Protein tyrosine phosphatases (PTPases), such as SHP-1 and SHP-2, that contain Src homology 2 (SH2) domains play important roles in growth factor and cytokine signal transduction pathways. A protein of ϳ115 to 120 kDa that interacts with SHP-1 and SHP-2 was purified from v-src-transformed rat fibroblasts (SR-3Y1 cells), and the corresponding cDNA was cloned. The predicted amino acid sequence of the encoded protein, termed SHPS-1 (SHP substrate 1), suggests that it is a glycosylated receptor-like protein with three immunoglobulin-like domains in its extracellular region and four YXX(L/V/I) motifs, potential tyrosine phosphorylation and SH2-domain binding sites, in its cytoplasmic region. Various mitogens, including serum, insulin, and lysophosphatidic acid, or cell adhesion induced tyrosine phosphorylation of SHPS-1 and its subsequent association with SHP-2 in cultured cells. Thus, SHPS-1 may be a direct substrate for both tyrosine kinases, such as the insulin receptor kinase or Src, and a specific docking protein for SH2-domain-containing PTPases. In addition, we suggest that SHPS-1 may be a potential substrate for SHP-2 and may function in both growth factor-and cell adhesion-induced cell signaling.

Cellular regulation of the ligand binding affinity of integrin adhesion receptors (integrin activation) depends on the integrin ␤ cytoplasmic domains (tails). The head domain of talin binds to several integrin ␤ tails and activates integrins. This head domain contains a predicted FERM domain composed of three subdomains (F1, F2, and F3). An integrin-activating talin fragment was predicted to contain the F2 and F3 subdomains. Both isolated subdomains bound specifically to the integrin ␤ 3 tail. However, talin F3 bound the ␤ 3 tail with a 4-fold higher affinity than talin F2. Furthermore, expression of talin F3 (but not F2) in cells led to activation of integrin ␣ IIb ␤ 3 . A molecular model of talin F3 indicated that it resembles a phosphotyrosine-binding (PTB) domain. PTB domains recognize peptide ligands containing ␤ turns, often formed by NPXY motifs. NPX(Y/F) motifs are highly conserved in integrin ␤ tails, and mutations that disrupt this motif interfere with both integrin activation and talin binding. Thus, integrin binding to talin resembles the interactions of PTB domains with peptide ligands. These resemblances suggest that the activation of integrins requires the presence of a ␤ turn at NPX(Y/F) motifs conserved in integrin ␤ cytoplasmic domains.Integrin adhesion receptors are essential for the development and survival of multicellular animals. Normal functioning of the Ͼ20 human integrins often requires dynamic cellular regulation of integrin ligand binding affinity (integrin activation). Activation of integrins is important in many biological processes, including cell migration, hemostasis, extracellular matrix assembly, tumor metastasis, and the immune response (1, 2). Integrin ␣␤ heterodimers generally possess two short cytoplasmic domains (tails). The integrin ␤ tail plays a central role in the activation process, probably by undergoing regulated interactions with certain cytoplasmic proteins (1, 3).Talin is an abundant and widely expressed 250-kDa integrin ␤ tail-binding protein implicated in integrin activation (4). Talin is composed of a 50-kDa head and 205-kDa rod domain. The head domain contains a major integrin-binding site (5-7), and expression of a 1071-residue fragment of talin containing the head domain in cells leads to activation of integrin ␣ IIb ␤ 3 (5). The capacity of this fragment to activate integrin ␣ IIb ␤ 3 is lost when the head domain is deleted from it or when the ␤ 3 cytoplasmic domain is truncated (5). Talin binding to integrin ␤ tails can be regulated via calpain proteolysis (6) or through the binding of phosphoinositides (8). Furthermore, the phosphorylation of either talin and/or integrin (9, 10) could provide additional mechanisms for regulation of integrin-talin interactions. Thus, the talin head domain is implicated in integrin activation, and modulation of its binding to integrins is likely to contribute to the regulation of integrin activation.The talin head domain contains a predicted FERM domain (band four-point-one/ezrin/radixin/moesin homology domain) (11). FERM ...

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...

SH-PTP2 is a nontransmembrane human protein-tyrosine phosphatase that contains two Src homology 2 (SH2) domains and binds to insulin receptor substrate 1 (IRS-1) via these domains in response to insulin. The expression of a catalytically inactive mutant of SH-PTP2 (containing the mutation Cys-459--Ser) in Chinese hamster ovary cells that overexpress human insulin receptors (CHO-IR cells) markedly attenuated insulinstimulated Ras activation. Expression of mutant SH-PTP2 also inhibited MAP kinase activation in response to insulin but not in response to 12-0-tetradecanoyl phorbol-13-acetate. In contrast, the insulin-induced association of phosphoinositide 3-kinase activity with IRS-1 was not affected by the expression of inactive SH-PTP2. Furthermore, the expression of mutant SH-PTP2 had no effect on the binding of Grb2 to IRS-1, on the tyrosine phosphorylation of Shc, or on the formation of the complex between Shc and Grb2 in response to insulin. However, the amount of SH-PTP2 bound to IRS-1 in insulin-treated CHO-IR cells expressing mutant SH-PTP2 was greater than that observed in CHO-IR cells overexpressing wild-type SH-PTP2. Recombinant SH-PTP2 specifically dephosphorylated a synthetic phosphopeptide corresponding to the sequence surrounding Tyr-1172 of IRS-1, a putative binding site for SH-PTP2. Additionally, phenylarsine oxide, an inhibitor of protein-tyrosine phosphatases, inactivated SH-PTP2 in vitro and increased the insulin-induced association of SH-PIP2 with IRS-1. These results suggest that SH-PTP2 may regulate an upstream element necessary for Ras activation in response to insulin and that this upstream element may be required for the Grb2-or Shc-dependent pathway. Furthermore, these results are consistent with the notion that SH-PTP2 may bind to IRS-1 through its SH2 domains in response to insulin and dephosphorylate the phosphotyrosine residue to which it binds, thereby regulating its association with IRS-1.

No abstract