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Biosynthesis, Structure, and Function of Plant Cell Walls

Higher plant cells are encased in cell walls that define their shape and contribute to the strength and structural integrity not only of individual cells, but also of the entire plant. Despite its necessary rigidity, the cell wall is a highly dynamic entity that is metabolically active and plays crucial roles in diverse cell activities such as growth, differentiation, cell-to-cell communication and transport, senescence, abscission, and plant-pathogen interactions. The wall can be described as a liquid crystal (Fig. 1): Microcrystalline cellulose is embedded in a hydrated matrix consisting of coextensive networks of complex heteropolysaccharides and sometimes glycoproteins. Cell walls also constitute renewable resources and are often present in by-products of industrial production, such as pulps. Genetic engineering of crop plant cell walls can identify biopolymers with novel functional properties, as well as simplify their extraction, thus increasing the value of these "waste-products." Cell walls will become more important in the future, as they are an abundant resource that can contribute to our biofuel needs.

Analytical Platform for the Microanalysis of Wall Polysaccharides

Currently mainly classical carbohydrate chemistry based methods are utilized in our lab to describe the structure of particular wall polysaccharides. These methods encompass solubilization of various wall polymers using sequential extraction procedures that make use of wall degrading enzymes and chemicals. The resulting fractions are then analysed using techniques such as monosaccharide composition and glycosidic linkage analysis. In addition, the presence of ester substituents (such as O-acetyl-substituents) is determined. However, such an analysis is rather labor-intensive and time consuming. Therefore, an oligosaccharide mass profiling method (OLIMP) using specific polysaccharide hydrolases in combination with mass spectrometry has been developed. The sensitivity of OLIMP allows for the rapid assessment of even minute amount of tissue-materials. A profile can be obtained from preparations of as little as 500 Arabidopsis cells prepared by a laser-dissection catapulting instrument.

One mean to identify wall mutants is the screening of populations of chemically mutagenised Arabidopsis seeds for novel structural wall mutants using OLIMP. This approach takes advantage of the speed of OLIMP and has lead to the identification of 60 distinct mutants with altered xyloglucan structures including the abundance of ester-substituents. Map-based cloning of the mutated genes should give valuable insights into biosynthesis, metabolism and function of structural variations of xyloglucan.

An additional approach is based on chemical genetics. Arabidopsis seeds can be germinated and grown in liquid culture with the addition of agents such as specific polysaccharide hydrolases. Due to the presence of the enzyme the seedlings exhibit distinct phenotypes (e.g., alterations in stomata development, growth and hypersensitive response phenotypes) that can be reversed when the agent, i.e. the enzyme, is removed. The enzyme containing medium was employed to screen mutagenized or T-DNA tagged Arabidopsis seed populations for the identification of mutants that exhibit a more severe phenotype ("hypersensitive mutants") or resembling the wild type phenotype grown without the enzyme ("resistant mutants"). Indeed, several mutants identified by their altered visible phenotype when subjected to the hydrolase exhibit an altered wall structure. Further characterization of the mutant phenotype as well as understanding the genetic basis for the phenotype should result not only in additional mutants with novel wall structures but also mutants impaired or altered in oligosaccharin signalling pathways.

Although information about the structural components of cell walls has considerably increased in recent years, very little is known about the biosynthesis of individual wall components on a molecular level. Thus, a reverse genetic approach is employed after the identification of candidate genes involved in this process. Currently, numerous novel genes involved in the synthesis of nucleotide sugars, the substrates for polysaccharide synthesis, have been identified through bioinformatic means by comparison to gene-sequences of well-characterized bacterial enzymes. The analysis of Arabidopsis insertional knock-out alleles of these candidate genes has already revealed, in some cases, a function for the gene (e.g., a UDP-rhamnose synthase), a plant with an altered polysaccharide composition (e.g., drastic reduction of rhamnogalacturonan I), and its effect on plant growth and morphology (e.g., detrimental seed development and morphology).

Training areas: Arabidopsis/tomato molecular biology, genetics, biochemistry, and carbohydrate chemistry.

