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PubMed1997

Activation of beta1 integrins on CML progenitors reveals cooperation between beta1 integrins and CD44 in the regulation of adhesion and proliferation.

Adhesion of normal colony-forming cells (CFC) to bone marrow (BM) stroma and the extracellular matrix (ECM) component fibronectin (FN) depends at least in part on the alpha4beta1 and alpha5beta1 integrins and the CD44 receptor. Aside from anchoring progenitors in the marrow microenvironment, beta1 integrin-dependent adhesion of normal CFC is associated with inhibition of their proliferation. In contrast to normal CFC, chronic myelogenous leukemia (CML) Ph+ CFC adhere significantly less to either stroma or FN. CML Ph+ CFC proliferation is also not inhibited by coculture with stroma or FN. However, equal numbers of alpha4, alpha5, and beta1 integrins and CD44 are present on CML and normal CD34+ cells. We have previously demonstrated that beta1-dependent adhesion to and subsequent proliferation inhibition by FN can be restored when CML Ph+ CFC are incubated with the beta1 integrin activating antibody, 8A2, and demonstrated a role for the alpha5beta1 integrin in this phenomenon. Since the integrin alpha4beta1 and the proteoglycan form of CD44 may cooperate in establishing normal CFC adhesion to FN, we examined if treatment of CML Ph+ CFC with 8A2 also restores the cooperativity between beta1 integrins and CD44. We demonstrate that 8A2 induces adhesion of CML Ph+ CFC not only to intact FN but also to alpha4beta1, alpha5beta1, and proteoglycan binding fragments of FN. 8A2-induced adhesion to these fragments and peptides also results in a significant inhibition of the proliferation of CML Ph+ CFC. Addition of antibodies to either the alpha5, alpha4, or beta1 integrins, antibodies against the CD44 receptor, or removal of chondroitin sulfate glycosaminoglycans from the surface of CML CD34+ HLA-DR+ cells significantly reduced the 8A2-induced adhesion to and adhesion-mediated inhibition of proliferation by FN. These studies demonstrate that activation of beta1 integrins on CML Ph+ CFC not only results in upregulation of beta1 integrin-dependent adhesion and adhesion-mediated inhibition of proliferation, but also in the restoration of cooperation between beta1 integrins and CD44. These studies suggest that decreased beta1 integrin avidity may also affect the function of the proteoglycan adhesion receptor CD44, both of which may contribute to the abnormal circulation and expansion of malignant progenitors in CML.

Antigens, CD↗

PubMed2003

Essential role for integrin linked kinase in Akt-mediated integrin survival signaling in hippocampal neurons.

Activation of integrin receptors in neurons can promote cell survival and synaptic plasticity, but the underlying signal transduction pathway(s) is unknown. We report that integrin signaling prevents apoptosis of embryonic hippocampal neurons by a mechanism involving integrin-linked kinase (ILK) that activates Akt kinase. Activation of integrins using a peptide containing the amino acid sequence EIKLLIS derived from the alpha chain of laminin protected hippocampal neurons from apoptosis induced by glutamate or staurosporine, and increased Akt activity in a beta1 integrin-dependent manner. Transfection of neurons with a plasmid encoding dominant negative Akt blocked the protective effect of the integrin-activating peptide, as did a chemical inhibitor of Akt. Although inhibitors of phosphoinositide-3 (PI3) kinase blocked the protective effect of the peptide, we found no increase in PI3 kinase activity following integrin stimulation suggesting that PI3 kinase was necessary for Akt activity but was not sufficient for the increase in Akt activity following integrin activation. Instead, we show a requirement for ILK in integrin receptor-induced Akt activation. ILK was activated following integrin stimulation and dominant negative ILK blocked integrin-mediated Akt activation and cell survival. Activation of ILK and Akt were also required for neuroprotection by substrate-associated laminin. These results establish a novel pathway that signals cell survival in neurons in response to integrin receptor activation.

Animals↗

PubMed1997

Ligation of integrin alpha5beta1 is required for internalization of vitronectin by integrin alphavbeta3.

