Centre for Vision and Vascular Science, Queen's University Belfast, Royal Victoria Hospital, Belfast BT12 6BA, Northern Ireland, United Kingdom.
J Biol Chem. 2010 Jul 23;285(30):23137-46. doi: 10.1074/jbc.M110.133827. Epub 2010 May 26.
The receptor for advanced glycation end products (RAGE) is a pattern-recognition receptor that binds to diverse ligands and initiates a downstream proinflammatory signaling cascade. RAGE activation has been linked to diabetic complications, Alzheimer disease, infections, and cancers. RAGE is known to mediate cell signaling and downstream proinflammatory gene transcription activation, although the precise mechanism surrounding receptor-ligand interactions is still being elucidated. Recent fluorescence resonance energy transfer evidence indicates that RAGE may form oligomers on the cell surface and that this could be related to signal transduction. To investigate whether RAGE forms oligomers, protein-protein interaction assays were carried out. Here, we demonstrate the interaction between RAGE molecules via their N-terminal V domain, which is an important region involved in ligand recognition. By protein cross-linking using water-soluble and membrane-impermeable cross-linker bis(sulfosuccinimidyl) suberate and nondenaturing gels, we show that RAGE forms homodimers at the plasma membrane, a process potentiated by S100B and advanced glycation end products. Soluble RAGE, the RAGE inhibitor, is also capable of binding to RAGE, similar to V peptide, as shown by surface plasmon resonance. Incubation of cells with soluble RAGE or RAGE V domain peptide inhibits RAGE dimerization, subsequent phosphorylation of intracellular MAPK proteins, and activation of NF-kappaB pathways. Thus, the data indicate that dimerization of RAGE represents an important component of RAGE-mediated cell signaling.
晚期糖基化终产物受体（RAGE）是一种模式识别受体，可与多种配体结合，并启动下游促炎信号级联反应。RAGE 的激活与糖尿病并发症、阿尔茨海默病、感染和癌症有关。已知 RAGE 介导细胞信号转导和下游促炎基因转录激活，尽管受体-配体相互作用的确切机制仍在阐明之中。最近的荧光共振能量转移证据表明，RAGE 可能在细胞表面形成寡聚体，这可能与信号转导有关。为了研究 RAGE 是否形成寡聚体，进行了蛋白-蛋白相互作用分析。在这里，我们通过其 N 端 V 结构域证明了 RAGE 分子之间的相互作用，V 结构域是参与配体识别的重要区域。通过使用水溶性和非膜渗透性交联剂双（磺基琥珀酰亚胺基）酯和非变性凝胶进行蛋白交联，我们表明 RAGE 在质膜上形成同源二聚体，S100B 和晚期糖基化终产物增强了这一过程。可溶性 RAGE（RAGE 抑制剂）也能够与 RAGE 结合，类似于 V 肽，这一点如表面等离子体共振所显示的那样。用可溶性 RAGE 或 RAGE V 结构域肽孵育细胞可抑制 RAGE 二聚化、细胞内 MAPK 蛋白的磷酸化以及 NF-κB 途径的激活。因此，数据表明 RAGE 的二聚化是 RAGE 介导的细胞信号转导的一个重要组成部分。
