The focal adhesion kinase p125Fak is a widely expressed cytosolic tyrosine kinase, which is involved in integrin signaling and in signal transduction of a number of growth factors. In contrast to tyrosine kinase receptors such as the platelet-derived growth factor and the hepatocyte growth factor receptors, which induce p125Fak phosphorylation, insulin has been shown to promote its dephosphorylation. Fak is dependent on the cell architecture, and hence the interaction of the insulin/IGF-I signaling system with the integrin system will vary accordingly. The focal adhesion kinase p125Fak is a cytosolic tyrosine kinase initially isolated from Src-transformed cells (1, 2). It is localized at focal adhesion plaques of cultured cells and binds to a number of proteins involved in the organization of the cytoskeleton and to signaling molecules, resulting in the formation of multicomponents complexes (reviewed in Refs. 3-6). Tyrosine phosphorylation of p125Fak occurs rapidly in response to integrin clustering or binding to the extracellular matrix, and this correlates with increased kinase activity (2, 7-9). For most cell types, cooperation of adhesion-mediated and growth factor-mediated signaling pathways is required for appropriate growth control, and it is now widely accepted that p125Fak may be a point of convergence in the actions of integrins and growth factors (10, 11). Indeed, p125Fak is not only activated by integrins, but also by mitogenic neuropeptides (12), thrombin (13), sphingosine (14), the bioactive lipid lysophosphatidic acid (15-17), and by ligands for tyrosine kinase receptors such as platelet-derived growth factor and hepatocyte growth factor (18 -20).In contrast to other tyrosine kinase receptors, which induce tyrosine phosphorylation of p125Fak , it has been reported that in fibroblasts insulin promotes a decrease in p125Fak phosphorylation (21-23).The insulin receptor is a heterotetrameric oligomer consisting of two extracellular 135-kDa ␣-subunits and two 95-kDa transmembrane ␤-subunits containing a tyrosine kinase (24,25). Insulin binding to the ␣-subunit stimulates autophosphorylation of the ␤-subunit cytoplasmic domain on multiple tyrosine residues. This activates the receptor kinase, leading to phosphorylation of several substrates including IRS-1, 1 IRS-2, Shc, and Gab-1 (26 -31). IRS-1/2 and Gab-1 carry multiple potential tyrosine phosphorylation sites, which upon phosphorylation become docking sites for SH2 domain-containing proteins. These include the p85 regulatory subunit of phosphatidylinositol 3-kinase, Grb2, and the phosphotyrosine phosphatase SHP-2 (32-34).IRS-1 is also found to be associated with Csk, the C-terminal Src kinase that is an inhibitor of the Src kinase family. It has been shown recently that the insulin-induced complex of IRS-1 and Csk could be involved in dephosphorylation of p125 Fak observed after insulin treatment of fibroblasts (35).The fact that insulin induces p125 Fak dephosphorylation suggests the existence of an antagonistic action of insulin on the integrin s...

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The cardiac remodeling process that accompanies the development of POH is a phase-dependent process associated with progressive deterioration of cardiac function.

3H-Labelled kappa-elastin peptides (kE:75 kDa molecular weight) were shown to bind to confluent human skin fibroblast (HSF) cultures in a time-dependent and saturable manner. Scatchard analysis indicated the presence of high affinity binding sites with kD = 2.7 x 10(-10) M and 19,000 sites per cell. Binding of kE to its receptor on HSF accelerates and intensifies the adhesion of insoluble elastin fibres (iE) to confluent HSF. Optimal effect was attained for a kE concentration of 0.3 x 10(-9) M close to kD. This stimulatory effect of kE on the binding of iE to HSF could be inhibited by neomycin, retinal and pertussis toxin, substances which act at different levels of the transduction mechanism following the activation of the receptor and the subsequent triggering of cell biological events (chemotaxis, modification of calcium fluxes). The stimulation of iE adhesion to HSF induced by kE as well as kE binding to the cells could be inhibited by lactose and laminin but not by Arg-Gly-Asp-Ser(RGDS) peptides. This indicates that the elastin peptide receptor on HSF possesses lectin-like properties and shares homology with the laminin receptor as also shown for other cell types. None of the substances tested, that is inhibitors of the transduction mechanism, lactose, laminin and Arg-Gly-Asp-Ser(RGDS) peptides were shown to interfere significantly with the binding of iE (in the absence of added kE) to confluent HSF. The proteins adhering strongly to elastin fibres were isolated by a sequential extraction procedure and the final hydrochloride guanidinium-DTT extract was analysed by SDS-PAGE under reducing conditions, Western blots using specific antibodies against several connective tissue proteins and affinity for [3H]-kE following nitrocellulose electro-transfer of proteins. Fibronectin, vitronectin, tropoelastin(s), and a 120 kDa cysteine rich glycoprotein previously designated as elastonectin were identified. Among these proteins, [3H]-kE was found to bind exclusively to a 65 kDa protein that could be eluted selectively from elastin fibres with a neutral buffer containing 100 mM lactose. Therefore the elastin peptide receptor on human skin fibroblasts shares properties with the elastin receptor characterized from other cell types. Conformational differences between elastin peptides and elastin fibres could explain the differences in the mechanisms of interactions between elastin fibres and elastin peptides with HSF in culture. The stimulatory effect of elastin-derived peptides on the adhesion of elastin fibres to HSF could have implications in the oriented biosynthesis of elastin fibres.