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Lysophosphatidic acid targets vascular and oncogenic pathways via RAGE signaling.

Rai V, Touré F, Chitayat S, Pei R, Song F, Li Q, Zhang J, Rosario R, Ramasamy R, Chazin WJ, Schmidt AM - J. Exp. Med. (2012)

Bottom Line: Hence, delineation of the full range of molecular mechanisms by which LPA exerts its broad effects is essential.In vivo, the administration of soluble RAGE or genetic deletion of RAGE mitigated LPA-stimulated vascular Akt signaling, autotaxin/LPA-driven phosphorylation of Akt and cyclin D1 in the mammary tissue of transgenic mice vulnerable to carcinogenesis, and ovarian tumor implantation and development.These findings identify novel roles for RAGE as a conduit for LPA signaling and suggest targeting LPA-RAGE interaction as a therapeutic strategy to modify the pathological actions of LPA.

View Article: PubMed Central - HTML - PubMed

Affiliation: Diabetes Research Program, Division of Endocrinology, Department of Medicine, New York University School of Medicine, New York, NY 10016, USA. vivek.raai@nyumc.org

ABSTRACT
The endogenous phospholipid lysophosphatidic acid (LPA) regulates fundamental cellular processes such as proliferation, survival, motility, and invasion implicated in homeostatic and pathological conditions. Hence, delineation of the full range of molecular mechanisms by which LPA exerts its broad effects is essential. We report avid binding of LPA to the receptor for advanced glycation end products (RAGE), a member of the immunoglobulin superfamily, and mapping of the LPA binding site on this receptor. In vitro, RAGE was required for LPA-mediated signal transduction in vascular smooth muscle cells and C6 glioma cells, as well as proliferation and migration. In vivo, the administration of soluble RAGE or genetic deletion of RAGE mitigated LPA-stimulated vascular Akt signaling, autotaxin/LPA-driven phosphorylation of Akt and cyclin D1 in the mammary tissue of transgenic mice vulnerable to carcinogenesis, and ovarian tumor implantation and development. These findings identify novel roles for RAGE as a conduit for LPA signaling and suggest targeting LPA-RAGE interaction as a therapeutic strategy to modify the pathological actions of LPA.

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Direct binding of LPA to RAGE: SPR and NMR. (A) Binding of 218 nM LPA to the immobilized sRAGE surface on CM5 sensor chip and SPR sensorgrams. (B) A monolayer of POPC liposomes was formed on the flow cell 1 (dark blue), and a monolayer of LPA-POPC liposomes was formed on the flow cell 2 (dark red). (C) Binding of 4 nM sRAGE to the immobilized LPA surface on HPA sensor chip. (D) sRAGE does not bind to POPC liposomes immobilized on flow cell 1 HPA sensor chip in sRAGE-LPA binding experiment (red curve). (E) SPR sensorgrams indicating that 1 µM sRAGE does not interact with immobilized S1P-POPC surface on flow cell 2 of the HPA sensor chip (dark red) and that sRAGE binds to LPA on flow cell 1 immobilized with LPA-POPC as the positive control for binding interaction (blue curve). (F) Binding of 9 nM RAGE V domain to the immobilized LPA surface. (G) Binding of 1 µM RAGE C2 domain to the immobilized LPA surface on HPA sensor chip. (H) Surface representation of the V domain (PDB 3CJJ) colored by electrostatic field, showing the highly basic (blue) character of the LPA binding surface. (I) NMR chemical shift perturbations mapped on the structure of the V domain (PDB 3CJJ) for LPA and Ca2+-loaded S100B. The significantly perturbed residues are highlighted in red. (J) 15N-1H HSQC NMR spectrum of 15N-enriched C2 domain in the absence (black) and presence (red) of LPA. (K) Complete 15N-1H HSQC NMR spectrum of 15N-enriched C2 domain in the absence (black) and presence (red) of LPA. (L) Binding of 1 µM sRAGE with 1 µM BSA or 1 µM S100B to the immobilized LPA liposomes on HPA sensor chip (blue and red, respectively). SPR assays results shown are representative of three independent experiments.
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fig1: Direct binding of LPA to RAGE: SPR and NMR. (A) Binding of 218 nM LPA to the immobilized sRAGE surface on CM5 sensor chip and SPR sensorgrams. (B) A monolayer of POPC liposomes was formed on the flow cell 1 (dark blue), and a monolayer of LPA-POPC liposomes was formed on the flow cell 2 (dark red). (C) Binding of 4 nM sRAGE to the immobilized LPA surface on HPA sensor chip. (D) sRAGE does not bind to POPC liposomes immobilized on flow cell 1 HPA sensor chip in sRAGE-LPA binding experiment (red curve). (E) SPR sensorgrams indicating that 1 µM sRAGE does not interact with immobilized S1P-POPC surface on flow cell 2 of the HPA sensor chip (dark red) and that sRAGE binds to LPA on flow cell 1 immobilized with LPA-POPC as the positive control for binding interaction (blue curve). (F) Binding of 9 nM RAGE V domain to the immobilized LPA surface. (G) Binding of 1 µM RAGE C2 domain to the immobilized LPA surface on HPA sensor chip. (H) Surface representation of the V domain (PDB 3CJJ) colored by electrostatic field, showing the highly basic (blue) character of the LPA binding surface. (I) NMR chemical shift perturbations mapped on the structure of the V domain (PDB 3CJJ) for LPA and Ca2+-loaded S100B. The significantly perturbed residues are highlighted in red. (J) 15N-1H HSQC NMR spectrum of 15N-enriched C2 domain in the absence (black) and presence (red) of LPA. (K) Complete 15N-1H HSQC NMR spectrum of 15N-enriched C2 domain in the absence (black) and presence (red) of LPA. (L) Binding of 1 µM sRAGE with 1 µM BSA or 1 µM S100B to the immobilized LPA liposomes on HPA sensor chip (blue and red, respectively). SPR assays results shown are representative of three independent experiments.

