Adaptor protein-2 exhibits alpha 1 beta 1 or alpha 6 beta 1 integrin-dependent redistribution in rhabdomyosarcoma cells

Downregulation of several signaling pathways, such as those stimulated by growth factor receptors, occurs by internalization of signaling receptors through clathrin-coated pits. The first step in internalization or endocytosis is interaction with AP-2, which results in coated pit formation by assembly of clathrin to AP-2. Changes in endocytosis are reflected in the distribution of AP-2 molecules at the cell surface. Integrins are receptors which mediate attachment to the extracellular matrix and also stimulate numerous intracellular signaling pathways; however, it is not known how signaling through integrins is terminated or downregulated. Endocytosis through clathrin-coated pits offers an attractive mechanism for this. This work explores the relationship between AP-2 and beta(1) integrins. RD cells grown for 24 h on collagen or laminin exhibit a redistribution of AP-2 to the cell periphery relative to those grown on fibronectin or polylysine. The total AP-2 protein levels in the cells are unaffected. Blocking alpha(1)beta(1) integrin ligand binding on collagen prevents this redistribution fully. On laminin where alpha(1)beta(1) and alpha(6)beta(1) integrins are engaged, both receptors must be simultaneously blocked to prevent AP-2 redistribution, confirming that the redistribution depends on the specific engagement of the receptors. Immunofluorescence reveals that the majority of alpha(1)beta(1) integrins colocalize with alpha(6)beta(1) integrins in linear structures identified as focal adhesions. A separate fraction of alpha(1)beta(1) integrins colocalize with AP-2 in coated pits. Interestingly, alpha(6)beta(1) integrins are not located in coated pits, demonstrating that integrin colocalization with AP-2 is not necessary to induce redistribution of AP-2.     

Activity-dependent regulation of synaptic AMPA receptor composition and abundance by beta3 integrins

At synapses, cell adhesion molecules (CAMs) provide the molecular framework for coordinating signaling events across the synaptic cleft. Among synaptic CAMs, the integrins, receptors for extracellular matrix proteins and counterreceptors on adjacent cells, are implicated in synapse maturation and plasticity and memory formation. However, little is known about the molecular mechanisms of integrin action at central synapses. Here, we report that postsynaptic beta3 integrins control synaptic strength by regulating AMPA receptors (AMPARs) in a subunit-specific manner. Pharmacological perturbation targeting beta3 integrins promotes endocytosis of GluR2-containing AMPARs via Rap1 signaling, and expression of beta3 integrins produces robust changes in the abundance and composition of synaptic AMPARs without affecting dendritic spine structure. Importantly, homeostatic synaptic scaling induced by activity deprivation elevates surface expression of beta3 integrins, and in turn, beta3 integrins are required for synaptic scaling. Our findings demonstrate a key role for integrins in the feedback regulation of excitatory synaptic strength.     

beta1 integrins expression in adult rat ventricular myocytes and its role in the regulation of beta-adrenergic receptor-stimulated apoptosis

We have shown that the stimulation of beta-adrenergic receptors (beta-AR) increases apoptosis in adult rat ventricular myocytes (ARVMs). Integrins, a family of alphabeta-heterodimeric cell surface receptors, are postulated to play a role in ventricular remodeling. Here, we show that norepinephrine (NE) increases beta1 integrins expression in ARVMs via the stimulation of alpha1-AR, not beta-AR. Inhibition of ERK1/2 using PD 98059, an inhibitor of ERK1/2 pathway, inhibited alpha1-AR-stimulated increases in beta1 integrins expression. Activation of beta1 integrins signaling pathway using laminin (LN) inhibited beta-AR-stimulated apoptosis as measured by terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL)-staining and flow cytometry. Likewise, ligation of beta1 integrins with anti-beta1 integrin antibodies prevented beta-AR-stimulated apoptosis. Treatment of cells using LN or anti-beta1 integrin antibodies activated ERK1/2 pathway. PD 98059 inhibited activation of ERK1/2 by LN, and prevented the anti-apoptotic effects of LN. Thus (1) stimulation of alpha1-AR regulates beta1 integrins expression via the activation of ERK1/2, (2) beta1 integrins signaling protects ARVMs from beta-AR-stimulated apoptosis, (3) activation of ERK1/2 plays a critical role in the anti-apoptotic effects of beta1-integrin signaling. These data suggest that beta1 integrin signaling protects ARVMs against beta-AR-stimulated apoptosis possibly via the involvement of ERK1/2.     

