SummaryWhile N-glycan synthesis in the endoplasmic reticulum (ER) is relatively well con-

The presence of nonmammalian core alpha(1,3)-fucose and core xylose glyco-epitopes on glycans N-linked to therapeutic glycoproteins produced in plants has raised the question of their immunogenicity in human therapy. We address this question by studying the distribution of these N-glycans in pea, rice, and maize (which are the crops intended for the production of therapeutic proteins) and by reinvestigating their immunogenicity in rodents. We found that immunization with a model glycoprotein, horseradish peroxidase, elicits in C57BL/6 mice and rats the production of antibodies (Abs) specific for core alpha(1,3)-fucose and core xylose epitopes. Furthermore, we demonstrated that about 50% of nonallergic blood donors contains in their sera Abs specific for core xylose, whereas 25% have Abs against core alpha(1,3)-fucose. These Abs probably result from sensitization to environmental antigens. Although the immunological significance of these data is too speculative at the moment, the presence of such Abs might introduce some limitations to the use of plant-derived biopharmaceutical glycoproteins, such as an accelerated clearance during human therapy.

Asn-linked glycans, or the glycan code, carry crucial information for protein folding, transport, sorting, and degradation. The biochemical pathway for generating such a code is highly conserved in eukaryotic organisms and consists of ordered assembly of a lipid-linked tetradeccasaccharide. Most of our current knowledge on glycan biosynthesis was obtained from studies of yeast asparagine-linked glycosylation (alg) mutants. By contrast, little is known about biosynthesis and biological functions of N-glycans in plants. Here, we show that loss-of-function mutations in the Arabidopsis thaliana homolog of the yeast ALG12 result in transfer of incompletely assembled glycans to polypeptides. This metabolic defect significantly compromises the endoplasmic reticulum-associated degradation of bri1-9 and bri1-5, two defective transmembrane receptors for brassinosteroids. Consequently, overaccumulated bri1-9 or bri1-5 proteins saturate the quality control systems that retain the two mutated receptors in the endoplasmic reticulum and can thus leak out of the folding compartment, resulting in phenotypic suppression of the two bri1 mutants. Our results strongly suggest that the complete assembly of the lipid-linked glycans is essential for successful quality control of defective glycoproteins in Arabidopsis.

SummaryTransgenic plants are attractive biological systems for the large-scale production of pharmaceutical proteins. In particular, seeds offer special advantages, such as ease of handling and long-term stable storage. Nevertheless, most of the studies of the expression of antibodies in plants have been performed in leaves. We report the expression of a In addition, we demonstrate that a plant-made antibody with triantennary highmannose-type N -glycans has similar Fab functionality to its counterpart with biantennary complex N -glycans, but the former antibody interacts with protein A in a stronger manner and is more immunogenic than the latter. Such differences could be related to a variable immunoglobulin G (IgG)-Fc folding that would depend on the size of the N -glycan.