Mentions: To test the hypothesis that LPA could induce signaling through RAGE, we first performed experiments to test for LPA–RAGE physical interaction. The extracellular portion of RAGE (soluble RAGE [sRAGE]) was immobilized on a carboxymethylated dextran CM5 chip and high affinity LPA binding was observed by surface plasmon resonance (SPR; Fig. 1 A). To confirm the interaction, we reversed the binding assay and examined the binding of sRAGE to immobilized LPA (18:1) 1-Palmitoyl-2-Oleoyl-sn-Glycero-3-Phosphocholine (POPC; POPC-LPA 10:1 wt/wt) and POPC liposomes as a reference on the HPA sensor chip (Fig. 1 B). The SPR binding response in these experiments validated that sRAGE bound LPA (Fig. 1 C).


Lysophosphatidic acid targets vascular and oncogenic pathways via RAGE signaling.

Rai V, Touré F, Chitayat S, Pei R, Song F, Li Q, Zhang J, Rosario R, Ramasamy R, Chazin WJ, Schmidt AM - J. Exp. Med. (2012)

Direct binding of LPA to RAGE: SPR and NMR. (A) Binding of 218 nM LPA to the immobilized sRAGE surface on CM5 sensor chip and SPR sensorgrams. (B) A monolayer of POPC liposomes was formed on the flow cell 1 (dark blue), and a monolayer of LPA-POPC liposomes was formed on the flow cell 2 (dark red). (C) Binding of 4 nM sRAGE to the immobilized LPA surface on HPA sensor chip. (D) sRAGE does not bind to POPC liposomes immobilized on flow cell 1 HPA sensor chip in sRAGE-LPA binding experiment (red curve). (E) SPR sensorgrams indicating that 1 µM sRAGE does not interact with immobilized S1P-POPC surface on flow cell 2 of the HPA sensor chip (dark red) and that sRAGE binds to LPA on flow cell 1 immobilized with LPA-POPC as the positive control for binding interaction (blue curve). (F) Binding of 9 nM RAGE V domain to the immobilized LPA surface. (G) Binding of 1 µM RAGE C2 domain to the immobilized LPA surface on HPA sensor chip. (H) Surface representation of the V domain (PDB 3CJJ) colored by electrostatic field, showing the highly basic (blue) character of the LPA binding surface. (I) NMR chemical shift perturbations mapped on the structure of the V domain (PDB 3CJJ) for LPA and Ca2+-loaded S100B. The significantly perturbed residues are highlighted in red. (J) 15N-1H HSQC NMR spectrum of 15N-enriched C2 domain in the absence (black) and presence (red) of LPA. (K) Complete 15N-1H HSQC NMR spectrum of 15N-enriched C2 domain in the absence (black) and presence (red) of LPA. (L) Binding of 1 µM sRAGE with 1 µM BSA or 1 µM S100B to the immobilized LPA liposomes on HPA sensor chip (blue and red, respectively). SPR assays results shown are representative of three independent experiments.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3526353&req=5