Characterization of the integrin alpha v beta 6 as a fibronectin-binding protein.

Integrins are a complex family of divalent cation-dependent cell adhesion receptors composed of one alpha and one beta subunit noncovalently bound to one another. A subset of integrins contains the alpha v subunit in association with one of several beta subunits (e.g. beta 3, beta 5, beta 1). We have recently identified a novel integrin beta subunit, beta 6, that is present in a number of epithelial cell lines. Using a polyclonal antibody raised against the carboxyl-terminal peptide of beta 6, we have now identified the integrin heterodimer, alpha v beta 6, on the surface of two human carcinoma cell lines. Using affinity chromatography of lysates from the pancreatic carcinoma cell line, FG-2, we demonstrate that alpha v beta 6 binds to fibronectin, but not to vitronectin or collagen I. In contrast, the alpha v beta 5 integrin, which is also expressed on FG-2 cells, binds exclusively to vitronectin. Immobilized collagen I does not interact with alpha v integrins, but binds beta 1-containing integrins. Both alpha v beta 6 and alpha v beta 5 are eluted from their respective immobilized ligands by a hexa-peptide containing the sequence Arg-Gly-Asp (RGD). RGD is highly effective in the presence of Ca2+, somewhat less effective in Mg2+, and virtually inactive in Mn2+. These results suggest that alpha v beta 6 functions as an RGD-dependent fibronectin receptor in FG-2 carcinoma cells. In agreement with this notion, cell adhesion assays show that FG-2 cell attachment to fibronectin is only partially inhibited by anti-beta 1 integrin antibodies, implying that other fibronectin receptors may be involved. Taken together with recent reports on the vitronectin receptor function of alpha v beta 5, our results suggest that the previously described carcinoma cell integrin, alpha v beta x (Cheresh, D. A., Smith, J. W., Cooper, H. M., and Quaranta, V. (1989) Cell 57, 59-69), is a mixture of at least two different receptors: alpha v beta 5, mediating adhesion to vitronectin, and alpha v beta 6, mediating adhesion to fibronectin.     

Sphingosine 1-phosphate-induced endothelial cell migration requires the expression of EDG-1 and EDG-3 receptors and Rho-dependent activation of alpha vbeta3- and beta1-containing integrins

Sphingosine 1-phosphate (SPP), a platelet-derived bioactive lysophospholipid, is a regulator of angiogenesis. However, molecular mechanisms involved in SPP-induced angiogenic responses are not fully defined. Here we report the molecular mechanisms involved in SPP-induced human umbilical vein endothelial cell (HUVEC) adhesion and migration. SPP-induced HUVEC migration is potently inhibited by antisense phosphothioate oligonucleotides against EDG-1 as well as EDG-3 receptors. In addition, C3 exotoxin blocked SPP-induced cell attachment, spreading and migration on fibronectin-, vitronectin- and Matrigel-coated surfaces, suggesting that endothelial differentiation gene receptor signaling via the Rho pathway is critical for SPP-induced cell migration. Indeed, SPP induced Rho activation in an adherence-independent manner, whereas Rac activation was dispensible for cell attachment and focal contact formation. Interestingly, both EDG-1 and -3 receptors were required for Rho activation. Since integrins are critical for cell adhesion, migration, and angiogenesis, we examined the effects of blocking antibodies against alpha(v)beta(3), beta(1), or beta(3) integrins. SPP induced Rho-dependent integrin clustering into focal contact sites, which was essential for cell adhesion, spreading and migration. Blockage of alpha(v)beta(3)- or beta(1)-containing integrins inhibited SPP-induced HUVEC migration. Together our results suggest that endothelial differentiation gene receptor-mediated Rho signaling is required for the activation of integrin alpha(v)beta(3) as well as beta(1)-containing integrins, leading to the formation of initial focal contacts and endothelial cell migration.     