Remodeling of the matrix by tumor cells is necessary for tumor invasion. We have shown previously that malignant astrocytomas, in contrast to normal astrocytes, synthesize vitronectin and express integrins alphavbeta3 and alphavbeta5. The activity states of these two integrins are differentially controlled. Thus, we investigated the regulation of the activity of integrins alphavbeta3 and alphavbeta5 with regard to their role in vitronectin internalization in U-251MG astrocytoma cell monolayers adherent to fibronectin, collagen, or laminin in serum-free conditions. Binding of [125I]vitronectin occurred in a specific, saturable manner that was partially inhibitable by monoclonal antibodies (mAbs) specific for integrins alphavbeta3 or alphavbeta5. Specific, lysosomally-mediated degradation of [125I]vitronectin was detectable at 1 h and increased over the 24-h assay period. The cell substrate affected the rate of turnover of [125I]vitronectin, which was 3.0 ng/min for cells plated on fibronectin but 0.35 ng/min for cells plated on collagen. Furthermore, although mAbs specific for either integrin alphavbeta3 or alphavbeta5 inhibited degradation (30%; combined effect 70%) of [125I]vitronectin by cells plated on fibronectin, only mAb anti-alphavbeta5 inhibited degradation (70-90%) by cells plated on collagen or laminin. To determine the requirement for integrin alpha5beta1 ligation in order for integrin alphavbeta3 to internalize its ligand, cells were plated on mAbs anti-integrin alpha5 or anti-integrin alpha3. When plated on mAb anti-alpha5, mAbs anti-alphavbeta3 and anti-alphavbeta5 both inhibited degradation. However, when plated on mAb anti-alpha3, mAb anti-alphavbeta3 had no effect whereas mAb anti-alphavbeta5 inhibited degradation. These data indicate that a signal from integrin alpha5beta1 is necessary for integrin alphavbeta3 to internalize vitronectin, whereas integrin alphavbeta5 constitutively internalizes vitronectin.

Astrocytoma↗

PubMed2001

Beta1 integrin activation on human neutrophils promotes beta2 integrin-mediated adhesion to fibronectin.

Although the importance of beta1 integrin-mediated binding to adhesion molecules and extracellular matrix (ECM) molecules is well established for most types of leukocytes, the expression patterns and functional importance of beta1 integrins on neutrophils have remained controversial. Using flow cytometry, we found that human neutrophils express the alpha4, alpha5, alpha9 and beta1 integrin subunits. To examine whether the integrins VLA-4 (alpha4/beta1) and VLA-5 (alpha5/beta1) have a functional role on neutrophils, we studied adhesion to their ligand fibronectin. Treatment of neutrophils with antibody 8A2, which specifically binds and activates beta1 integrins, resulted in increased binding to fibronectin. However, addition of blocking mAb revealed that 8A2-induced adhesion did not depend on beta1 integrins, but on the beta2 integrin CD11b/CD18. Similarly, activation of beta1 integrins by 8A2 resulted in CD11b-dependent binding of neutrophils to fibrinogen. 8A2 treatment increased expression of an activation epitope of CD11b/CD18, which depended on phosphoinositide 3-OH kinase activity and an adequate concentration of intracellular free Ca2+. These data suggest that engagement of beta1 integrins on neutrophils results in a cross-talk signal that leads to activation of the beta2 integrin CD11b/CD18, followed by CD11b-mediated adhesion. As transmigrated neutrophils are surrounded by both beta1 and beta2 ligands in the ECM, this integrin cross-talk could play a role in modifying migration and cellular activation in inflamed tissues.

Androstadienes↗

PubMed1989

A novel integrin (alpha E beta 4) from human epithelial cells suggests a fourth family of integrin adhesion receptors.

A new member of the integrin superfamily of adhesion receptors was isolated from human epithelial cells. Analogously to other integrins, this molecule is a heterodimer comprised of structurally unrelated subunits, both glycosylated. Unequivocal amino-acid sequence homologies were observed between these subunits and integrin alpha and beta chain sequences, indicating that this epithelial heterodimer is a novel integrin. No obvious serologic cross-reactivities were detected with other integrins. The beta chain of the epithelial integrin displayed a mol. wt significantly higher than other integrin beta chains, possibly due to a large sialic acid content. Integrin heterodimers are grouped into three families, based on which of three beta chains (beta 1, beta 2 and beta 3) they contain. Therefore, the epithelial integrin may represent the prototype of a fourth integrin family, because it contains a structurally distinct beta chain. The designation alpha E beta 4 is proposed for this novel human integrin.

