Structural characterisation of the pectic polysaccharide rhamnogalacturonan II using an acidic fingerprinting methodology
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Rhamnogalacturonan II (RG-II) is a structurally complex cell wall pectic polysaccharide. Despite its complexity, both the structure of RG-II and its ability to dimerise via a borate diester are conserved in vascular plants suggesting that RG-II has a fundamental role in primary cell wall organisation and function. The selection and analysis of new mutants affected in RG-II formation represents a promising strategy to unravel these functions and to identify genes encoding enzymes involved in RG-II biosynthesis. In this paper, a novel fingerprinting strategy is described for the screening of RG-II mutants based on the mild acid hydrolysis of RG-II coupled to the analysis of the resulting fragments by mass spectrometry. This methodology was developed using RG-II fractions isolated from citrus pectins and then validated for RG-II isolated from the Arabidopsis mur1 mutant and irx10 irx10-like double mutant.
KeywordsCell wall mutants Fingerprinting Mass spectrometry Rhamnogalacturonan II
1,2 Diamino-4,5-methylene dioxybenzene
Electrospray ionisation mass spectrometry
Matrix-assisted laser desorption ionisation-time of flight
Oligosaccharide mass profiling method
The Nottingham Arabidopsis Stock Center and Gregory Mouille, INRA Versailles, France are acknowledged for providing seeds for the Arabidopsis insertion lines and mutants. The authors thank Muriel Bardor, EA4358 University of Rouen, France for MALDI TOF analyses. This research was supported in France by the CNRS, the Région Haute-Normandie and the University of Rouen. Research at UPSC Sweden (A.M., A.-M.W.) was supported by the Swedish Foundation for Scientific Research (SSF) and Formas.
- Delmas F, Petit J, Joubès J, Seveno M, Paccalet T, Hernould M, Lerouge P, Mouras M, Chevalier C (2003) The gene expression and enzyme activity of plant 3-deoxy-D-manno-2-octulosonic acid-8-phosphate synthase are preferentially associated with cell division in a cell cycle-dependent manner. Plant Physiol 133:348–360PubMedCrossRefGoogle Scholar
- Egelund EJ, Petersen BL, Motawia MS, Damager I, Faik A, Olsen CE, Ishii T, Clausen H, Ulvskov P, Geshi N (2006) Arabidopsis thaliana RGXT1 and RGXT2 encode Golgi-localized (1, 3)-α-D-xylosyltransferases involved in the synthesis of pectic rhamnogalacturonan-II. Plant Cell 18:2593–2607PubMedCrossRefGoogle Scholar
- Fagard M, Desnos T, Desprez T, Goubet F, Refregier G, Mouille G, McCann M, Rayon C, Vernhettes S, Höfte H (2000) PROCUSTE1 encodes a cellulose synthase required for normal cell elongation specifically in roots and dark-grown hypocotyls of Arabidopsis. Plant Cell 12:2409–2423PubMedCrossRefGoogle Scholar
- Matsunaga T, Ishii T, Matsumoto S, Higuchi M, Darvill A, Albersheim P, O’Neill M (2004) Occurence of the primary cell wall polysaccharide rhamnogalacturonan II in Pteridophytes, Lycophytes, and Bryophytes. Implications for the evolution of vascular plants. Plant Physiol 134:339–351PubMedCrossRefGoogle Scholar
- Reuhs BL, Glenn J, Stephens SB, Kim JS, Christie DB, Glushka JG, Zablackis E, Albersheim P, Darvill A, O’Neill MA (2004) L-galactose replaces L-fucose in the pectic polysaccharide rhamnogalacturonan II synthesized by the L-fucose-deficient mur1 Arabidopsis mutant. Planta 219:147–157PubMedCrossRefGoogle Scholar