Journal Article

Altered nitrogen metabolism associated with de‐differentiated suspension cultures derived from root cultures of <i>Datura stramonium</i> studied by heteronuclear multiple bond coherence (HMBC) NMR spectroscopy

Ophélie Fliniaux, François Mesnard, Sophie Raynaud‐Le Grandic, Sylvie Baltora‐Rosset, Christophe Bienaimé, Richard J. Robins and Marc‐André Fliniaux

in Journal of Experimental Botany

Published on behalf of Society for Experimental Biology

Volume 55, issue 399, pages 1053-1060
Published in print May 2004 | ISSN: 0022-0957
Published online May 2004 | e-ISSN: 1460-2431 | DOI: http://dx.doi.org/10.1093/jxb/erh119
Altered nitrogen metabolism associated with de‐differentiated suspension cultures derived from root cultures of Datura stramonium studied by heteronuclear multiple bond coherence (HMBC) NMR spectroscopy

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De‐differentiation of transformed root cultures of Datura stramonium has previously been shown to cause a loss of tropane alkaloid synthetic capacity. This indicates a marked shift in physiological status, notably in the flux of primary metabolites into tropane alkaloids. Nitrogen metabolism in transformed root cultures of D. stramonium (an alkaloid‐producing system) and de‐differentiated suspension cultures derived therefrom (a non‐producing system) has been compared using Nuclear Magnetic Resonance (NMR) spectroscopy. 15N‐Labelled precursors [(15NH4)2SO4 and K15NO3] were fed and their incorporation into nitrogenous metabolites studied using Heteronuclear Multiple Bond Coherence (HMBC) NMR spectroscopy. In both cultures, the same amino acids were resolved in the HMBC spectra. However, marked differences were found in the intensity of labelling of a range of nitrogenous compounds. In differentiated root cultures, cross‐peaks corresponding to secondary metabolites, such as tropine, were observed, whereas these were absent in the de‐differentiated cultures. By contrast, N‐ acetylputrescine and γ‐aminobutyric acid (GABA) accumulated in the de‐differentiated cultures to a much larger extent than in the root cultures. It can therefore be suggested that the loss of alkaloid biosynthesis was compensated by the diversion of putrescine metabolism away from the tropane pathway and toward the synthesis of GABA via N‐acetylputrescine.

Keywords: Datura stramonium, HMBC NMR, nitrogen metabolism, NMR, transformed cultures (root), transformed cultures (de‐differentiated), tropane alkaloids.

Journal Article.  4701 words.  Illustrated.

Subjects: Plant Sciences and Forestry

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