Amino Acid Sequence↗

PubMedDate not supplied

Adenovirus type 5 uptake by lung adenocarcinoma cells in culture correlates with Ad5 fibre binding is mediated by alpha(v)beta1 integrin and can be modulated by changes in beta1 integrin function.

BACKGROUND: Recombinant adenoviruses (Ad) have been employed as vectors for a wide variety of gene therapy applications, but their use has been hindered by problems relating to efficacy and safety. The efficiency of Ad-mediated gene transfer depends on the interaction of the fibre and penton base proteins with their corresponding cell receptors. Ad infection is initiated by the formation of a high affinity complex between the fibre protein and a host cell protein that for most Ad serotypes is CAR (the coxsackie B virus and Ad receptor). A second molecule, the MHC class I, may also be involved in Ad type 2 and Ad type 5 uptake. Ad internalization results from the interaction of the penton base protein with cell surface integrins alpha(v)beta3 and alpha(v)beta5. In this study, we addressed the interaction between Ad type 5 (Ad5) and its receptors on lung derived adenocarcinoma cells in culture. METHODS: Using flow cytometry, we determined the level of expression of attachment and internalization receptors that are expressed on the cell surface of A549, H322 and H441 lung-derived adenocarcinoma cells in culture. The level of alpha(v)beta1 cell surface integrin was assessed by immunoprecipitation. Measuring the level of luciferase gene expression at different viral titres quantitated Ad5 uptake by these cells. The kinetics of binding of Ad5 fibre knobs to A549, H322 and H441 cells was assessed in direct binding studies using 125I labelling of purified recombinant Ad5 fibre-knob domains. In order to assess the functionality of integrins, adhesion assays were performed in the presence or absence of activators of integrin function. In competition experiments, prior to exposure to the virus, the cells were pre-incubated with purified recombinant Ad5 fibre-knob domains, function blocking anti-integrin antibodies, or integrin activating agents, prior to the introduction of luciferase expressing Ad5. RESULTS: We found that Ad5-mediated gene transfer in A549, H322 and H441 adenocarcinoma cells in culture is highly variable and that this variation correlates with specific binding of Ad5 fibre-knob domain binding to the cell surface. We also found, for the first time, that Ad5 infection is mediated by integrin alpha(v)beta1 and that functional activation of beta1 integrin by means of the specific anti-beta1 monoclonal antibody, TS2/16, induced increased A549 cell adhesion to fibronectin and vitronectin and also enhanced Ad5 uptake by these cells. CONCLUSIONS: Our studies demonstrate that the Ad5 fibre-knob domain interaction with CAR represents a major determinant of Ad5-mediated gene transfer to lung-derived adenocarcinoma cells in culture. The finding that integrin alpha(v)beta1 is involved in Ad5 infection has implications for the use of recombinant Ad5 vectors for cancer gene therapy, since alpha(v)beta1 is expressed at high levels and acts as an alternative vitronectin receptor in many epithelial and some melanoma tumours which express no alpha(v)beta3 and constant low levels of alpha(v)beta5. The fact that the beta1 integrin-activating antibody TS2/16 can enhance alpha(v)beta1-mediated Ad5 infection suggests that the efficacy of Ad5-mediated gene transfer might be influenced not only by the level of cell surface expression of integrins but also by their state of activation.

Adenocarcinoma↗

PubMed2000

Integrin expression by primary and immortalized human chondrocytes: evidence of a differential role for alpha1beta1 and alpha2beta1 integrins in mediating chondrocyte adhesion to types II and VI collagen.

