Abstract
To study the effect of short N-acetylglucosamine (GlcNAc) oligosaccharides on the physiology of plants, N-ACETYLGLUCOSAMINYLTRANSFERASE (NodC) of Azorhizobium caulinodans was expressed in Arabidopsis (Arabidopsis thaliana). The corresponding enzyme catalyzes the polymerization of GlcNAc and, accordingly, β-1,4-GlcNAc oligomers accumulated in the plant. A phenotype characterized by difficulties in developing an inflorescence stem was visible when plants were grown for several weeks under short-day conditions before transfer to long-day conditions. In addition, a positive correlation between the oligomer concentration and the penetrance of the phenotype was demonstrated. Although NodC overexpression lines produced less cell wall compared with wild-type plants under nonpermissive conditions, no indications were found for changes in the amount of the major cell wall polymers. The effect on the cell wall was reflected at the transcriptome level. In addition to genes encoding cell wall-modifying enzymes, a whole set of genes encoding membrane-coupled receptor-like kinases were differentially expressed upon GlcNAc accumulation, many of which encoded proteins with an extracellular Domain of Unknown Function26. Although stress-related genes were also differentially expressed, the observed response differed from that of a classical chitin response. This is in line with the fact that the produced chitin oligomers were too small to activate the chitin receptor-mediated signal cascade. Based on our observations, we propose a model in which the oligosaccharides modify the architecture of the cell wall by acting as competitors in carbohydrate-carbohydrate or carbohydrate-protein interactions, thereby affecting noncovalent interactions in the cell wall or at the interface between the cell wall and the plasma membrane.
Highlights
Bartel Vanholme*, Ruben Vanholme2, Halbay Turumtay2, Geert Goeminne, Igor Cesarino, Florence Goubet, Kris Morreel, Jorge Rencoret, Vincent Bulone, Cortwa Hooijmaijers, Riet De Rycke, Godelieve Gheysen, John Ralph, Marc De Block, Frank Meulewaeter, and Wout Boerjan
GlcNAc oligomer levels could be artificially increased in NodC OE lines with tunicamycin, an inhibitor of GlcNAc phosphotransferase, which catalyzes the first step in N-glycosylation, the transfer of GlcNAc phosphate from UDP-GlcNAc to dolichol phosphate (Koizumi et al, 1999)
The maximum degree of polymerization (DP) of the oligomers detected was 5 (Fig. 1), which is in agreement with the described function of this gene in the bacterial symbiont A. caulinodans (Geremia et al, 1994), where the GlcNAc pentamer forms the backbone for the lipooligosaccharide commonly known as the Nod factor
Summary
Bartel Vanholme*, Ruben Vanholme, Halbay Turumtay, Geert Goeminne, Igor Cesarino, Florence Goubet, Kris Morreel, Jorge Rencoret, Vincent Bulone, Cortwa Hooijmaijers, Riet De Rycke, Godelieve Gheysen, John Ralph, Marc De Block, Frank Meulewaeter, and Wout Boerjan. The in vitro production of these exopolymers has been described (Shirai et al, 1997; Lee et al, 2001; Liu et al, 2001; He et al, 2009; de Mesquita et al, 2012), no experimental data have been provided on the production of chitin or chitosan in plants or on their incorporation into plant cell walls Such in vivo production could have important benefits, as it would avoid the disadvantages related to the need in various applications for postisolation modification of cellulose fibers, such as low yield or quality, and the occurrence of undesirable side reactions, which are difficult to control. This suggests that shorter oligomers could be used to modify cellulose properties in the plant while avoiding the activation of stress responses
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