Arabinogalactan proteins is an umbrella term applied to a highly diverse class of cell surface glycoproteins, many of which contain glycosylphosphatidylinositol lipid anchors. The structures of protein and glycan moieties of arabinogalactan proteins are overwhelmingly diverse while the "hydroxproline contiguity hypothesis" predicts arabinogalactan modification of members of many families of extracellular proteins. Descriptive studies using monoclonal antibodies reacting with carbohydrate epitopes on arabinogalactan proteins and experimental work using beta-Yariv reagent implicate arabinogalactan proteins in many biological processes of cell proliferation and survival, pattern formation and growth, and in plant microbe interaction. Advanced structural understanding of arabinogalactan proteins and an emerging molecular genetic definition of biological roles of individual arabinogalactan protein species, in conjunction with potentially analogous extracellular matrix components of animals, stimulate hypotheses about their mode of action. Arabinogalactan proteins might be soluble signals, or might act as modulators and coreceptors of apoplastic morphogens; their amphiphilic molecular nature makes them prime candidates of mediators between the cell wall, the plasma membrane, and the cytoplasm.

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In eukaryotes, class I a-mannosidases are involved in early N-glycan processing reactions and in N-glycan-dependent quality control in the endoplasmic reticulum (ER). To investigate the role of these enzymes in plants, we identified the ERtype a-mannosidase I (MNS3) and the two Golgi-a-mannosidase I proteins (MNS1 and MNS2) from Arabidopsis thaliana. All three MNS proteins were found to localize in punctate mobile structures reminiscent of Golgi bodies. Recombinant forms of the MNS proteins were able to process oligomannosidic N-glycans. While MNS3 efficiently cleaved off one selected a1,2-mannose residue from Man 9 GlcNAc 2 , MNS1/2 readily removed three a1,2-mannose residues from Man 8 GlcNAc 2 . Mutation in the MNS genes resulted in the formation of aberrant N-glycans in the mns3 single mutant and Man 8 GlcNAc 2 accumulation in the mns1 mns2 double mutant. N-glycan analysis in the mns triple mutant revealed the almost exclusive presence of Man 9 GlcNAc 2 , demonstrating that these three MNS proteins play a key role in N-glycan processing. The mns triple mutants displayed short, radially swollen roots and altered cell walls. Pharmacological inhibition of class I a-mannosidases in wildtype seedlings resulted in a similar root phenotype. These findings show that class I a-mannosidases are essential for early N-glycan processing and play a role in root development and cell wall biosynthesis in Arabidopsis.

Cell and cell wall growth are mutually dependent processes that must be tightly coordinated and controlled. LRR-extensin1 (LRX1) of Arabidopsis thaliana is a potential regulator of cell wall development, consisting of an N-terminal leucine-rich repeat domain and a C-terminal extensin-like domain typical for structural cell wall proteins. LRX1 is expressed in root hairs, and lrx1 mutant plants develop distorted root hairs that often swell, branch, or collapse. The aberrant cell wall structures found in lrx1 mutants point toward a function of LRX1 during the establishment of the extracellular matrix. To identify genes that are involved in an LRX1-dependent developmental pathway, a suppressor screen was performed on the lrx1 mutant, and two independent rol1 (for repressor of lrx1) alleles were isolated. ROL1 is allelic to Rhamnose Biosynthesis1, which codes for a protein involved in the biosynthesis of rhamnose, a major monosaccharide component of pectin. The rol1 mutations modify the pectic polysaccharide rhamnogalacturonan I and, for one allele, rhamnogalacturonan II. Furthermore, the rol1 mutations cause a change in the expression of a number of cell wall-related genes. Thus, the lrx1 mutant phenotype is likely to be suppressed by changes in pectic polysaccharides or other cell wall components.

Interactions between cell wall polymers are critical for establishing cell wall integrity and cell-cell adhesion. Here, we exploit the Arabidopsis (Arabidopsis thaliana) seed coat mucilage system to examine cell wall polymer interactions. On hydration, seeds release an adherent mucilage layer strongly attached to the seed in addition to a nonadherent layer that can be removed by gentle agitation. Rhamnogalacturonan I (RG I) is the primary component of adherent mucilage, with homogalacturonan, cellulose, and xyloglucan constituting minor components. Adherent mucilage contains rays composed of cellulose and pectin that extend above the center of each epidermal cell. CELLULOSE SYNTHASE5 (CESA5) and the arabinogalactan protein SALT-OVERLY SENSITIVE5 (SOS5) are required for mucilage adherence through unknown mechanisms. SOS5 has been suggested to mediate adherence by influencing cellulose biosynthesis. We, therefore, investigated the relationship between SOS5 and CESA5. cesa5-1 seeds show reduced cellulose, RG I, and ray size in adherent mucilage. In contrast, sos5-2 seeds have wild-type levels of cellulose but completely lack adherent RG I and rays. Thus, relative to each other, cesa5-1 has a greater effect on cellulose, whereas sos5-2 mainly affects pectin. The double mutant cesa5-1 sos5-2 has a much more severe loss of mucilage adherence, suggesting that SOS5 and CESA5 function independently. Doublemutant analyses with mutations in MUCILAGE MODIFIED2 and FLYING SAUCER1 that reduce mucilage release through pectin modification suggest that only SOS5 influences pectin-mediated adherence. Together, these findings suggest that SOS5 mediates adherence through pectins and does so independently of but in concert with cellulose synthesized by CESA5.

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Five Arabidopsis thaliana genes that encode UDP-glucose 4-epimerase (UGE) and represent two ancient plant UGE clades might be involved in the regulation of cell wall carbohydrate biosynthesis. We tested this hypothesis in a genome-wide reverse genetic study. Despite significant contributions of each gene to total UGE activity, none was essential for normal growth on soil. uge2 uge4 displayed dramatic general growth defects, while other mutant combinations were partially aberrant. UGE2 together with UGE3 influenced pollen development. UGE2 and UGE4 synergistically influenced cell wall galactose content, which was correlated with shoot growth. UGE2 strongly and UGE1 and UGE5 lightly supported UGE4 in influencing root growth and cell wall galactose content by affecting galactan content. By contrast, only UGE4 influenced xyloglucan galactosylation in roots. Secondary hypocotyl thickening and arabinogalactan protein carbohydrate structure in xylem parenchyma depended on the combination of UGE2 and UGE4. As opposed to cell wall galactose content, tolerance to external galactose strictly paralleled total UGE activity. We suggest a gradual recruitment of individual UGE isoforms into specific roles. UGE2 and UGE4 influence growth and cell wall carbohydrate biosynthesis throughout the plant, UGE3 is specialized for pollen development, and UGE1 and UGE5 might act in stress situations.

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