Specification of the direction of adhesive signaling by the integrin beta cytoplasmic domain

Integrin adhesion receptors can signal in two directions: first, they can regulate cellular behaviors by modulating cellular signaling enzymes ("outside-in signaling"); second, cells can regulate the affinity of integrins ("inside-out signaling") by such pathways. Integrin beta cytoplasmic domains (tails) mediate both types of signaling, and Src family kinases (SFKs) and talin, which bind to beta tails, are important for integrin signaling. Here, we utilized "homology scanning" mutagenesis to identify beta tail mutants selectively defective in c-Src binding and found that amino acid exchanges affecting a combination of an Arg and Thr residue in the integrin beta3 tail control the binding specificity for SFKs but have no effect on talin binding. Using beta tail mutants at these residues, we found that SFK binding to integrin beta tails is dispensable for inside-out signaling but is obligatory for cell spreading, a marker of outside-in signaling. Conversely, we found that point mutations that disrupt talin binding abolish integrin activation, but they do not inhibit SFK binding to the beta3 tail or the initiation of outside-in signaling once the integrins are in a high affinity form. Thus, we show that inside-out and outside-in integrin signaling are mediated by distinct and separable interactions of the integrin beta tails. Furthermore, based on our results, it is possible to discern the relative contributions of the direction of integrin signaling on biological functions in cell culture and, ultimately, in vivo.     

The collagen receptor integrins have distinct ligand recognition and signaling functions.

Distinct collagen subtypes are recognized by specific cell surface receptors. Two of the best known collagen receptors are members of the integrin family and are named alpha1beta1 and alpha2beta1. Integrin alpha1beta1 is abundant on smooth muscle cells, whereas the alpha2beta1 integrin is the major collagen receptor on epithelial cells and platelets. Many cell types, such as fibroblasts, osteoblasts, chondrocytes, endothelial cells, and lymphocytes may concomitantly express both of the receptors. We have studied the cell biology of these integrins at two levels. First, we have analyzed their ligand binding mechanism and specificity. Second, we have studied their signaling function inside three-dimensional collagen gels. This mini-review summarizes our most recent results. In conclusion, our data indicate that alpha1beta1 and alpha2beta1 integrins have differences in their ligand binding specificity. Furthermore, the two receptors are connected to distinct signaling pathways and their ligation may lead to opposite cellular responses.     

Beta 1 integrins signal lipid second messengers required during cell adhesion.

Clustering of integrin receptors during cell adhesion stimulates signal transduction across the cell membrane. Second messengers are generated, activating cytosolic proteins and causing cytoskeletal assembly and rearrangement. HeLa cell adhesion to a collagen substrate has been shown to initiate an arachidonic acid-mediated signaling pathway, leading to the activation of protein kinase C (PKC) and cell spreading. To determine the role of integrin receptors in triggering this signaling pathway, monoclonal antibodies to beta 1 integrins were used to either cluster integrins on the cell surface or to provide an integrin-dependent substrate for cell adhesion. Using this approach, we have defined a pathway required for cell spreading that can be initiated by the ligation of integrins and leads to the activation of PKC. Specifically, our results indicate that clustering beta 1 integrins results in the activation of phospholipase A2 leading to the production of arachidonic acid and the activation of PKC.     

Expression of chicken integrin beta 1 subunit in rat PC12 cells

We transfected rat pheochromocytoma (PC12) cells with a cDNA encoding chicken integrin beta 1 subunit. The chicken integrin beta 1 subunit produced in stable transfectants associated with two major alpha subunits of rat integrins to form interspecific chimeric receptors. These receptors mediated cell spreading and initial neurite outgrowth on laminin as did corresponding endogenous integrins, although they were slightly less effective in inducing cell adhesion to laminin. These results indicate that chicken integrin beta 1 may functionally substitute for beta 1 subunit of rat integrins in PC12 cells. Apparently, the structure of the integrin beta 1 subunit is highly conserved in the evolution of these species.     

beta1 integrins regulate keratinocyte adhesion and differentiation by distinct mechanisms

In keratinocytes, the beta1 integrins mediate adhesion to the extracellular matrix and also regulate the initiation of terminal differentiation. To explore the relationship between these functions, we stably infected primary human epidermal keratinocytes and an undifferentiated squamous cell carcinoma line, SCC4, with retroviruses encoding wild-type and mutant chick beta1 integrin subunits. We examined the ability of adhesion-blocking chick beta1-specific antibodies to inhibit suspension-induced terminal differentiation of primary human keratinocytes and the ability of the chick beta1 subunit to promote spontaneous differentiation of SCC4. A D154A point mutant clustered in focal adhesions but was inactive in the differentiation assays, showing that differentiation regulation required a functional ligand-binding domain. The signal transduced by beta1 integrins in normal keratinocytes was "do not differentiate" (transduced by ligand-occupied receptors) as opposed to "do differentiate" (transduced by unoccupied receptors), and the signal depended on the absolute number, rather than on the proportion, of occupied receptors. Single and double point mutations in cyto-2 and -3, the NPXY motifs, prevented focal adhesion targeting without inhibiting differentiation control. However, deletions in the proximal part of the cytoplasmic domain, affecting cyto-1, abolished the differentiation-regulatory ability of the beta1 subunit. We conclude that distinct signaling pathways are involved in beta1 integrin-mediated adhesion and differentiation control in keratinocytes.     