Selected Publications

Jose SBS, Barton CJ, Taylor NG, Ryden P, Neumetzler L,Pauly M Roberts K, Seifert GJ (2006) Interactions between MUR10/CesA7 dependent secondary cellulose biosynthesis and primary cell wall structure, Plant Physiol 142: 1353-1363 Abstract

Carrai F, Baxter C, Urbanczyk-Wochniak E, Zanor MI, Nunes-Nesi A, Usadel B, Nikiforova V, Centero D, Ratzka A,Pauly M, Sweetlove L, Fernie AR (2006) Integrated analysis of metabolite and transcript levels reveals tight co-ordinate regulation of metabolism underlies tomato fruit development. Plant Physiol 142: 1380-1396 Abstract

Immerzeel P, Pauly M (2006) Profiling methods for the analysis of cell wall polysaccharides, New Zeal J Forest Sci 35: 55-67

Mouille G, Witucka-Wall H, Bryant MP, Loudet O, Rihouey C, Lerouxel O, Lerouge P, Hoefte H, Pauly M (2006) Quantitative Trait Loci analysis of primary cell wall composition in Arabidopsis thaliana. Plant Physiol 141: 1035-1044 Abstract

Diet A, Link B, Seifert GJ, Schellenberg B, Wagner U, Pauly M, Reiter WD, Ringli C (2006) The Arabidopsis root hair cell wall formation mutant lrx1 is suppressed by mutations in the RHM1 gene encoding a UDP-L-rhamnose synthase. Plant Cell 18: 1630-1641 Abstract

Harholt J, Jensen JK, Sorensen SO, Orfila C, Pauly M, Scheller HV (2006) Arabinan deficient 1 is a putative arabinosyltransferase involved in biosynthesis of pectin arabinan in Arabidopsis. Plant Physiol 140: 49-58 Abstract

Abdulrazzak N, Pollet B, Ehlting J, Larsen K, Asnaghi C, Ronseau S, Proux C, Erhardt M, Seltzer V, Renau JP, Ullmann P, Pauly M, Lapierre C, Werck-Reichhart D (2006) A coumaroyl-ester-3-hydroxylase insertion mutant reveals the existence of non redundant meta-hydroxylation pathways and essential roles for phenolic precursors in cell expansin and plant growth. Plant Physiol 140: 30-48 Abstract

Usadel B, Kuschinsky AM, Steinhauser D, Pauly M (2005) Transcriptional co-response analysis as a tool to identify new components of the wall biosynthetic machinery. Plant Biosystems 139: 69-73 Abstract

Scheible WR, Pauly M (2004) Glycosyltransferases and cell wall biosynthesis: novel players and insights. Curr Opin Plant Biol 7: 285-295 Abstract

Usadel B, Kuschinsky AM, Rosso M, Eckermann N, Pauly M (2004) RHM2 is involved in mucilage pectin synthesis and is required for the development of the seed coat in Arabidopsis thaliana. Plant Physiol 134: 286-285 Abstract

Lerouxel O, Choo TZ, Seveno M, Usadel B, Faye L, Lerouge P, Pauly M (2002) Rapid structural phenotyping of plant cell wall mutants by enzymatic oligosaccharide fingerprinting. Plant Physiol 130: 1754-1763 Abstract

Oxenbøll Sørensen SO, Pauly M, Bush MS, Skjøt M, McCann MC, Borkhardt B, Ulvskov P. (2000) Pectin engineering: Modification of potato pectin by in vivo expression of an Endo-1,4-galactanase. Proc Natl Acad Sci USA 97: 7639-7644 Abstract

Pauly M, Albersheim P, Darvill AG, York WS (1999) Molecular domains of the cellulose/xyloglucan network in the cell walls of higher plants. Plant J 20: 629-639 Abstract

Zablackis E, York WS, Pauly M, Hantus S, Reiter WD, Chapple CCS, Albersheim P, Darvill A (1996) Substitution of L-fucose by L-galactose in cell walls of Arabidopsis mur1. Science 272: 1808-1810 Abstract

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