The Shb adaptor protein causes Src-dependent cell spreading and activation of focal adhesion kinase in murine brain endothelial cells.
The mechanisms leading to focal adhesion kinase (FAK) activation remain obscure. We have investigated the role of the adaptor protein Shb in cell spreading and the regulation of FAK phosphorylation in immortalised brain endothelial (IBE) cells. Fibroblast growth factor 2 (FGF-2) stimulation lead to a direct association between Shb and FAK, which was mediated by the phosphotyrosine binding (PTB) domain of Shb. IBE cells overexpressing wild-type or R522K Shb (with an inactive Src homology 2 (SH2) domain) displayed increased FAK phosphorylation as well as enhanced spreading when seeded on collagen. FGF-2-induced tyrosine phosphorylation of Shb was dependent upon Src activity but independent of FAK activation. The use of Rat-1 fibroblasts overexpressing a temperature sensitive v-Src (tsLA29) confirmed that active Src enhanced Shb phosphorylation. The data indicate that Shb binds directly to FAK and regulates its phosphorylation leading to enhanced cell spreading in a Src-dependent manner.
Holmqvist
Kristina
K
Department of Medical Cell Biology, Biomedical Centre, Uppsala University, P.O. Box 571, S-751 23, Uppsala, Sweden.
Cross
Michael
M
Riley
Debbie
D
Welsh
Michael
M
D033942:Adaptor Proteins, Vesicular Transport / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000166:cytology / Q000201:enzymology*; D002478:Cells, Cultured; D003094:Collagen; D003598:Cytoskeletal Proteins / Q000378:metabolism; D004727:Endothelium / Q000166:cytology / Q000201:enzymology; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D015870:Gene Expression; D051379:Mice; D008822:Mice, Transgenic; D051419:Paxillin; D051966:Phospholipase C gamma; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000378:metabolism; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism*
D033942:Adaptor Proteins, Vesicular Transport; D003598:Cytoskeletal Proteins; D051419:Paxillin; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; C496062:Pxn protein, mouse; C085634:Shb protein, mouse; C490082:Shb protein, rat; D003094:Collagen; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; D019061:src-Family Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D020741:rho GTP-Binding Proteins