Transformation of Chicken Embryo Fibroblasts by v-src Uncouples β1 Integrin-Mediated Outside-in but Not Inside-out Signaling

Datta, Anirban; Shi, Qi Xian; Boettiger, David

doi:10.1128/mcb.21.21.7295-7306.2001

Cited by 23 publications

(19 citation statements)

References 73 publications

(99 reference statements)

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Section: Fak Y397 Phosphorylation Is a Read-out Of The Level Of Fibromentioning

confidence: 71%

“…There were no significant increases in the levels of phosphorylation of Y407 or Y577 as a function of increased fibronectin-bound ␣5␤1. No phosphorylation of Y925 was detected under these conditions although this antibody did detect pY925 in v-srctransformed cells (Datta et al, 2001).To determine the generality of these results, similar analyses were performed using IMR90 and HOS cells. Both these cell types gave essentially the same results, and those for HOS cells are shown in Figure 4.…”

mentioning

confidence: 79%

“…This provides a means for the cell to measure the mechanical and biochemical environment. The reason that we do not include Y861 as a primary measure of ␣5␤1 bonds is that the coupling between Y861 phosphorylation and the number of bonds was weaker in other cell types: including, chicken embryo fibroblasts (Datta et al, 2001), IMR90 human fibroblasts, and HOS osteosarcoma cells, and because it responded to alternate signaling mechanisms. When HT1080 cells were plated on fibronectin, both tethering and clustering occur as part of the normal process.…”

Section: Discussionmentioning

confidence: 99%

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A Novel Mode for Integrin-mediated Signaling: Tethering Is Required for Phosphorylation of FAK Y397

Shi

Boettiger

2003

MBoC

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133

113

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The common model for integrin mediated signaling is based on integrin clustering and the potential for that clustering to recruit signaling molecules including FAK and src. The clustering model for transmembrane signaling originated with the analysis of the EGF receptor signaling and remains the predominant model. The roles for substrate-bound ligand and ligand occupancy in integrin-mediated signaling are less clear. A kinetic model was established using HT1080 cells in which there was a linear relationship between the strength of adhesion, the proportion of ␣5␤1 integrin that could be chemically cross-linked, and the number of receptor-ligand bonds. This graded signal produced a similarly graded response measured by the level of specific phosphorylation of FAK Y397. FAK Y397 phosphorylation could also be induced by antibody bound to the substrate. In contrast, clustering of ␣5␤1 on suspended cells with either antibody to ␤1 or by clustering of soluble ligand bound to ␣5␤1 induced the phosphorylation of FAK Y861 but not Y397. There were no differences in signaling when activating antibodies were compared with blocking antibodies, presence or absence of ligand. Only tethering of ␣5␤1 to the substrate was required for induction of FAK Y397 phosphorylation. INTRODUCTIONThere are several classes of transmembrane receptors that are involved in the transmission of signals from the extracellular space across the plasma membrane. It is a logical requirement of these systems that ligand binding to the extracellular domain results in a change in the cytoplasmic domain. How the signal is transferred from the extracellular domain is basic to the generation of the intracellular downstream signals. Allosteric proteins have been described in which the occupation of a binding site on one side of the protein can result in a conformational change that affects other binding sites. The interposition of a lipid bilayer between the receiving and effector domains of the transmembrane receptors poses limitations in the application of the allosteric model. Although a few of the receptor tyrosine kinase receptor systems have been analyzed in detail, the mechanism for transmembrane signal transduction by other receptor classes is less well understood.The EGF receptor is the best understood model. Receptor dimerization is initiated by the binding of EGF to extracellular domain 1 altering the conformation of the extracellular domain to generate a dimerization of receptors mediated through domain 2. This dimerization brings the cytoplasmic domains of two EGF receptors into proximity and allows the cross-phosphorylation of cytoplasmic domains by the encoded tyrosine kinase (Schlessinger, 2000). The generation of signals through receptor dimerization is a general theme. Among the tyrosine kinase receptors the mechanisms of generating the dimer vary from the use of bivalent ligands for growth hormone and erythropoietin (Kossiakoff and de Vos, 1998;Jiang and Hunter, 1999), to dimeric ligands for PDGF and VEGF (Wiesmann et al., 1997), to complexe...

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Section: Fak Y397 Phosphorylation Is a Read-out Of The Level Of Fibromentioning

confidence: 71%

mentioning

confidence: 79%

Section: Discussionmentioning

confidence: 99%

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A Novel Mode for Integrin-mediated Signaling: Tethering Is Required for Phosphorylation of FAK Y397

Shi

Boettiger

2003

MBoC

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133

113

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“…V-Src-mediated phosphorylation of b1A integrin was found to occur on tyrosines 783 and 795 (Sakai et al, 2001). V-Src expression induced a reduction in adhesion of cells expressing wild-type b1A integrin, but could less effectively impair adhesion of cells expressing a b1A variant with both of these tyrosine residues mutated to phenylalanine (Datta et al, 2001). In the second, indirect mechanism of regulating adhesion, activated Src or v-Src interacts with and phosphorylates a member of the Ras family of GTPases (Zou et al, 2002).…”

