Sanford Children's Health Research Center, Sanford-Burnham Medical Research Institute, La Jolla, California 92037, USA.
J Cell Biochem. 2010 Jun 1;110(3):645-59. doi: 10.1002/jcb.22575.
The receptor for advanced glycation end products (RAGE) is a signaling receptor protein of the immunoglobulin superfamily implicated in multiple pathologies. It binds a diverse repertoire of ligands, but the structural basis for the interaction of different ligands is not well understood. We earlier showed that carboxylated glycans on the V-domain of RAGE promote the binding of HMGB1 and S100A8/A9. Here we study the role of these glycans on the binding and intracellular signaling mediated by another RAGE ligand, S100A12. S100A12 binds carboxylated glycans, and a subpopulation of RAGE enriched for carboxylated glycans shows more than 10-fold higher binding potential for S100A12 than total RAGE. When expressed in mammalian cells, RAGE is modified by complex glycans predominantly at the first glycosylation site (N25IT) that retains S100A12 binding. Glycosylation of RAGE and maximum binding sites for S100A12 on RAGE are also cell type dependent. Carboxylated glycan-enriched population of RAGE forms higher order multimeric complexes with S100A12, and this ability to multimerize is reduced upon deglycosylation or by using non-glycosylated sRAGE expressed in E. coli. mAbGB3.1, an antibody against carboxylated glycans, blocks S100A12-mediated NF-kappaB signaling in HeLa cells expressing full-length RAGE. These results demonstrate that carboxylated N-glycans on RAGE enhance binding potential and promote receptor clustering and subsequent signaling events following oligomeric S100A12 binding.
晚期糖基化终产物受体（RAGE）是免疫球蛋白超家族的信号转导受体蛋白，与多种病理过程有关。它结合了一系列不同的配体，但不同配体相互作用的结构基础尚不清楚。我们之前表明，RAGE V 结构域上的羧基化聚糖促进了 HMGB1 和 S100A8/A9 的结合。在这里，我们研究了这些聚糖在另一种 RAGE 配体 S100A12 的结合和细胞内信号转导中的作用。S100A12 结合羧基化聚糖，富含羧基化聚糖的 RAGE 亚群对 S100A12 的结合潜力比总 RAGE 高 10 多倍。当在哺乳动物细胞中表达时，RAGE 被复杂聚糖修饰，主要在第一个糖基化位点（N25IT），该位点保留了 S100A12 的结合。RAGE 的糖基化和 S100A12 在 RAGE 上的最大结合位点也依赖于细胞类型。富含羧基化聚糖的 RAGE 亚群与 S100A12 形成更高阶的多聚体复合物，而这种多聚化能力在去糖基化或使用在大肠杆菌中表达的非糖基化 sRAGE 时会降低。针对羧基化聚糖的 mAbGB3.1 抗体可阻断全长 RAGE 表达的 HeLa 细胞中 S100A12 介导的 NF-kappaB 信号转导。这些结果表明，RAGE 上的羧基化 N-聚糖增强了结合潜力，并促进了寡聚 S100A12 结合后受体的聚类和随后的信号事件。
