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Contact inhibition of VEGF-induced proliferation requires vascular endothelial cadherin, beta-catenin, and the phosphatase DEP-1/CD148.

Grazia Lampugnani M, Zanetti A, Corada M, Takahashi T, Balconi G, Breviario F, Orsenigo F, Cattelino A, Kemler R, Daniel TO, Dejana E - J. Cell Biol. (2003)

Bottom Line: Comparing isogenic endothelial cells differing for vascular endothelial cadherin (VE-cadherin) expression only, we found that the presence of this protein attenuates VEGF-induced VEGF receptor (VEGFR) 2 phosphorylation in tyrosine, p44/p42 MAP kinase phosphorylation, and cell proliferation.A dominant-negative mutant of high cell density-enhanced PTP 1 (DEP-1)//CD148 as well as reduction of its expression by RNA interference partially restore VEGFR-2 phosphorylation and MAP kinase activation.In sparse cells or in VE-cadherin- cells, this phenomenon cannot occur and the receptor is fully activated by the growth factor.

View Article: PubMed Central - PubMed

Affiliation: FIRC Institute of Molecular Oncology, 20139 Milan, Italy.

ABSTRACT
Confluent endothelial cells respond poorly to the proliferative signals of VEGF. Comparing isogenic endothelial cells differing for vascular endothelial cadherin (VE-cadherin) expression only, we found that the presence of this protein attenuates VEGF-induced VEGF receptor (VEGFR) 2 phosphorylation in tyrosine, p44/p42 MAP kinase phosphorylation, and cell proliferation. VE-cadherin truncated in beta-catenin but not p120 binding domain is unable to associate with VEGFR-2 and to induce its inactivation. beta-Catenin- endothelial cells are not contact inhibited by VE-cadherin and are still responsive to VEGF, indicating that this protein is required to restrain growth factor signaling. A dominant-negative mutant of high cell density-enhanced PTP 1 (DEP-1)//CD148 as well as reduction of its expression by RNA interference partially restore VEGFR-2 phosphorylation and MAP kinase activation. Overall the data indicate that VE-cadherin-beta-catenin complex participates in contact inhibition of VEGF signaling. Upon stimulation with VEGF, VEGFR-2 associates with the complex and concentrates at cell-cell contacts, where it may be inactivated by junctional phosphatases such as DEP-1. In sparse cells or in VE-cadherin- cells, this phenomenon cannot occur and the receptor is fully activated by the growth factor.

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Related in: MedlinePlus

Inhibition of DEP-1 enhances phosphorylation of p44/42 MAP kinase and BrdU nuclear incorporation in endothelial cells. (A) Phosphorylation of p44/42 MAP kinase in response to VEGF (80 ng/ml for 10 min) is increased by 80–90% in VEC-positive cells expressing DEP C/S in comparison with control VEC positive. A similar increase is observed after DEP siRNA transfection of the cells. The graph represents the ratio between phosphorylated and total p44/42 MAP kinase as fold increase over untreated VEC- cells. The values are means ± SD of four independent experiments. Plus and minus indicate the presence or the absence of the protein indicated on the left. (B) Nuclear incorporation of BrdU was stimulated by 80–100% in VEC-positive cells expressing C/S DEP or after DEP siRNA. BrdU incorporation was assayed as in Fig. 1. Data are means ± SD of four experiments.
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fig10: Inhibition of DEP-1 enhances phosphorylation of p44/42 MAP kinase and BrdU nuclear incorporation in endothelial cells. (A) Phosphorylation of p44/42 MAP kinase in response to VEGF (80 ng/ml for 10 min) is increased by 80–90% in VEC-positive cells expressing DEP C/S in comparison with control VEC positive. A similar increase is observed after DEP siRNA transfection of the cells. The graph represents the ratio between phosphorylated and total p44/42 MAP kinase as fold increase over untreated VEC- cells. The values are means ± SD of four independent experiments. Plus and minus indicate the presence or the absence of the protein indicated on the left. (B) Nuclear incorporation of BrdU was stimulated by 80–100% in VEC-positive cells expressing C/S DEP or after DEP siRNA. BrdU incorporation was assayed as in Fig. 1. Data are means ± SD of four experiments.

Mentions: Both DEP-1 C/S and siRNA had a partial, but significant, effect in restoring MAP kinase activation and BrdU uptake induced by VEGF in VEC-positive cells (Fig. 10, A and B). Wild-type DEP-1 (DEPwt) and the scrambled RNA did not have any effect.


