Spatial distribution and hormonal regulation of gene products from methyl erythritol phosphate and monoterpene-secoiridoid pathways in Catharanthus roseus

Abstract

The monoterpene indole alkaloids (MIAs) from Madagascar periwinkle (Catharanthus roseus) are secondary metabolites of high interest due to their therapeutical values. Secologanin, the monoterpenoid moiety incorporated into MIAs, is derived from the plastidial methyl-D: -erythritol 4-phosphate (MEP) pathway. Here, we have cloned a cDNA encoding hydroxymethylbutenyl diphosphate synthase (HDS), a MEP pathway enzyme, and generated antibodies to investigate the distribution of transcripts and protein in MIA-producing aerial tissues. Consistent with our earlier work, transcripts for the genes encoding the so-called early steps in monoterpenoid biosynthesis (ESMB) enzymes (HDS, others MEP pathway enzymes and geraniol 10-hydroxylase) were preferentially co-localized to internal phloem associated parenchyma (IPAP) cells. By contrast, transcripts for the enzyme catalysing the last biosynthetic step to secologanin, secologanin synthase, were found in the epidermis. A coordinated response of ESMB genes was also observed in cell cultures stimulated to synthesise MIAs by hormone treatment, whereas no changes in SLS expression were detected under the same experimental conditions. Immunocytolabelling studies with the HDS-specific serum demonstrated the localisation of HDS to the plastid stroma and revealed that HDS proteins were most abundant in IPAP cells but could also be found in other cell types, including epidermal and mesophyll cells. Besides showing the existence of post-transcriptional mechanisms regulating the levels of HDS in C. roseus cells, our results support that intercellular translocation likely plays an important role during monoterpene-secoiridoid assembly.

A Mutant Impaired in the Production of Plastome-Encoded Proteins Uncovers a Mechanism for the Homeostasis of Isoprenoid Biosynthetic Enzymes inArabidopsisPlastids

“…Several putative substrates, including a variety of chaperones and components required for the synthesis of plastome-encoded proteins, were recently identified by searching for stromal proteins differentially accumulated in chloroplasts from transgenic plants with reduced levels of Clp protease activity (Sjö gren et al, 2004(Sjö gren et al, , 2006Rudella et al, 2006). Our results (Figures 8 and 9) support the hypothesis that some of the MEP pathway enzymes, which are also localized in the stroma (Carretero-Paulet et al, 2002;Oudin et al, 2007), could be targets of the plastidic Clp protease machinery as well. However, current evidence does not allow us to conclude whether these proteins are direct targets (actual substrates) of this protease.…”

Growth regulators and essential oil production

“…This enzyme acts in the terpene moiety of indolic alkaloids as ajmalicine, serpentine and catharantine. Oudin et al (2007) verified that adding cytokinin to the medium increased the production of alkaloids with terpenic moiety. In in vitro culture of Lavandula dentata L. application of cytokinin (0.1 mg L -1 BA) had a positive effect on production and/or accumulation of essential oil.…”

“…GA ∅ x Activation of compounds conversion Zhang et al 2005 Catharanthus roseus CYT + x Increase of enzyme activity Decendit et al 1993;Papon et al 2005;Oudin et al 2007 Chrysanthemum cinerariaefolium ET ∅ x Enzyme activation Haque et al 2007 Cymbopogon citratus CYT + ∅ Source of hormone Craveiro et al 1989 C. citratus …”

The Plastidial 2-C-Methyl-d-Erythritol 4-Phosphate Pathway Provides the Isoprenyl Moiety for Protein Geranylgeranylation in Tobacco BY-2 Cells

“…However, exogenous sucrose reverses this effect and restores the MEP pathway, GGPP synthesis, and GFP-CaM53 geranylgeranylation. Indeed, evidence abounds for transport of MEP pathway intermediates between cells and tissues (Burlat et al, 2004;Gutié rrez-Nava et al, 2004;Oudin et al, 2007).…”