title: Molecular cloning and functional characterization of Catharanthus roseus hydroxymethylbutenyl 4-diphosphate synthase gene promoter from the methyl erythritol phosphate pathway.
abstract: The Madagascar periwinkle produces monoterpenoid indole alkaloids (MIA) of high interest due to their therapeutical values. The terpenoid moiety of MIA is derived from the methyl erythritol phosphate (MEP) and seco-iridoid pathways. These pathways are regarded as the limiting branch for MIA biosynthesis in C. roseus cell and tissue cultures. In previous studies, we demonstrated a coordinated regulation at the transcriptional and spatial levels of genes from both pathways. We report here on the isolation of the 5'-flanking region (1,049 bp) of the hydroxymethylbutenyl 4-diphosphate synthase (HDS) gene from the MEP pathway. To investigate promoter transcriptional activities, the HDS promoter was fused to GUS reporter gene. Agrobacterium-mediated transformation of young tobacco leaves revealed that the cloned HDS promoter displays a tissue-specific GUS staining restricted to the vascular region of the leaves and limited to a part of the vein that encompasses the phloem in agreement with the previous localization of HDS transcripts in C. roseus aerial organs. Further functional characterizations in stably or transiently transformed C. roseus cells allowed us to identify the region that can be consider as the minimal promoter and to demonstrate the induction of HDS promoter by several hormonal signals (auxin, cytokinin, methyljasmonate and ethylene) leading to MIA production. These results, and the bioinformatic analysis of the HDS 5'-region, suggest that the HDS promoter harbours a number of cis-elements binding specific transcription factors that would regulate the flux of terpenoid precursors involved in MIA biosynthesis.
authors.affiliation: Université François Rabelais de Tours, EA 2106, Biomolécules et Biotechnologies Végétales, 31 avenue Monge, 37200, Tours, France.
authors.forename: Olivia
authors.initials: O
authors.lastname: Ginis
authors.forename: Vincent
authors.initials: V
authors.lastname: Courdavault
authors.forename: Céline
authors.initials: C
authors.lastname: Melin
authors.forename: Arnaud
authors.initials: A
authors.lastname: Lanoue
authors.forename: Nathalie
authors.initials: N
authors.lastname: Giglioli-Guivarc'h
authors.forename: Benoit
authors.initials: B
authors.lastname: St-Pierre
authors.forename: Martine
authors.initials: M
authors.lastname: Courtois
authors.forename: Audrey
authors.initials: A
authors.lastname: Oudin
mesh_terms: D024506:5' Flanking Region / Q000235:genetics; D000085:Acetates / Q000494:pharmacology; D001483:Base Sequence; D053898:Biosynthetic Pathways / Q000187:drug effects; D030008:Catharanthus / Q000166:cytology / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics*; D003001:Cloning, Molecular; D003517:Cyclopentanes / Q000494:pharmacology; D003583:Cytokinins / Q000494:pharmacology; D004798:Enzymes / Q000235:genetics* / Q000378:metabolism; D004896:Erythritol / Q000031:analogs & derivatives* / Q000378:metabolism*; D005030:Ethylenes / Q000494:pharmacology; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D017343:Genes, Plant / Q000235:genetics*; D005966:Glucuronidase / Q000378:metabolism; D008969:Molecular Sequence Data; D059372:Nucleotide Motifs / Q000235:genetics; D009928:Organ Specificity / Q000187:drug effects / Q000235:genetics; D054883:Oxylipins / Q000494:pharmacology; D018515:Plant Leaves / Q000187:drug effects / Q000201:enzymology / Q000235:genetics; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D046948:Secologanin Tryptamine Alkaloids / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion / Q000235:genetics; D013403:Sugar Phosphates / Q000378:metabolism*; D013535:Suspensions; D014026:Nicotiana / Q000187:drug effects / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects
chemical_list: C114232:2-C-methylerythritol 4-phosphate; D000085:Acetates; D003517:Cyclopentanes; D003583:Cytokinins; D004798:Enzymes; D005030:Ethylenes; D054883:Oxylipins; D046948:Secologanin Tryptamine Alkaloids; D013403:Sugar Phosphates; D013535:Suspensions; C421242:hydroxymethylbutenyl 4-diphosphate synthase; C072239:methyl jasmonate; C036216:ethylene; D005966:Glucuronidase; D004896:Erythritol