Regulation of the pp72syk protein tyrosine kinase by platelet integrin alpha IIb beta 3.

pp72syk is essential for development and function of several hematopoietic cells, and it becomes activated through tandem SH2 interaction with ITAM motifs in immune response receptors. Since Syk is also activated through integrins, which do not contain ITAMs, a CHO cell model system was used to study Syk activation by the platelet integrin, alpha IIb beta 3. As in platelets, Syk underwent tyrosine phosphorylation and activation during CHO cell adhesion to alpha IIb beta 3 ligands, including fibrinogen. This involved Syk autophosphorylation and the tyrosine kinase activity of Src, and it exhibited two novel features. Firstly, unlike alpha IIb beta 3-mediated activation of pp125FAK, Syk activation could be triggered by the binding of soluble fibrinogen and abolished by truncation of the alpha IIb or beta 3 cytoplasmic tail, and it was resistant to inhibition by cytochalasin D. Secondly, it did not require phosphorylated ITAMs since it was unaffected by disruption of an ITAM-interaction motif in the SH2(C) domain of Syk or by simultaneous overexpression of the tandem SH2 domains. These studies demonstrate that Syk is a proximal component in alpha IIb beta 3 signaling and is regulated as a consequence of intimate functional relationships with the alpha IIb beta 3 cytoplasmic tails and with Src or a closely related kinase. Furthermore, there are fundamental differences in the activation of Syk by alpha IIb beta 3 and immune response receptors, suggesting a unique role for integrins in Syk function.     

The N-terminal domains of talin cooperate with the phosphotyrosine binding-like domain to activate beta1 and beta3 integrins

The activation of integrin adhesion receptors from low to high affinity in response to intracellular cues controls cell adhesion and signaling. Binding of the cytoskeletal protein talin to the beta3 integrin cytoplasmic tail is required for beta3 activation, and the integrin-binding PTB-like F3 domain of talin is sufficient to activate beta3 integrins. Here we report that, whereas the conserved talin-integrin interaction is also required for beta1 activation, and talin F3 binds beta1 and beta3 integrins with comparable affinity, expression of the talin F3 domain is not sufficient to activate beta1 integrins. beta1 integrin activation could, however, be detected following expression of larger talin fragments that included the N-terminal and F1 domains, and mutagenesis indicates that these domains cooperate with talin F3 to mediate beta1 activation. This effect is not due to increased affinity for the integrin beta tail and we hypothesize that the N-terminal domains function by targeting or orienting talin in such a way as to optimize the interaction with the integrin tail. Analysis of beta3 integrin activation indicates that inclusion of the N-terminal and F1 domains also enhances F3-mediated beta3 activation. Our results therefore reveal a role for the N-terminal and F1 domains of talin during integrin activation and highlight differences in talin-mediated activation of beta1 and beta3 integrins.     

Polymerization of type I and III collagens is dependent on fibronectin and enhanced by integrins alpha 11beta 1 and alpha 2beta 1

Polymerization of the ECM proteins fibronectin and laminin has been shown to take place in close vicinity to the cell surface and be facilitated by beta(1) integrins (Lohikangas, L., Gullberg, D., and Johansson, S. (2001) Exp. Cell Res. 265, 135-144 and Wennerberg, K., Lohikangas, L., Gullberg, D., Pfaff, M., Johansson, S., and Fassler, R. (1996) J. Cell Biol. 132, 227-238). We have studied the role of collagen receptors, integrins alpha(11)beta(1) and alpha(2)beta(1), and fibronectin in collagen polymerization using fibronectin-deficient mouse embryonic fibroblast cell lines. In contrast to the earlier belief that collagen polymerization occurs via self-assembly of collagen molecules we show that a preformed fibronectin matrix is essential for collagen network formation and that collagen-binding integrins strongly enhance this process. Thus, collagen deposition is regulated by the cells, both indirectly through integrin alpha(5)beta(1)-dependent polymerization of fibronectin and directly through collagen-binding integrins.     

