Circular dichroism spectrometry was used on oligogalacturonides (OGAs) and showed the existence of a calcium/sodium-induced conformational state that is intermediate between single-isolated chains and calcium-associated multimer chains. This conformation is interpreted as being egg box dimers. Using the 2F4 monoclonal antibody that specifically binds such an egg box dimer conformation of pectin, the stability of OGA dimers was investigated over a period of 24 hours. The extent to which egg box dimers were recognized by the antibody was dependent on the temperature and duration of preincubation of the OGA. This suggests a "maturation" process of the egg-box structure that consists in a progressive increase in the length of the junction sequences between two chains that slide along each other in order to form a maximum number of calcium bridges and dimer ends. The maturation of egg boxes induced both a significant increase in their binding to wall-associated kinase 1 (WAK1) and an increased extracellular alkalinization when applied to Arabidopsis thaliana cell suspensions. The chemical modification of the reducing end of the OGAs largely diminished their elicitating activity but did not hinder either dimerization or binding of these end-reduced egg boxes to WAK1. We conclude that there are at least two different perception systems for egg box dimers. One binds egg box junctions and the other binds egg box ends. The relevance of these results is discussed in terms of pectic signal perception and plant-pathogen interaction.

Plant cell walls undergo remodeling during growth and development and are the first target of many invading pathogens. Acidic pectin (homogalacturonans) binds calcium and forms chain dimers called egg boxes and even multimers at higher calcium ion concentrations. Chitosan, the deacetylated form of chitin produced by fungi when invading plant tissues, is a cationic polymer that can interact with negatively charged pectin. The interaction between chitosan oligomers (COS) and pectic egg boxes was investigated using 2F4, a monoclonal antibody specific for calcium-associated dimers of pectin. Depending on the size of the pectic molecules, the COS to pectin ratio, the degree of polymerization and the degree of acetylation of COS in the mixture, the calcium-induced egg box conformation of oligogalacturonides (OGA) was strongly stabilized or destroyed. The biological activity of COS-stabilized egg boxes was assayed on Arabidopsis cell suspensions. COS-OGA egg boxes strongly enhanced extracellular alkalinization and decreased potassium fluxes compared to control COS and OGA alone. Furthermore, OGA rescued Arabidopsis from cell death induced by higher concentrations of deacetylated COS. The stabilized COS-OGA egg boxes could constitute a combined emergency signal that informs plant cells on both cell wall degradation and pathogen presence, triggering a much stronger response than individual components alone.