fig1: Direct binding of LPA to RAGE: SPR and NMR. (A) Binding of 218 nM LPA to the immobilized sRAGE surface on CM5 sensor chip and SPR sensorgrams. (B) A monolayer of POPC liposomes was formed on the flow cell 1 (dark blue), and a monolayer of LPA-POPC liposomes was formed on the flow cell 2 (dark red). (C) Binding of 4 nM sRAGE to the immobilized LPA surface on HPA sensor chip. (D) sRAGE does not bind to POPC liposomes immobilized on flow cell 1 HPA sensor chip in sRAGE-LPA binding experiment (red curve). (E) SPR sensorgrams indicating that 1 µM sRAGE does not interact with immobilized S1P-POPC surface on flow cell 2 of the HPA sensor chip (dark red) and that sRAGE binds to LPA on flow cell 1 immobilized with LPA-POPC as the positive control for binding interaction (blue curve). (F) Binding of 9 nM RAGE V domain to the immobilized LPA surface. (G) Binding of 1 µM RAGE C2 domain to the immobilized LPA surface on HPA sensor chip. (H) Surface representation of the V domain (PDB 3CJJ) colored by electrostatic field, showing the highly basic (blue) character of the LPA binding surface. (I) NMR chemical shift perturbations mapped on the structure of the V domain (PDB 3CJJ) for LPA and Ca2+-loaded S100B. The significantly perturbed residues are highlighted in red. (J) 15N-1H HSQC NMR spectrum of 15N-enriched C2 domain in the absence (black) and presence (red) of LPA. (K) Complete 15N-1H HSQC NMR spectrum of 15N-enriched C2 domain in the absence (black) and presence (red) of LPA. (L) Binding of 1 µM sRAGE with 1 µM BSA or 1 µM S100B to the immobilized LPA liposomes on HPA sensor chip (blue and red, respectively). SPR assays results shown are representative of three independent experiments.
Mentions: To test the hypothesis that LPA could induce signaling through RAGE, we first performed experiments to test for LPA–RAGE physical interaction. The extracellular portion of RAGE (soluble RAGE [sRAGE]) was immobilized on a carboxymethylated dextran CM5 chip and high affinity LPA binding was observed by surface plasmon resonance (SPR; Fig. 1 A). To confirm the interaction, we reversed the binding assay and examined the binding of sRAGE to immobilized LPA (18:1) 1-Palmitoyl-2-Oleoyl-sn-Glycero-3-Phosphocholine (POPC; POPC-LPA 10:1 wt/wt) and POPC liposomes as a reference on the HPA sensor chip (Fig. 1 B). The SPR binding response in these experiments validated that sRAGE bound LPA (Fig. 1 C).

Bottom Line: Hence, delineation of the full range of molecular mechanisms by which LPA exerts its broad effects is essential.In vivo, the administration of soluble RAGE or genetic deletion of RAGE mitigated LPA-stimulated vascular Akt signaling, autotaxin/LPA-driven phosphorylation of Akt and cyclin D1 in the mammary tissue of transgenic mice vulnerable to carcinogenesis, and ovarian tumor implantation and development.These findings identify novel roles for RAGE as a conduit for LPA signaling and suggest targeting LPA-RAGE interaction as a therapeutic strategy to modify the pathological actions of LPA.

View Article: PubMed Central - HTML - PubMed

Affiliation: Diabetes Research Program, Division of Endocrinology, Department of Medicine, New York University School of Medicine, New York, NY 10016, USA. vivek.raai@nyumc.org

ABSTRACT
The endogenous phospholipid lysophosphatidic acid (LPA) regulates fundamental cellular processes such as proliferation, survival, motility, and invasion implicated in homeostatic and pathological conditions. Hence, delineation of the full range of molecular mechanisms by which LPA exerts its broad effects is essential. We report avid binding of LPA to the receptor for advanced glycation end products (RAGE), a member of the immunoglobulin superfamily, and mapping of the LPA binding site on this receptor. In vitro, RAGE was required for LPA-mediated signal transduction in vascular smooth muscle cells and C6 glioma cells, as well as proliferation and migration. In vivo, the administration of soluble RAGE or genetic deletion of RAGE mitigated LPA-stimulated vascular Akt signaling, autotaxin/LPA-driven phosphorylation of Akt and cyclin D1 in the mammary tissue of transgenic mice vulnerable to carcinogenesis, and ovarian tumor implantation and development. These findings identify novel roles for RAGE as a conduit for LPA signaling and suggest targeting LPA-RAGE interaction as a therapeutic strategy to modify the pathological actions of LPA.

Show MeSH
Related in: MedlinePlus