A Medicago truncatula ABC transporter belonging to subfamily G modulates the level of isoflavonoids.

Full-sized ATP-binding cassette (ABC) transporters of the G subfamily (ABCG) are considered to be essential components of the plant immune system. These proteins have been proposed to be implicated in the active transmembrane transport of various secondary metabolites. Despite the importance of ABCG-based transport for plant-microbe interactions, these proteins are still poorly recognized in legumes. The experiments described here demonstrated that the level of Medicago truncatula ABCG10 (MtABCG10) mRNA was elevated following application of fungal oligosaccharides to plant roots. Spatial expression pattern analysis with a reporter gene revealed that the MtABCG10 promoter was active in various organs, mostly within their vascular tissues. The corresponding protein was located in the plasma membrane. Silencing of MtABCG10 in hairy roots resulted in lower accumulation of the phenylpropanoid pathway-derived medicarpin and its precursors. PCR-based experiments indicated that infection with Fusarium oxysporum, a root-infecting pathogen, progressed faster in MtABCG10-silenced composite plants (consisting of wild-type shoots on transgenic roots) than in the corresponding controls. Based on the presented data, it is proposed that in Medicago, full-sized ABCG transporters might modulate isoflavonoid levels during the defence response associated with de novo synthesis of phytoalexins.

LC/MS profiling of flavonoid glycoconjugates isolated from hairy roots, suspension root cell cultures and seedling roots of Medicago truncatula.

Hairy roots and suspension cell cultures are commonly used in deciphering different problems related to the biochemistry and physiology of plant secondary metabolites. Here, we address about the issue of possible differences in the profiles of flavonoid compounds and their glycoconjugates derived from various plant materials grown in a standard culture media. We compared profiles of flavonoids isolated from seedling roots, hairy roots, and suspension root cell cultures of a model legume plant, Medicago truncatula. The analyses were conducted with plant isolates as well as the media. The LC/MS profiles of target natural products obtained from M. truncatula seedling roots, hairy roots, and suspension root cell cultures differed substantially. The most abundant compounds in seedlings roots were mono- and diglucuronides of isoflavones and/or flavones. This type of glycosylation was not observed in hairy roots or suspension root cell cultures. The only recognized glycoconjugates in the latter samples were glucose derivatives of isoflavones. Application of a high-resolution mass spectrometer helped evaluate the elemental composition of protonated molecules, such as [M + H](+). Comparison of collision-induced dissociation MS/MS spectra registered with a quadrupole time-of-flight analyzer for tissue extracts and standards allowed us to estimate the aglycone structure on the basis of the pseudo-MS(3) experiment. Structures of these natural products were described according to the registered mass spectra and literature data. The analyses conducted represent an overview of flavonoids and their conjugates in different types of plant material representing the model legume, M. truncatula. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11306-011-0287-2) contains supplementary material, which is available to authorized users.

LC-MSMS profiling of flavonoid conjugates in wild Mexican lupine, Lupinus reflexus.

Profiles of flavonoid conjugates present in the root and leaf tissues of the Mexican wild lupine, Lupinus reflexus, were established using two LC-MSMS systems in the positive and negative ion modes. The ion trap mass spectrometer and quadrupole time-of flight instrument provided sequential MS(n) spectra and MSMS spectra with accurate m/z values of [M + H](+) and [M - H] (-) ions, respectively. Sixty-two flavone and isoflavone glycoconjugates were found and tentatively identified. Numerous isomeric or isobaric compounds with the same molecular mass could be differentiated. Isomeric di- and mono glucosides of biochanin A, genistein, 2'-hydroxygenistein, luteone, and 2,3-didehydrokievitone were distinguished on the basis of relative abundances of product ions. The studied flavonoid glycoconjugates were acylated with dicarboxylic aliphatic acids and their methyl esters at either the aglycone or glycosidic moiety.