title: Integrin-dependent control of inositol lipid synthesis in vascular endothelial cells and smooth muscle cells.
abstract: Extracellular matrix (ECM) molecules, such as fibronectin (FN), regulate fibroblast sensitivity to soluble growth factors, in part, by controlling cellular levels of phosphatidylinositol bis-phosphate (PIP2), the substrate for phospholipase C-gamma (McNamee et al., 1993, J. Cell Biol. 121, 673-678). In the present study, we extended these investigations by exploring whether cells of the vascular wall also exhibit this response and analyzing the mechanism by which adhesion to ECM regulates intracellular PIP2 mass. Capillary endothelial cells, pulmonary vascular smooth muscle cells, and C3H 101/2 fibroblasts were all found to exhibit a similar two- to threefold increase in PIP2 mass within 3 h after binding to dishes coated with FN. Furthermore, similar effects were observed using dishes coated with a variety of different ECM molecules, including collagen types I and IV as well as a synthetic RGD-containing peptide. An increase in PIP2 mass also was produced when suspended cells bound to microbeads (4.5 micron diameter; coated with RGD-peptide or anti-integrin beta 1 antibody) that induce local integrin clustering and focal adhesion formation, independently of cell spreading. In contrast, neither binding of soluble FN nor binding of microbeads coated with ligands for other transmembrane surface receptors (e.g., acetylated low-density lipoprotein, antibodies against heparan sulfate) had any effect on PIP2 mass. While these results suggest that integrin clustering stimulates PIP2 synthesis, no change in total cellular or cytoskeletal-associated phosphatidylinositol-4-phosphate kinase (PIP kinase) activity could be detected when cells bound to immobilized integrin ligands. However, when focal adhesion complexes were isolated from these cells using a magnetic procedure (G. Plopper and D. E. Ingber, 1993, Biochem. Biophys. Res. Commun. 193, 571-578), this subfraction of the cytoskeleton was found to be enriched for PIP kinase activity by more than twofold relative to the whole cytoskeleton. These data suggest that ECM binding may increase PIP2 mass in vascular cells by clustering cell surface integrin receptors and activating cytoskeletal-associated PIP kinases locally within the focal adhesion complex.
authors.affiliation: Department of Pathology, Children's Hospital and Harvard Medical School, Boston Massachusetts 02115, USA.
authors.forename: H P
authors.initials: HP
authors.lastname: McNamee
authors.forename: H G
authors.initials: HG
authors.lastname: Liley
authors.forename: D E
authors.initials: DE
authors.lastname: Ingber
mesh_terms: D000818:Animals; D002417:Cattle; D002448:Cell Adhesion; D004730:Endothelium, Vascular / Q000166:cytology / Q000254:growth & development*; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D019012:Integrin beta1 / Q000378:metabolism*; D051379:Mice; D024510:Muscle Development; D009130:Muscle, Smooth / Q000166:cytology / Q000254:growth & development*; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates / Q000096:biosynthesis*; D015398:Signal Transduction
chemical_list: D016326:Extracellular Matrix Proteins; D019012:Integrin beta1; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates