Towards Elucidating Carnosic Acid Biosynthesis in Lamiaceae: Functional Characterization of the Three First Steps of the Pathway in Salvia fruticosa and Rosmarinus officinalis

Autor: Božić, Dragana, Papaefthimiou, Dimitra, Brückner, Kathleen, Vos, Ric C.H. de, Tsoleridis, Constantinos A., Katsarou, Dimitra, Papanikolaou, Antigoni, Pateraki, Irene, Chatzopoulou, Fani M., Dimitriadou, Eleni, Kostas, Stefanos, Manzano, David, Scheler, Ulschan, Ferrer, Albert, Tissier, Alain, Makris, Antonios M., Kampranis, Sotirios C., Kanellis, Angelos K.
Přispěvatelé: Universitat de Barcelona, European Commission, Netherlands Genomics Initiative
Rok vydání: 2015
Předmět:
0106 biological sciences
Labiades
Leaves
lcsh:Medicine
01 natural sciences
Biochemistry
Rosmarinus
chemistry.chemical_compound
Salvia fruticosa
subcellular-localization
Salvia
Cloning
Molecular
lcsh:Science
Plant Proteins
0303 health sciences
Multidisciplinary
biology
Trichomes
Polymerase chain reaction
Ferruginol
creticus subsp creticus
diphosphate synthase
BIOS Applied Metabolic Systems
Gas chromatography-mass spectrometry
Research Article
Biosíntesi
Bioquímica
Biosynthesis
l
Copalyl diphosphate synthase
03 medical and health sciences
expression
Hexanes
030304 developmental biology
Gas-chromatography-mass-spectrometry
Yeast
Sequence motif analysis
Alkyl and Aryl Transferases
Lamiaceae
Sequence Analysis
RNA
Gene Expression Profiling
lcsh:R
Carnosic acid
biology.organism_classification
gene-cluster
Plant Leaves
in-vitro antioxidant
chemistry
Abietanes
biology.protein
phenolic diterpenes
lcsh:Q
rosemary
Diterpene
metabolism
010606 plant biology & botany
Zdroj: PLOS ONE
PLoS ONE 10 (2015) 5
Recercat. Dipósit de la Recerca de Catalunya
instname
PLoS ONE, 10(5)
PLOS ONE, 10(5):e0124106
Dipòsit Digital de Documents de la UAB
Universitat Autònoma de Barcelona
Dipòsit Digital de la UB
Universidad de Barcelona
PLoS ONE, Vol 10, Iss 5, p e0124106 (2015)
PLoS ONE
Digital.CSIC. Repositorio Institucional del CSIC
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0124106
Popis: Carnosic acid (CA) is a phenolic diterpene with anti-tumour, anti-diabetic, antibacterial and neuroprotective properties that is produced by a number of species from several genera of the Lamiaceae family, including Salvia fruticosa (Cretan sage) and Rosmarinus officinalis (Rosemary). To elucidate CA biosynthesis, glandular trichome transcriptome data of S. fruticosa were mined for terpene synthase genes. Two putative diterpene synthase genes, namely SfCPS and SfKSL, showing similarities to copalyl diphosphate synthase and kaurene synthase-like genes, respectively, were isolated and functionally characterized. Recombinant expression in Escherichia coli followed by in vitro enzyme activity assays confirmed that SfCPS is a copalyl diphosphate synthase. Coupling of SfCPS with SfKSL, both in vitro and in yeast, resulted in the synthesis miltiradiene, as confirmed by 1D and 2D NMR analyses (1H, 13C, DEPT, COSY H-H, HMQC and HMBC). Coupled transient in vivo assays of SfCPS and SfKSL in Nicotiana benthamiana further confirmed production of miltiradiene in planta. To elucidate the subsequent biosynthetic step, RNA-Seq data of S. fruticosa and R. officinalis were searched for cytochrome P450 (CYP) encoding genes potentially involved in the synthesis of the first phenolic compound in the CA pathway, ferruginol. Three candidate genes were selected, SfFS, RoFS1 and RoFS2. Using yeast and N. benthamiana expression systems, all three where confirmed to be coding for ferruginol synthases, thus revealing the enzymatic activities responsible for the first three steps leading to CA in two Lamiaceae genera.
This work was supported by EU Grant agreement KBBE-227448 for the TERPMED project (http://www.terpmed.eu). RCHdV acknowledges the Netherlands Metabolomics Centre, which is part of the Netherlands Genomics Initiative/Netherlands Organization for additional financial support. Support from Cost Action FA1006 is greatly appreciated. Funding for having this publication made available Open Access has been provided by the European Commission through its FP7 post-grant Gold Open Access Pilot.
Databáze: OpenAIRE
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