Section: Integrins and Cadherinsmentioning

confidence: 96%

The interplay between Src and integrins in normal and tumor biology

2004

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Src family nonreceptor protein tyrosine kinases transduce signals that control normal cellular processes such as cell proliferation, adhesion and motility. Normally, cellular Src is held in an inactive state, but in several cancer types, abnormal events lead to elevated kinase activity of the protein and cause pleiotropic cellular responses inducing transformation and metastasis. A prerequisite of the ability of a cancer cell to undergo metastasis into distant tissues is to penetrate surrounding extracellular matrices. These processes are facilitated by the integrin family of cell adhesion molecules. As is the case with Src, altered integrin activity or substrate affinity can contribute to the neoplastic phenotype. Therefore, understanding the interplay between Src and integrin function has been of intense interest over the past few years. This review focuses on the role of Src and integrin signaling in normal cells and how this is deregulated in human cancer. We will identify the key players in the integrin-mediated signaling pathways involved in cell motility and apoptosis, such as FAK, paxillin and p130 CAS , and discuss how Src signaling affects the formation of focal adhesions and the extracellular matrix.

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“…Tyrosine phosphorylation of the NPXY motifs in integrin ␤ tails by Src family kinases is important in cell migration (41), hemostasis (42), and transformation (43). PTB domain recognition of peptide ligands can be promoted or inhibited by NPXY phosphorylation (21).…”

Section: Molecular Modeling and Mutagenesis Establish That Integrin ␤mentioning

confidence: 99%

Integrin β cytoplasmic domain interactions with phosphotyrosine-binding domains: A structural prototype for diversity in integrin signaling

Calderwood

Fujioka

Pereda

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

379

344

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The cytoplasmic domains (tails) of heterodimeric integrin adhesion receptors mediate integrins' biological functions by binding to cytoplasmic proteins. Most integrin ␤ tails contain one or two NPXY͞F motifs that can form ␤ turns. These motifs are part of a canonical recognition sequence for phosphotyrosine-binding (PTB) domains, protein modules that are present in a wide variety of signaling and cytoskeletal proteins. Indeed, talin and ICAP1-␣ bind to integrin ␤ tails by means of a PTB domain-NPXY ligand interaction. To assess the generality of this interaction we examined the binding of a series of recombinant PTB domains to a panel of short integrin ␤ tails. In addition to the known integrin-binding proteins, we found that Numb (a negative regulator of Notch signaling) and Dok-1 (a signaling adaptor involved in cell migration) and their isolated PTB domain bound to integrin tails. Furthermore, Dok-1 physically associated with integrin ␣IIb␤3. Mutations of the integrin ␤ tails confirmed that these interactions are canonical PTB domain-ligand interactions. First, the interactions were blocked by mutation of an NPXY motif in the integrin tail. Second, integrin class-specific interactions were observed with the PTB domains of Dab, EPS8, and tensin. We used this specificity, and a molecular model of an integrin ␤ tail-PTB domain interaction to predict critical interacting residues. The importance of these residues was confirmed by generation of gain-and loss-of-function mutations in ␤7 and ␤3 tails. These data establish that short integrin ␤ tails interact with a large number of PTB domain-containing proteins through a structurally conserved mechanism. I ntegrin adhesion receptors are heterodimers of ␣ and ␤ subunits, which combine to form a large extracellular domain, two transmembrane domains (one for each subunit), and a cytoplasmic domain typically composed of the short ␣ and ␤ C-terminal cytoplasmic tails (1). Bidirectional signal transduction through integrin adhesion receptors is essential for a wide variety of functions, including cell adhesion and migration, and assembly and remodeling of the extracellular matrix. Binding of intracellular proteins to integrin cytoplasmic tails is an important step in the transduction of signals to and from integrin-adhesion receptors (2). Integrin ␤ cytoplasmic tails, with the exception of those of ␤4 and ␤8, are short (Ͻ60 residues) and contain one or two NPXY or NPXY-like motifs (Fig. 1A), the first of which has the propensity to form a ␤ turn (3). Such ␤ turn-forming sequences frequently serve to bind to phosphotyrosine-binding (PTB) domains (4). NXXY motif-dependent binding of the Shc PTB domain to the large (Ͼ1,000 residues) ␤4 cytoplasmic tail has been observed (5) and molecular modeling studies suggested that the interaction of integrin cytoplasmic domain-associated protein (ICAP)1-␣ with ␤1A (6), and of talin with ␤3 (7), are mediated by PTB domain-like interactions. The solved crystal structure of a complex of a talin fragment with part of the ␤3 tail verified th...

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Transformation of Chicken Embryo Fibroblasts by v-src Uncouples β1 Integrin-Mediated Outside-in but Not Inside-out Signaling

Cited by 23 publications

References 73 publications

A Novel Mode for Integrin-mediated Signaling: Tethering Is Required for Phosphorylation of FAK Y397

A Novel Mode for Integrin-mediated Signaling: Tethering Is Required for Phosphorylation of FAK Y397

The interplay between Src and integrins in normal and tumor biology

Integrin β cytoplasmic domain interactions with phosphotyrosine-binding domains: A structural prototype for diversity in integrin signaling

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