In-situ analysis of pectic polysaccharides in seed mucilage and at the root surface of Arabidopsis thaliana

Pectic polysaccharides are a complex set of macromolecules of the primary cell wall matrix with distinct structural domains. The biosynthesis, organisation and function of these domains within cell wall matrices are poorly understood. An immersion immunofluorescence labelling technique was developed for the in-situ analysis of pectic polysaccharides at the surface of seeds and seedlings of Arabidopsis thaliana (L.) Heynh., and used to investigate the occurrence of pectic homogalacturonan (HG) and rhamnogalacturonan-I (RG-I) epitopes. Seed mucilage appeared to consist of two regions: a highly methyl-esterified HG was a major component throughout the mucilage, while an inner region with relatively low porosity was stabilized by calcium-based HG cross-linking. The small size and transparency of Arabidopsis roots allowed the occurrence of pectic HG and RG-I epitopes at root surfaces to be directly determined on whole-mount preparations. Pectic epitopes were not distributed evenly over root surfaces and were notably absent from lateral root apices and from the surface of root hairs. The use of defined antibody probes in the immersion immunolabelling protocol will be useful for the analysis of the influence of growth conditions and genetic factors on pectic polysaccharides in Arabidopsis.