Tyrosine phosphorylation of the integrin beta 3 subunit regulates beta 3 cleavage by calpain

Outside-in signaling of beta(3) integrins induces and requires phosphorylation at tyrosine 747 (Tyr(747)) and tyrosine 759 (Tyr(759)) of the beta(3) subunit, but the mechanism for this requirement is unclear. On the other hand, a key consequence of integrin signaling, cell spreading, is inhibited by calpain cleavage of beta(3) cytoplasmic domain. Here we show that beta(3) tyrosine phosphorylation inhibits calpain cleavage. Mutating both tyrosines to phenylalanine sensitizes beta(3) to calpain cleavage. Furthermore, phosphorylation at Tyr(747) and Tyr(759) of beta(3) in the focal adhesion sites and the leading edge of spreading platelets was differentially regulated. Selective dephosphorylation of Tyr(759) is associated with calpain cleavage at Tyr(759). Thus, one mechanism by which tyrosine phosphorylation promotes integrin signaling and cell spreading is its inhibition of calpain cleavage of the beta(3) cytoplasmic domain.     

Expression of dominant-negative and chimeric subunits reveals an essential role for beta1 integrin during myelination

Myelination represents a remarkable example of cell specialization and cell-cell interaction in development. During this process, axons are wrapped by concentric layers of cell membrane derived either from central nervous system (CNS) oligodendrocytes or peripheral nervous system Schwann cells. In the CNS, oligodendrocytes elaborate a membranous extension with an area of more than 1000 times that of the cell body. The mechanisms regulating this change in cell shape remain poorly understood. Signaling mechanisms regulated by cell surface adhesion receptors of the integrin family represent likely candidates. Integrins link the extracellular environment of the cell with both intracellular signaling molecules and the cytoskeleton and have been shown to regulate the activity of GTPases implicated in the control of cell shape. Our previous work has established that oligodendrocytes and their precursors express a limited repertoire of integrins. One of these, the alpha6beta1 laminin receptor, can interact with laminin-2 substrates to enhance oligodendrocyte myelin membrane formation in cell culture. However, these experiments do not address the important question of integrin function during myelination in vivo, nor do they define the respective roles of the alpha and beta subunits in the signaling pathways involved. Here, we use a dominant-negative approach to provide, for the first time, evidence that beta1 integrin function is required for myelination in vivo and use chimeric integrins to dissect apart the roles of the extracellular and cytoplasmic domains of the alpha6 subunit in the signaling pathways of myelination.     

Cytoplasmic tails of beta 1, beta 2, and beta 7 integrins differentially regulate LFA-1 function in K562 cells.

The beta 2 integrin lymphocyte function-associated antigen 1 (LFA-1) mediates activation-dependent adhesion of lymphocytes. To investigate whether lymphocyte-specific elements are essential for LFA-1 function, we expressed LFA-1 in the erythroleukemic cell line K562, which expresses only the integrin very late antigen 5. We observed that LFA-1-expressing K562 cannot bind to intercellular adhesion molecule 1-coated surfaces when stimulated by phorbol 12-myristate 13-acetate (PMA), whereas the LFA-1-activating antibody KIM185 markedly enhanced adhesion. Because the endogenously expressed beta 1 integrin very late antigen 5 is readily activated by PMA, we investigated the role of the cytoplasmic domain of distinct beta subunits in regulating LFA-1 function. Transfection of chimeric LFA-1 receptors in K562 cells reveals that replacement of the beta 2 cytoplasmic tail with the beta 1 but not the beta 7 cytoplasmic tail completely restores PMA responsiveness of LFA-1, whereas a beta 2 cytoplasmic deletion mutant of LFA-1 is constitutively active. Both deletion of the beta 2 cytoplasmic tail or replacement by the beta 1 cytoplasmic tail alters the localization of LFA-1 into clusters, thereby regulating LFA-1 activation and LFA-1-mediated adhesion to intercellular adhesion molecule 1. These data demonstrate that distinct signaling routes activate beta 1 and beta 2 integrins through the beta-chain and hint at the involvement of lymphocyte-specific signal transduction elements in beta 2 and beta 7 integrin activation that are absent in the nonlymphocytic cell line K562.     

Phospholipase Cgamma binds alpha1beta1 integrin and modulates alpha1beta1 integrin-specific adhesion.

