Induction of RAGE shedding by activation of G protein-coupled receptors

PLoS One. 2012;7(7):e41823. doi: 10.1371/journal.pone.0041823. Epub 2012 Jul 30.


The multiligand Receptor for Advanced Glycation End products (RAGE) is involved in various pathophysiological processes, including diabetic inflammatory conditions and Alzheimers disease. Full-length RAGE, a cell surface-located type I membrane protein, can proteolytically be converted by metalloproteinases ADAM10 and MMP9 into a soluble RAGE form. Moreover, administration of recombinant soluble RAGE suppresses activation of cell surface-located RAGE by trapping RAGE ligands. Therefore stimulation of RAGE shedding might have a therapeutic value regarding inflammatory diseases. We aimed to investigate whether RAGE shedding is inducible via ligand-induced activation of G protein-coupled receptors (GPCRs). We chose three different GPCRs coupled to distinct signaling cascades: the V2 vasopressin receptor (V2R) activating adenylyl cyclase, the oxytocin receptor (OTR) linked to phospholipase Cβ, and the PACAP receptor (subtype PAC1) coupled to adenylyl cyclase, phospholipase Cβ, calcium signaling and MAP kinases. We generated HEK cell lines stably coexpressing an individual GPCR and full-length RAGE and then investigated GPCR ligand-induced activation of RAGE shedding. We found metalloproteinase-mediated RAGE shedding on the cell surface to be inducible via ligand-specific activation of all analyzed GPCRs. By using specific inhibitors we have identified Ca(2+) signaling, PKCα/PKCβI, CaMKII, PI3 kinases and MAP kinases to be involved in PAC1 receptor-induced RAGE shedding. We detected an induction of calcium signaling in all our cell lines coexpressing RAGE and different GPCRs after agonist treatment. However, we did not disclose a contribution of adenylyl cyclase in RAGE shedding induction. Furthermore, by using a selective metalloproteinase inhibitor and siRNA-mediated knock-down approaches, we show that ADAM10 and/or MMP9 are playing important roles in constitutive and PACAP-induced RAGE shedding. We also found that treatment of mice with PACAP increases the amount of soluble RAGE in the mouse lung. Our findings suggest that pharmacological stimulation of RAGE shedding might open alternative treatment strategies for Alzheimers disease and diabetes-induced inflammation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ADAM Proteins / antagonists & inhibitors
  • ADAM Proteins / genetics
  • ADAM Proteins / metabolism
  • ADAM10 Protein
  • ADAM17 Protein
  • Adenylyl Cyclases / metabolism
  • Amyloid Precursor Protein Secretases / antagonists & inhibitors
  • Amyloid Precursor Protein Secretases / genetics
  • Amyloid Precursor Protein Secretases / metabolism
  • Animals
  • Calcium Signaling
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Dipeptides / pharmacology
  • Gene Knockdown Techniques
  • HEK293 Cells
  • Humans
  • Hydroxamic Acids / pharmacology
  • Lung / metabolism
  • MAP Kinase Signaling System
  • Male
  • Matrix Metalloproteinase 2 / genetics
  • Matrix Metalloproteinase 2 / metabolism
  • Matrix Metalloproteinase 9 / genetics
  • Matrix Metalloproteinase 9 / metabolism
  • Membrane Proteins / antagonists & inhibitors
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism
  • Mice
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phosphatidylinositol 3-Kinases / physiology
  • Pituitary Adenylate Cyclase-Activating Polypeptide / pharmacology
  • Pituitary Adenylate Cyclase-Activating Polypeptide / physiology
  • Protease Inhibitors / pharmacology
  • Proteolysis
  • RNA Interference
  • Receptor for Advanced Glycation End Products
  • Receptors, Immunologic / metabolism*
  • Receptors, Oxytocin / metabolism*
  • Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I / metabolism*
  • Receptors, Vasopressin / metabolism*


  • 3-(formylhydroxyamino)-2-(3-phenyl-1-propyl)butanoic acid (2,2-dimethyl-1-methylcarbamoyl-1-propyl)amide
  • Dipeptides
  • Hydroxamic Acids
  • Membrane Proteins
  • N-(2(R)-2-(hydroxamidocarbonylmethyl)-4-methylpentanoyl)-L-tryptophan methylamide
  • OXTR protein, human
  • Pituitary Adenylate Cyclase-Activating Polypeptide
  • Protease Inhibitors
  • Receptor for Advanced Glycation End Products
  • Receptors, Immunologic
  • Receptors, Oxytocin
  • Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I
  • Receptors, Vasopressin
  • Phosphatidylinositol 3-Kinases
  • Cyclic AMP-Dependent Protein Kinases
  • Amyloid Precursor Protein Secretases
  • ADAM Proteins
  • MMP2 protein, human
  • Matrix Metalloproteinase 2
  • MMP9 protein, human
  • Matrix Metalloproteinase 9
  • ADAM10 Protein
  • ADAM10 protein, human
  • ADAM17 Protein
  • Adenylyl Cyclases

Grant support

This work was supported by the Alzheimer Forschung Initiative e.V. (Düsseldorf, Germany), grant 07807 (E.K.) and the Dr. Georg Scheuing-Stiftung (Mainz, Germany to R.P.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.