Role of tyrosine phosphatase SHP-1 in the mechanism of endorepellin angiostatic activity.
Endorepellin, the C-terminal domain of perlecan, is a powerful angiogenesis inhibitor. To dissect the mechanism of endorepellin-mediated endothelial silencing, we used an antibody array against multiple tyrosine kinase receptors. Endorepellin caused a widespread reduction in phosphorylation of key receptors involved in angiogenesis and a concurrent increase in phosphatase activity in endothelial cells and tumor xenografts. These effects were efficiently hampered by function-blocking antibodies against integrin alpha2beta1, the functional endorepellin receptor. The Src homology-2 protein phosphatase-1 (SHP-1) coprecipitated with integrin alpha2 and was phosphorylated in a dynamic fashion after endorepellin stimulation. Genetic evidence was provided by lack of an endorepellin-evoked phosphatase response in microvascular endothelial cells derived from integrin alpha2beta1(-/-) mice and by response to endorepellin in cells genetically engineered to express the alpha2beta1 integrin, but not in cells either lacking this receptor or expressing a chimera harboring the integrin alpha2 ectodomain fused to the alpha1 intracellular domain. siRNA-mediated knockdown of integrin alpha2 caused a dose-dependent reduction of SHP-1. Finally, the levels of SHP-1 and its enzymatic activity were substantially reduced in multiple organs from alpha2beta1(-/-) mice. Our results show that SHP-1 is an essential mediator of endorepellin activity and discover a novel functional interaction between the integrin alpha2 subunit and SHP-1.
Department of Pathology, Anatomy, and Cell Biology, and Cancer Cell Biology and Signaling Program, Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, Philadelphia, PA 19107, USA.
Alexander
A
Nyström
Zabeena P
ZP
Shaik
Donald
D
Gullberg
Thomas
T
Krieg
Beate
B
Eckes
Roy
R
Zent
Ambra
A
Pozzi
Renato V
RV
Iozzo
D000818:Animals; D018827:Carcinoma, Lewis Lung / Q000098:blood supply; D002448:Cell Adhesion / Q000187:drug effects; D004730:Endothelium, Vascular / Q000502:physiology; D019812:Heparan Sulfate Proteoglycans / Q000494:pharmacology / Q000502:physiology*; D006801:Humans; D039423:Integrin alpha1 / Q000235:genetics; D038982:Integrin alpha2beta1 / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009389:Neovascularization, Pathologic / Q000188:drug therapy; D018919:Neovascularization, Physiologic / Q000502:physiology*; D010446:Peptide Fragments / Q000494:pharmacology / Q000502:physiology*; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000032:analysis; D025941:Protein Interaction Mapping; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017434:Protein Structure, Tertiary; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6 / Q000737:chemistry / Q000172:deficiency / Q000378:metabolism / Q000502:physiology*; D020794:Receptor Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000502:physiology; D023041:Xenograft Model Antitumor Assays
D019812:Heparan Sulfate Proteoglycans; D039423:Integrin alpha1; D038982:Integrin alpha2beta1; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C071927:perlecan; D019000:Phosphotyrosine; D020794:Receptor Protein-Tyrosine Kinases; C509249:PTPN6 protein, human; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; C509248:Ptpn6 protein, mouse