Syndecan-1 regulates αvβ3 and αvβ5 integrin activation during angiogenesis and is blocked by synstatin, a novel peptide inhibitor

Abstract

Syndecan-1 (Sdc1) is a matrix receptor shown to associate via its extracellular domain with the αvβ3 and αvβ5 integrins, potentially regulating cell adhesion, spreading, and invasion of cells expressing these integrins. Using Sdc1 deletion mutants expressed in human mammary carcinoma cells, we identified the active site within the Sdc1 core protein and derived a peptide inhibitor called synstatin (SSTN) that disrupts Sdc1's interaction with these integrins. Because the αvβ3 and αvβ5 integrins are critical in angiogenesis, a process in which a role for Sdc1 has been uncertain, we used human vascular endothelial cells in vitro to show that the Sdc1 regulatory mechanism is also required for integrin activation on these cells. We found Sdc1 expressed in the vascular endothelium during microvessel outgrowth from aortic explants in vitro and in mouse mammary tumors in vivo. Moreover, we show that SSTN blocks angiogenesis in vitro or when delivered systemically in a mouse model of angiogenesis in vivo, and impairs mammary tumor growth in an orthotopic mouse tumor model. Thus, Sdc1 is a critical regulator of these two important integrins during angiogenesis and tumorigenesis, and is inhibited by the novel SSTN peptide.

Syndecan-1 in Breast Cancer Stroma Fibroblasts Regulates Extracellular Matrix Fiber Organization and Carcinoma Cell Motility

“…The Sdc1 regulates the activities of several integrins, including ␣v␤3, ␣v␤5, and ␤4 integrins. [43][44][45][46] Although the FN matrix assembly appears to be initiated mainly by Arg-Gly-Asp-binding integrin ␣5␤1, 47,48 alternative FN-binding integrin receptors (eg, ␣v␤3, ␣4␤1, and ␣v␤1) can also support this process once properly activated. 33,49 -52 It is possible that Sdc1 induction activates integrins other than ␣5␤1 and initiates an alternative assembly pathway, thus resulting in an ECM that is structurally/compositionally different from that assembled under the control of ␣5␤1.…”

Syndecan-1 couples the insulin-like growth factor-1 receptor to inside-out integrin activation

“…Sdc1 regulates the activation of avb3 and avb5 integrins on mammary carcinoma cells, fibroblasts and endothelial cells Beauvais et al, 2009;Beauvais and Rapraeger, 2003;McQuade et al, 2006). Activation of avb3 and/or avb5 integrin stimulates adhesion, spreading and migration of tumor and endothelial cells, with clear consequences on tumor progression.…”

Interaction of Syndecan and α6β4 Integrin Cytoplasmic Domains

“…Co-immunoprecipitation Assays-Immunoprecipitations were carried in a manner similar to that of Beauvais et al (30,31). Cells (8 ϫ 10 6 ) were washed once with washing buffer (50 mM Hepes, 50 mM NaCl and 10 mM EDTA, pH 7.4) and then lysed for 20 min on ice in 1% Triton X-100 containing a 1:1000 dilution of protease inhibitor mixture set III (Calbiochem) in washing buffer.…”

“…To measure cell surface syndecan expression, suspended cells were incubated for 1 h on ice with 1 g of primary antibody per 3 ϫ 10 5 cells and then washed and counterstained with Alexa-488-conjugated secondary antibodies and scanned on a FACSCalibur bench top cytometer. Cell scatter and propidium iodide staining profiles were used to gate live, single-cell events for data analysis (30,31).…”