OBJECTIVE: Chondrocytes have been shown to express beta1-containing integrins both in vitro and in situ, but their role in regulating chondrocyte function is poorly understood. The objective of this study was to determine how the relative expression of different integrins may be modulated in relation to the differentiated state and proliferative capacity of the chondrocyte. DESIGN: Integrin expression by four different cell lines of human chondrocytes immortalized with Simian virus 40 large T-antigen (SV40-TAg) was studied and compared to primary chondrocytes. Differences in alpha1 and alpha2 integrin subunit expression were utilized to further study the role of these integrins in mediating adhesion to types II and VI collagen. RESULTS: The overall cell-surface levels of beta1-containing integrins were higher on all four immortalized cell lines which expressed over 10-fold higher levels of alpha2 and alpha3 integrin subunits compared to primary cells. However, primary cells expressed higher levels of the alpha1 integrin subunit which was not expressed by T/C28a4 cells and expressed at variable and lower levels in the other lines. Levels of the alpha3 integrin subunit were significantly greater on the highly proliferative juvenile costal chondrocyte lines (T/C-28a4, C-2812, and C-20a4) compared to primary articular chondrocytes and tsT/AC-62 cells which were derived from adult articular chondrocytes. Expression of alpha5 was similar among primary cells and cell lines except on C-20/A4 cells which had an average of over 4-fold higher levels. None of the primary or immortalized chondrocytes tested expressed significant levels of alpha4. Cell adhesion assays revealed that both alpha1beta1 and alpha2beta1 could serve as chondrocyte adhesion receptors for types II and VI collagen. In cell lines expressing both integrins, alpha1beta1 was the preferential receptor for type VI collagen while alpha2beta1 was the preferential receptor for type II collagen. Rather than inhibiting adhesion, incubation with the alpha3 blocking antibody P1B5 increased adhesion of C-28/12 cells to both fibronectin and type II collagen by 67% and 100% respectively. CONCLUSIONS: Immortalization with SV40-TAg results in altered integrin expression by chondrocytes. Changes in the relative levels of alpha1, alpha2, and alpha3 subunits may significantly alter the manner in which chondrocytes interact with types II and VI collagen in the extracellular matrix.

Adult↗

PubMed1999

Steroid regulation of human placental integrins: suppression of alpha2 integrin expression in cytotrophoblasts by glucocorticoids.

Maintenance of uterine-placental attachment during human pregnancy may depend at least partly on adhesive interactions between cytotrophoblasts and their extracellular matrix (ECM). Such interactions are often mediated by integrins, signal-transducing heterodimeric transmembrane glycoproteins. We previously showed that glucocorticoid (GC) suppressed the expression of collagen and laminin in human placenta; here we show that GC also modulates the expression by human cytotrophoblasts of the integrin subunits alpha2 and beta1, components of a known receptor for these ECM ligands. Cytotrophoblasts were isolated from human term placentas, cultured up to 4 days in the presence of 0-1000 nM dexamethasone (DEX), and assayed for 1) integrin messenger RNA (mRNA) levels by Northern hybridization, 2) integrin subunit synthesis after [35S]methionine labeling, or 3) cell surface integrin levels after 125I labeling by lactoperoxidase. In four independent experiments, 100 nM DEX reduced mRNA levels for integrin alpha2 to 6+/-1% of the control value. This effect was similar between 1-4 days of treatment and was dose dependent between 1-1000 nM DEX. Cortisol treatment (100 nM) inhibited levels of integrin alpha2 mRNA, but 100 nM testosterone, estradiol, and progesterone were less effective, suggesting that this response was specific to GC. In immunoprecipitation studies, treatment of cytotrophoblasts with 100 nM DEX for 2 days reduced the rates of synthesis of the alpha2 integrin subunit as well as its expression on the cell surface to 1-10% of control levels. DEX effects on the beta1 integrin subunit were less dramatic. DEX reduced beta1 mRNA levels to only 69+/-8% of control levels, a smaller reduction compared with effects on alpha2 integrin mRNA. DEX inhibited beta1 protein synthesis and cell surface expression to 60-70% of control levels. In all experiments, DEX had no effect on total protein synthesis. Thus, our results demonstrate that GC treatment specifically and markedly down-regulates expression of alpha2 integrin subunit by human cytotrophoblasts. This finding is consistent with the concept that uterine-placental adherence across gestation may be regulated by coordinate effects on ECM ligands and cellular adhesion receptors.

Antigens, CD↗

PubMed1998

Hepatocyte growth factor enables enhanced integrin-cytoskeleton linkage by affecting integrin expression in subconfluent epithelial cells.