Contact inhibition of VEGF-induced proliferation requires vascular endothelial cadherin, beta-catenin, and the phosphatase DEP-1/CD148.

Grazia Lampugnani M, Zanetti A, Corada M, Takahashi T, Balconi G, Breviario F, Orsenigo F, Cattelino A, Kemler R, Daniel TO, Dejana E - J. Cell Biol. (2003)

Inhibition of DEP-1 enhances phosphorylation of p44/42 MAP kinase and BrdU nuclear incorporation in endothelial cells. (A) Phosphorylation of p44/42 MAP kinase in response to VEGF (80 ng/ml for 10 min) is increased by 80–90% in VEC-positive cells expressing DEP C/S in comparison with control VEC positive. A similar increase is observed after DEP siRNA transfection of the cells. The graph represents the ratio between phosphorylated and total p44/42 MAP kinase as fold increase over untreated VEC- cells. The values are means ± SD of four independent experiments. Plus and minus indicate the presence or the absence of the protein indicated on the left. (B) Nuclear incorporation of BrdU was stimulated by 80–100% in VEC-positive cells expressing C/S DEP or after DEP siRNA. BrdU incorporation was assayed as in Fig. 1. Data are means ± SD of four experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199373&req=5

fig10: Inhibition of DEP-1 enhances phosphorylation of p44/42 MAP kinase and BrdU nuclear incorporation in endothelial cells. (A) Phosphorylation of p44/42 MAP kinase in response to VEGF (80 ng/ml for 10 min) is increased by 80–90% in VEC-positive cells expressing DEP C/S in comparison with control VEC positive. A similar increase is observed after DEP siRNA transfection of the cells. The graph represents the ratio between phosphorylated and total p44/42 MAP kinase as fold increase over untreated VEC- cells. The values are means ± SD of four independent experiments. Plus and minus indicate the presence or the absence of the protein indicated on the left. (B) Nuclear incorporation of BrdU was stimulated by 80–100% in VEC-positive cells expressing C/S DEP or after DEP siRNA. BrdU incorporation was assayed as in Fig. 1. Data are means ± SD of four experiments.
Mentions: Both DEP-1 C/S and siRNA had a partial, but significant, effect in restoring MAP kinase activation and BrdU uptake induced by VEGF in VEC-positive cells (Fig. 10, A and B). Wild-type DEP-1 (DEPwt) and the scrambled RNA did not have any effect.

Bottom Line: Comparing isogenic endothelial cells differing for vascular endothelial cadherin (VE-cadherin) expression only, we found that the presence of this protein attenuates VEGF-induced VEGF receptor (VEGFR) 2 phosphorylation in tyrosine, p44/p42 MAP kinase phosphorylation, and cell proliferation.A dominant-negative mutant of high cell density-enhanced PTP 1 (DEP-1)//CD148 as well as reduction of its expression by RNA interference partially restore VEGFR-2 phosphorylation and MAP kinase activation.In sparse cells or in VE-cadherin- cells, this phenomenon cannot occur and the receptor is fully activated by the growth factor.

View Article: PubMed Central - PubMed

Affiliation: FIRC Institute of Molecular Oncology, 20139 Milan, Italy.

ABSTRACT
Confluent endothelial cells respond poorly to the proliferative signals of VEGF. Comparing isogenic endothelial cells differing for vascular endothelial cadherin (VE-cadherin) expression only, we found that the presence of this protein attenuates VEGF-induced VEGF receptor (VEGFR) 2 phosphorylation in tyrosine, p44/p42 MAP kinase phosphorylation, and cell proliferation. VE-cadherin truncated in beta-catenin but not p120 binding domain is unable to associate with VEGFR-2 and to induce its inactivation. beta-Catenin- endothelial cells are not contact inhibited by VE-cadherin and are still responsive to VEGF, indicating that this protein is required to restrain growth factor signaling. A dominant-negative mutant of high cell density-enhanced PTP 1 (DEP-1)//CD148 as well as reduction of its expression by RNA interference partially restore VEGFR-2 phosphorylation and MAP kinase activation. Overall the data indicate that VE-cadherin-beta-catenin complex participates in contact inhibition of VEGF signaling. Upon stimulation with VEGF, VEGFR-2 associates with the complex and concentrates at cell-cell contacts, where it may be inactivated by junctional phosphatases such as DEP-1. In sparse cells or in VE-cadherin- cells, this phenomenon cannot occur and the receptor is fully activated by the growth factor.

Show MeSH
Related in: MedlinePlus