Sugar availability modulates polyisoprenoid and phytosterol profiles in Arabidopsis thaliana hairy root culture

Abstract

Sugars are recognized as signaling molecules regulating the biosynthesis of secondary metabolites in plants. Here, a modulatory effect of sugars on dolichol and phytosterol profiles was noted in the hairy roots of Arabidopsis thaliana. Arabidopsis roots contain a complex dolichol mixture comprising three groups ('families') of dolichols differing in the chainlength. These dolichols, especially the longest ones are accompanied by considerable amounts of polyprenols of the same length. The spectrum of polyisoprenoid alcohols, i.e. dolichols and polyprenols, was dependent on sugar type (glucose or sucrose) and its concentration in the medium. Among the long-chain dolichols Dol/Pren-20 (dolichol or prenol molecule composed of 20 isoprene residues) and Dol/Pren-23 were the main components at 0.5% and 2% glucose, respectively. Moreover, the ratio of polyprenols versus respective dolichols was also modulated by sugar in this group of polyisoprenoids, with polyprenols dominating at 3% sucrose and dolichols at 2% glucose. Glucose concentration affected the expression level of genes encoding cis-prenyltransferases, enzymes responsible for elongation of the polyisoprenoid chain.The most abundant phytosterols of the A. thaliana roots, β-sitosterol, stigmasterol and campesterol, were accompanied by corresponding stanols and traces of brassicasterol, stigmast-4,22-dien-3-one and stigmast-4-en-3-one. Similarly to the polyisoprenoids, sterol profile responded to the sugar present in the medium, β-sitosterol dominating in roots grown on 3% or lower glucose concentrations and stigmasterol in 3% sucrose. These results indicate on involvement of sugar signaling in the regulation of cis-prenyltransferases and phytosterol pathway enzymes.

Modeling of Dolichol Mass Spectra Isotopic Envelopes as a Tool to Monitor Isoprenoid Biosynthesis

“…S1 and S2), suggesting dedicated roles of individual CPTs in plant responses to different stresses and implying a role of reactive oxygen species (ROS) signaling in Dol generation. Analysis of the content of Prens, a-unsaturated counterparts accompanying Dols in Arabidopsis hairy roots (Jozwiak et al, 2013), performed in parallel, revealed a modest decrease of their total content with the concomitant increase of Dols upon sorbitol treatment (Fig. 1).…”

“…The complexity of these mass spectra reflects the dispersion of the probability of formation of differentially labeled [ 13 C]Dol isotopomers/isotopologs (Skorupinska-Tudek et al, 2008). One should also note that, upon MS analysis, Dols form monovalent ions (z = 1); thus, the recorded m/z corresponds to the mass of the ion (Skorupinska-Tudek et al, 2003;Jozwiak et al, 2013). Moreover, the signal of each isotopomer (of a given m/z) in fact corresponds to various isotopologs (Dol ions containing the same number of 13 C atoms differently localized along the polyisoprenoid chain).…”

“…Dols, Prens, and phytosterols were isolated as described earlier (Jozwiak et al, 2013) with modifications. Briefly, to isolate Dols, unsaponifiable lipids were purified on a silica gel column eluted with hexane containing increasing concentrations of diethyl ether (0%-25%).…”

“…In vitro culture of Arabidopsis (Arabidopsis thaliana) hairy roots was propagated as described previously (Jozwiak et al, 2013). For metabolic labeling with a general precursor, roots were grown in one-half-strength Murashige and Skoog medium supplemented with either 2% [1-13 C]Glc or 2% [U-13 C 6 ]Glc used as a sole carbon source.…”

“…1). Arabidopsis (Arabidopsis thaliana) hairy roots contain a complex Dol mixture comprising three groups (families) differing in chain length (Jozwiak et al, 2013). Their biosynthesis is catalyzed by cisprenyltransferases (CPTs), and six out of nine putative CPT-encoding genes of Arabidopsis are expressed in roots (Surmacz and Swiezewska, 2011).…”