Thrombospondin stimulates focal adhesion disassembly through Gi- and phosphoinositide 3-kinase-dependent ERK activation.
The matricellular protein thrombospondin (TSP) stimulates stress fiber and focal adhesion disassembly through a sequence (hep I) in its heparin-binding domain. TSP/hep I signals focal adhesion disassembly by binding cell surface calreticulin (CRT) and activating phosphoinositide 3-kinase (PI3K). However, other components of this signaling pathway have not been identified. We now show that TSP induces focal adhesion disassembly through activation of pertussis toxin (PTX)-sensitive G proteins and ERK phosphorylation. PTX pretreatment inhibits TSP/hep I-mediated focal adhesion disassembly as well as PI3K activation. In addition, membrane-permeable Galpha(i2)- and Gbetagamma-blocking peptides inhibit hep I-mediated focal adhesion disassembly. Hep I stimulates a transient increase in ERK activation, which is abrogated by both PTX and PI3K inhibitors. Inhibiting ERK activation with MEK inhibitors blocks hep I-mediated focal adhesion disassembly, indicating that ERK activation is required for cytoskeletal reorganization. G protein signals and ERK phosphorylation are induced by TSP binding to cell surface CRT, because CRT null mouse embryonic fibroblasts (MEF) fail to stimulate ERK phosphorylation in response to TSP/hep I treatment. These data show that G(i) protein and ERK, in concert with PI3K, are stimulated by TSP.CRT interactions at the cell surface to induce de-adhesive changes in the cytoskeleton.
Orr
Anthony Wayne
AW
Department of Pathology, University of Alabama, Birmingham, Alabama 35294-0019, USA.
Pallero
Manuel Antonio
MA
Murphy-Ullrich
Joanne E
JE
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002448:Cell Adhesion / Q000502:physiology*; D002460:Cell Line; D004789:Enzyme Activation; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008969:Molecular Sequence Data; D037342:Pertussis Toxin; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D019699:Thrombospondins / Q000502:physiology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D019699:Thrombospondins; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D019869:Phosphatidylinositol 3-Kinases; D020928:Mitogen-Activated Protein Kinases; D020962:Heterotrimeric GTP-Binding Proteins