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EMMPRIN/CD147 is a novel coreceptor of VEGFR-2 mediating its activation by VEGF.

Khayati F, Pérez-Cano L, Maouche K, Sadoux A, Boutalbi Z, Podgorniak MP, Maskos U, Setterblad N, Janin A, Calvo F, Lebbé C, Menashi S, Fernandez-Recio J, Mourah S - Oncotarget (2015)

Bottom Line: EMMPRIN/CD147 is mainly known for its protease inducing function but a role in promoting tumor angiogenesis has also been demonstrated.This study provides evidence that EMMPRIN is a new coreceptor for the VEGFR-2 tyrosine kinase receptor in both endothelial and tumor cells, as it directly interacts with it and regulates its activation by its VEGF ligand, signalling and functional consequences both in vitro and in vivo.EMMPRIN is overexpressed in cancer and hence is able to further potentiate VEGFR-2 activation, suggesting that a combinatory therapy of an antiangiogenic drug together with an inhibitor of EMMPRIN/VEGFR-2 interaction may have a greater impact on inhibiting angiogenesis and malignancy.

View Article: PubMed Central - PubMed

Affiliation: INSERM UMR-S 976, Hôpital Saint-Louis, Paris, France.

ABSTRACT
EMMPRIN/CD147 is mainly known for its protease inducing function but a role in promoting tumor angiogenesis has also been demonstrated. This study provides evidence that EMMPRIN is a new coreceptor for the VEGFR-2 tyrosine kinase receptor in both endothelial and tumor cells, as it directly interacts with it and regulates its activation by its VEGF ligand, signalling and functional consequences both in vitro and in vivo. Computational docking analyses and mutagenesis studies identified a molecular binding site in the extracellular domain of EMMPRIN located close to the cell membrane and containing the amino acids 195/199. EMMPRIN is overexpressed in cancer and hence is able to further potentiate VEGFR-2 activation, suggesting that a combinatory therapy of an antiangiogenic drug together with an inhibitor of EMMPRIN/VEGFR-2 interaction may have a greater impact on inhibiting angiogenesis and malignancy.

No MeSH data available.


Related in: MedlinePlus

EMMPRIN/CD147 interacts with VEGFR-2 and VEGF in endothelial and tumor cells(A) VEGFR-2 and VEGF from HMEC and M10 cell lysates were immunoprecipitated (IP) with anti-VEGFR-2 and anti-VEGF antibody respectively; western blotting was performed using anti-EMMPRIN antibody. Non immune IgG was used as controls. Representative blots of three independent experiments are shown. (B)In situ Proximity ligation assay (PLA) detection of EMMPRIN-VEGFR-2 and EMMPRIN-VEGF heterodimers (red dots). Negative controls without primary antibody are also shown. Nuclei were stained with DAPI (blue), magnification x 63. Representative images of three independent experiments are shown. (C) Direct interaction between the recombinant EMMPRIN and the recombinant VEGFR-2 in vitro. VEGFR-2 was first incubated with protein G beads prior to the addition of the recombinant EMMPRIN. Bound proteins were subsequently analyzed by Western blotting. Non-immune IgG served as a negative control and interaction between VEGF and VEGFR-2 served as a positive control. (D) Cells (HMEC, MDA-MB-231 and M10) were transfected for 24 hours with EMMPRIN siRNA or scrambled control siRNA at 33nmol/L concentration, and then subjected to IP assays using antibodies against VEGFR-2 and VEGF. Western blotting was performed using anti-EMMPRIN antibody. Representative blots of three independent experiments are shown.
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Figure 1: EMMPRIN/CD147 interacts with VEGFR-2 and VEGF in endothelial and tumor cells(A) VEGFR-2 and VEGF from HMEC and M10 cell lysates were immunoprecipitated (IP) with anti-VEGFR-2 and anti-VEGF antibody respectively; western blotting was performed using anti-EMMPRIN antibody. Non immune IgG was used as controls. Representative blots of three independent experiments are shown. (B)In situ Proximity ligation assay (PLA) detection of EMMPRIN-VEGFR-2 and EMMPRIN-VEGF heterodimers (red dots). Negative controls without primary antibody are also shown. Nuclei were stained with DAPI (blue), magnification x 63. Representative images of three independent experiments are shown. (C) Direct interaction between the recombinant EMMPRIN and the recombinant VEGFR-2 in vitro. VEGFR-2 was first incubated with protein G beads prior to the addition of the recombinant EMMPRIN. Bound proteins were subsequently analyzed by Western blotting. Non-immune IgG served as a negative control and interaction between VEGF and VEGFR-2 served as a positive control. (D) Cells (HMEC, MDA-MB-231 and M10) were transfected for 24 hours with EMMPRIN siRNA or scrambled control siRNA at 33nmol/L concentration, and then subjected to IP assays using antibodies against VEGFR-2 and VEGF. Western blotting was performed using anti-EMMPRIN antibody. Representative blots of three independent experiments are shown.

Mentions: The potential interaction between EMMPRIN and VEGFR-2 was investigated by immunoprecipitation (IP) assays in endothelial cells HMEC and melanoma cells M10. Complex formation was identified by the immunoprecipitation of either VEGFR-2 or VEGF followed by EMMPRIN immunoblotting (Figure 1A). IgG was used as a negative control. The fluorescent red spots observed using in situ proximity ligation assay (PLA) (Figure 1B) and confocal microscopy, a method which highlights protein/protein closely colocalized in cells, confirmed the proximity between EMMPRIN and VEGFR-2, and to a lesser extent between EMMPRIN and VEGF, at the cell surface.


