthetic pathway occurs at both t...
Autores principales: | , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7238098/ https://www.ncbi.nlm.nih.gov/pubmed/32272573 http://dx.doi.org/10.3390/plants9040462 |
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author | Guirimand, Grégory Guihur, Anthony Perello, Catalina Phillips, Michael Mahroug, Samira Oudin, Audrey Dugé de Bernonville, Thomas Besseau, Sébastien Lanoue, Arnaud Giglioli-Guivarc’h, Nathalie Papon, Nicolas St-Pierre, Benoit Rodríguez-Concepcíon, Manuel Burlat, Vincent Courdavault, Vincent |
author_facet | Guirimand, Grégory Guihur, Anthony Perello, Catalina Phillips, Michael Mahroug, Samira Oudin, Audrey Dugé de Bernonville, Thomas Besseau, Sébastien Lanoue, Arnaud Giglioli-Guivarc’h, Nathalie Papon, Nicolas St-Pierre, Benoit Rodríguez-Concepcíon, Manuel Burlat, Vincent Courdavault, Vincent |
author_sort | Guirimand, Grégory |
collection | PubMed |
description | The Madagascar periwinkle (Catharanthus roseus) synthesizes the highly valuable monoterpene indole alkaloids (MIAs) through a long metabolic route initiated by the 2C-methyl-D-erythritol 4-phosphate (MEP) pathway. In leaves, a complex compartmentation of the MIA biosynthetic pathway occurs at both the cellular and subcellular levels, notably for some gene products of the MEP pathway. To get a complete overview of the pathway organization, we cloned four genes encoding missing enzymes involved in the MEP pathway before conducting a systematic analysis of transcript distribution and protein subcellular localization. RNA in situ hybridization revealed that all MEP pathway genes were coordinately and mainly expressed in internal phloem-associated parenchyma of young leaves, reinforcing the role of this tissue in MIA biosynthesis. At the subcellular level, transient cell transformation and expression of fluorescent protein fusions showed that all MEP pathway enzymes were targeted to plastids. Surprisingly, two isoforms of 1-deoxy-D-xylulose 5-phosphate synthase and 1-deoxy-D-xylulose 5-phosphate reductoisomerase initially exhibited an artifactual aggregated pattern of localization due to high protein accumulation. Immunogold combined with transmission electron microscopy, transient transformations performed with a low amount of transforming DNA and fusion/deletion experiments established that both enzymes were rather diffuse in stroma and stromules of plastids as also observed for the last six enzymes of the pathway. Taken together, these results provide new insights into a potential role of stromules in enhancing MIA precursor exchange with other cell compartments to favor metabolic fluxes towards the MIA biosynthesis. |
format | Online Article Text |
id | pubmed-7238098 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-72380982020-05-28 Cellular and Subcellular Compartmentation of the 2C-Methyl-D-Erythritol 4-Phosphate Pathway in the Madagascar Periwinkle Guirimand, Grégory Guihur, Anthony Perello, Catalina Phillips, Michael Mahroug, Samira Oudin, Audrey Dugé de Bernonville, Thomas Besseau, Sébastien Lanoue, Arnaud Giglioli-Guivarc’h, Nathalie Papon, Nicolas St-Pierre, Benoit Rodríguez-Concepcíon, Manuel Burlat, Vincent Courdavault, Vincent Plants (Basel) Article The Madagascar periwinkle (Catharanthus roseus) synthesizes the highly valuable monoterpene indole alkaloids (MIAs) through a long metabolic route initiated by the 2C-methyl-D-erythritol 4-phosphate (MEP) pathway. In leaves, a complex compartmentation of the MIA biosynthetic pathway occurs at both the cellular and subcellular levels, notably for some gene products of the MEP pathway. To get a complete overview of the pathway organization, we cloned four genes encoding missing enzymes involved in the MEP pathway before conducting a systematic analysis of transcript distribution and protein subcellular localization. RNA in situ hybridization revealed that all MEP pathway genes were coordinately and mainly expressed in internal phloem-associated parenchyma of young leaves, reinforcing the role of this tissue in MIA biosynthesis. At the subcellular level, transient cell transformation and expression of fluorescent protein fusions showed that all MEP pathway enzymes were targeted to plastids. Surprisingly, two isoforms of 1-deoxy-D-xylulose 5-phosphate synthase and 1-deoxy-D-xylulose 5-phosphate reductoisomerase initially exhibited an artifactual aggregated pattern of localization due to high protein accumulation. Immunogold combined with transmission electron microscopy, transient transformations performed with a low amount of transforming DNA and fusion/deletion experiments established that both enzymes were rather diffuse in stroma and stromules of plastids as also observed for the last six enzymes of the pathway. Taken together, these results provide new insights into a potential role of stromules in enhancing MIA precursor exchange with other cell compartments to favor metabolic fluxes towards the MIA biosynthesis. MDPI 2020-04-07 /pmc/articles/PMC7238098/ /pubmed/32272573 http://dx.doi.org/10.3390/plants9040462 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Guirimand, Grégory Guihur, Anthony Perello, Catalina Phillips, Michael Mahroug, Samira Oudin, Audrey Dugé de Bernonville, Thomas Besseau, Sébastien Lanoue, Arnaud Giglioli-Guivarc’h, Nathalie Papon, Nicolas St-Pierre, Benoit Rodríguez-Concepcíon, Manuel Burlat, Vincent Courdavault, Vincent Cellular and Subcellular Compartmentation of the 2C-Methyl-D-Erythritol 4-Phosphate Pathway in the Madagascar Periwinkle |
title | Cellular and Subcellular Compartmentation of the 2C-Methyl-D-Erythritol 4-Phosphate Pathway in the Madagascar Periwinkle |
title_full | Cellular and Subcellular Compartmentation of the 2C-Methyl-D-Erythritol 4-Phosphate Pathway in the Madagascar Periwinkle |
title_fullStr | Cellular and Subcellular Compartmentation of the 2C-Methyl-D-Erythritol 4-Phosphate Pathway in the Madagascar Periwinkle |
title_full_unstemmed | Cellular and Subcellular Compartmentation of the 2C-Methyl-D-Erythritol 4-Phosphate Pathway in the Madagascar Periwinkle |
title_short | Cellular and Subcellular Compartmentation of the 2C-Methyl-D-Erythritol 4-Phosphate Pathway in the Madagascar Periwinkle |
title_sort | cellular and subcellular compartmentation of the 2c-methyl-d-erythritol 4-phosphate pathway in the madagascar periwinkle |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7238098/ https://www.ncbi.nlm.nih.gov/pubmed/32272573 http://dx.doi.org/10.3390/plants9040462 |
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