Integrin adhesion receptors have been implicated in bidirectional signal transduction. The dynamic regulation of integrin affinity and avidity as well as post-ligand effects involved in outside-in signaling depends on the interaction of integrins with cytoskeletal and signaling proteins. In this study, we attempted to identify cytoplasmic binding partners of alpha(1)beta(1) integrin. We were able to show that cell adhesion to alpha(1)beta(1)-specific substrates results in the association of phospholipase Cgamma (PLCgamma) with the alpha(1)beta(1) integrin independent of PLCgamma tyrosine phosphorylation. Using peptide-binding assays, the membrane proximal sequences within the alpha(1)beta(1) integrin subunits were identified as binding sites for PLCgamma. In particular, the conserved sequence of beta(1) subunit binds the enzyme very efficiently. Because purified PLCgamma also binds the integrin peptides, binding seems to be direct. Inhibition of PLC by leads to reduced cell adhesion on alpha(1)beta(1)-specific substrates. Cells lacking the conserved domain of the alpha(1) subunit fail to respond to the PLC inhibition, indicating that this domain is necessary for PLC-dependent adhesion modulation of alpha(1)beta(1) integrin.     

Threonine phosphorylation sites in the beta 2 and beta 7 leukocyte integrin polypeptides

The cytoplasmic domains of integrins play a key role in a variety of integrin-mediated events including adhesion, migration, and signaling. The molecular mechanisms that enhance integrin function are still incompletely understood. Because protein kinases are known to be involved in the signaling and the activation of integrins, the role of phosphorylation has been studied by several groups. The beta(2) leukocyte integrin subunit has previously been shown to become phosphorylated in leukocytes on cytoplasmic serine and functionally important threonine residues. We have now mapped the phosphorylated threonine residues in activated T cells. After phorbol ester stimulation, all three threonine residues (758-760) of the threonine triplet became phosphorylated but only two at a time. CD3 stimulation leads to a strong threonine phosphorylation of the beta(2) integrin, but differed from phorbol ester activation in that phosphorylation occurred only on threonine 758. The other leukocyte-specific integrin, beta(7), has also been shown to need the cytoplasmic domain and leukocyte-specific signal transduction elements for integrin activation. Cell activation with phorbol ester, and interestingly, through the TCR-CD3 complex, caused beta(7) integrin binding to VCAM-1. Additionally, cell activation led to increased phosphorylation of the beta(7) subunit, and phosphoamino acid analysis revealed that threonine residues became phosphorylated after cell activation. Sequence analysis by manual radiosequencing by Edman degradation established that threonine phosphorylation occurred in the same threonine triplet as in beta(2) phosphorylation.     

Human ether-a-go-go-related gene 1 channels are physically linked to beta1 integrins and modulate adhesion-dependent signaling

Adhesive receptors of the integrin family are primarily involved in cell-extracellular matrix adhesion. Additionally, integrins trigger multiple signaling pathways that are involved in cell migration, proliferation, survival, and differentiation. We previously demonstrated that the activation of integrins containing the beta(1) subunit leads to a selective increase in potassium currents carried by the human ether-a-go-go-related gene (hERG) channels in neuroblastoma and leukemia cells; this current activation modulates adhesion-dependent differentiation in these cells. We hypothesized that the cross-talk between integrins and hERG channels could be traced back to the assembly of a macromolecular signaling complex comprising the two proteins. We tested this hypothesis in both SH-SY5Y neuroblastoma cells and in human embryonic kidney 293 cells stably transfected with hERG1 and, therefore, expressing only the full-length hERG1 protein on the plasma membrane. The beta(1) integrin and hERG1 coprecipitate in these cells and colocalize in both intracellular and surface membrane compartments. The two proteins also coprecipitate with caveolin-1, suggesting the localization of the complex in lipid rafts/caveolae. hERG1-transfected cells undergo an activation of hERG currents after beta(1) integrin-mediated adhesion to fibronectin; concomitant with this activation, the focal adhesion kinase associates with the hERG1 protein and becomes tyrosine phosphorylated. Using hERG1-specific inhibitors, we show that the tyrosine phosphorylation of focal adhesion kinase is strictly dependent on hERG channel activity. Similarly, the activity of the small GTPase Rac1 turned out to be dependent on hERG currents. On the whole, these data indicate that the hERG1 protein associates with beta(1) integrins and modulates adhesion receptor signaling.