EMMPRIN/CD147 is a novel coreceptor of VEGFR-2 mediating its activation by VEGF.

Khayati F, Pérez-Cano L, Maouche K, Sadoux A, Boutalbi Z, Podgorniak MP, Maskos U, Setterblad N, Janin A, Calvo F, Lebbé C, Menashi S, Fernandez-Recio J, Mourah S - Oncotarget (2015)

EMMPRIN/CD147 interacts with VEGFR-2 and VEGF in endothelial and tumor cells(A) VEGFR-2 and VEGF from HMEC and M10 cell lysates were immunoprecipitated (IP) with anti-VEGFR-2 and anti-VEGF antibody respectively; western blotting was performed using anti-EMMPRIN antibody. Non immune IgG was used as controls. Representative blots of three independent experiments are shown. (B)In situ Proximity ligation assay (PLA) detection of EMMPRIN-VEGFR-2 and EMMPRIN-VEGF heterodimers (red dots). Negative controls without primary antibody are also shown. Nuclei were stained with DAPI (blue), magnification x 63. Representative images of three independent experiments are shown. (C) Direct interaction between the recombinant EMMPRIN and the recombinant VEGFR-2 in vitro. VEGFR-2 was first incubated with protein G beads prior to the addition of the recombinant EMMPRIN. Bound proteins were subsequently analyzed by Western blotting. Non-immune IgG served as a negative control and interaction between VEGF and VEGFR-2 served as a positive control. (D) Cells (HMEC, MDA-MB-231 and M10) were transfected for 24 hours with EMMPRIN siRNA or scrambled control siRNA at 33nmol/L concentration, and then subjected to IP assays using antibodies against VEGFR-2 and VEGF. Western blotting was performed using anti-EMMPRIN antibody. Representative blots of three independent experiments are shown.
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Figure 1: EMMPRIN/CD147 interacts with VEGFR-2 and VEGF in endothelial and tumor cells(A) VEGFR-2 and VEGF from HMEC and M10 cell lysates were immunoprecipitated (IP) with anti-VEGFR-2 and anti-VEGF antibody respectively; western blotting was performed using anti-EMMPRIN antibody. Non immune IgG was used as controls. Representative blots of three independent experiments are shown. (B)In situ Proximity ligation assay (PLA) detection of EMMPRIN-VEGFR-2 and EMMPRIN-VEGF heterodimers (red dots). Negative controls without primary antibody are also shown. Nuclei were stained with DAPI (blue), magnification x 63. Representative images of three independent experiments are shown. (C) Direct interaction between the recombinant EMMPRIN and the recombinant VEGFR-2 in vitro. VEGFR-2 was first incubated with protein G beads prior to the addition of the recombinant EMMPRIN. Bound proteins were subsequently analyzed by Western blotting. Non-immune IgG served as a negative control and interaction between VEGF and VEGFR-2 served as a positive control. (D) Cells (HMEC, MDA-MB-231 and M10) were transfected for 24 hours with EMMPRIN siRNA or scrambled control siRNA at 33nmol/L concentration, and then subjected to IP assays using antibodies against VEGFR-2 and VEGF. Western blotting was performed using anti-EMMPRIN antibody. Representative blots of three independent experiments are shown.
Mentions: The potential interaction between EMMPRIN and VEGFR-2 was investigated by immunoprecipitation (IP) assays in endothelial cells HMEC and melanoma cells M10. Complex formation was identified by the immunoprecipitation of either VEGFR-2 or VEGF followed by EMMPRIN immunoblotting (Figure 1A). IgG was used as a negative control. The fluorescent red spots observed using in situ proximity ligation assay (PLA) (Figure 1B) and confocal microscopy, a method which highlights protein/protein closely colocalized in cells, confirmed the proximity between EMMPRIN and VEGFR-2, and to a lesser extent between EMMPRIN and VEGF, at the cell surface.

Bottom Line: EMMPRIN/CD147 is mainly known for its protease inducing function but a role in promoting tumor angiogenesis has also been demonstrated.This study provides evidence that EMMPRIN is a new coreceptor for the VEGFR-2 tyrosine kinase receptor in both endothelial and tumor cells, as it directly interacts with it and regulates its activation by its VEGF ligand, signalling and functional consequences both in vitro and in vivo.EMMPRIN is overexpressed in cancer and hence is able to further potentiate VEGFR-2 activation, suggesting that a combinatory therapy of an antiangiogenic drug together with an inhibitor of EMMPRIN/VEGFR-2 interaction may have a greater impact on inhibiting angiogenesis and malignancy.

View Article: PubMed Central - PubMed

Affiliation: INSERM UMR-S 976, Hôpital Saint-Louis, Paris, France.

ABSTRACT
EMMPRIN/CD147 is mainly known for its protease inducing function but a role in promoting tumor angiogenesis has also been demonstrated. This study provides evidence that EMMPRIN is a new coreceptor for the VEGFR-2 tyrosine kinase receptor in both endothelial and tumor cells, as it directly interacts with it and regulates its activation by its VEGF ligand, signalling and functional consequences both in vitro and in vivo. Computational docking analyses and mutagenesis studies identified a molecular binding site in the extracellular domain of EMMPRIN located close to the cell membrane and containing the amino acids 195/199. EMMPRIN is overexpressed in cancer and hence is able to further potentiate VEGFR-2 activation, suggesting that a combinatory therapy of an antiangiogenic drug together with an inhibitor of EMMPRIN/VEGFR-2 interaction may have a greater impact on inhibiting angiogenesis and malignancy.

No MeSH data available.


Related in: MedlinePlus