Hepatocyte growth factor (HGF) exerts mitogenic and motogenic effects in different cell types. In the epithelial cell line mHepR1 we found that HGF induced pronounced alterations in cell morphology and promoted cell adhesion and spreading. To analyze the mechanisms how HGF affects these integrin mediated functions we studied the physical linkage of integrins with the cytoskeleton. First we found that HGF increased the expression of different integrin subunits in subconfluent cells and influenced the distribution of integrins on the cell surface. To address the physical association of integrins with the cytoskeleton we analyzed Triton X-100-extracted cell fractions using flow cytometry. Here we show that cultivation of the cells with HGF for 24 h prior to integrin cross-linking significantly enhanced the cytoskeletal anchorage of integrins. To further find out whether HGF directly induces an integrin-cytoskeleton link without subsequent cross-linking we added HGF to suspended cells but failed to detect cytoskeletally immobilized integrins in the detergent-insoluble cell fraction which could be related to the absence of a calcium response induced by HGF. Overall, the results indicate that HGF promotes the physical linkage of integrins to the cytoskeleton which requires additional stimulation of integrins.

Animals↗

PubMed2002

Modulation of integrins and integrin signaling molecules in the pressure-loaded murine ventricle.

Integrins are heterodimeric cell-surface receptors that link the extracellular matrix and the intracellular cytoskeleton and function as mechanotransducers. Signaling through integrins is important for cell growth, migration, and survival. Extracellular matrix is altered in the myocardium during hypertrophic induction and the transition to heart failure. The role of integrins in this process is poorly understood. Recently, integrin subunits have been identified that are dominantly expressed in striated muscle. We tested the hypothesis that since integrins are mechanotransducers, their expression and signaling would be modulated with murine left ventricular hemodynamic loading. The acute and chronic effects of pressure overload on changes in the expression of integrins, as well as related integrin-mediated signaling events were studied. Acute pressure loading increased phosphorylation of focal adhesion kinase, p42 and p44 extracellular signal-regulated kinase. Chronic loading: (1) increased expression of alpha1, alpha5, and beta1 integrin transcripts and (2) increased protein expression of integrin subunits which are dominantly expressed in striated muscle (alpha7 and beta1D) both by western blotting and immunofluorescent microscopy. These results show that adaptive responses of the myocardium to pressure overload include acute modulation of integrin-related signaling molecules and more chronic changes effect expression of integrin subunits, including ones dominantly expressed in muscle.

Animals↗

PubMed2001

The linkage between beta1 integrin and the actin cytoskeleton is differentially regulated by tyrosine and serine/threonine phosphorylation of beta1 integrin in normal and cancerous human breast cells.

BACKGROUND: Structural requirements for the beta1 integrin functions in cell adhesion, spreading and signaling have been well documented mainly for fibroblasts. In this study, we examined the reason for the reduced surface expression of beta1 integrin in human breast cancer MCF-7 cells compared to normal human breast epithelial (HBE) cells, both of which adhered to collagen type IV. RESULTS: The beta1 integrin immunoprecipitates from either HBE or MCF-7 cells involved alpha-actinin while actin coprecipitated with beta1 integrin from HBE cells but not from MCF-7 cells. Immunoblotting using the anti-phosphotyrosine (PY) antibody indicated the phosphorylation of beta1 integrin at least at tyrosine in both cells. Dephosphorylation of beta1 integrin from HBE cells by protein tyrosine phosphatase (PTP), but not by protein serine/threonine phosphatase (PP), caused dissociation of actin from beta1 integrin, although dephosphorylation of it from MCF-7 cells by either PTP or PP caused association of the two proteins. In MCF-7 cells beta1 integrin coprecipitated doublet of proteins having the Ca2+/calmodulin-dependent protein kinase (CaMK) II activity that was susceptible to KN-62, a specific inhibitor of CaMKII. CONCLUSION: The results suggest that beta1 integrin is tyrosine phosphorylated and links with actin via alpha-actinin in HBE cells but prevented from linking with actin in MCF-7 cells by phosphorylation at both tyrosine and serine/threonine of beta1 integrin which forms a complex with alpha-actinin and CaMKII. Thus the linkage formation of beta1 integrin with actin may be differentially regulated by its tyrosine and serine/threonine phosphorylation in normal HBE cells and breast cancer MCF-7 cells.

Actin Cytoskeleton↗

PubMed2004

A specific alpha5beta1-integrin conformation promotes directional integrin translocation and fibronectin matrix formation.

Integrin adhesion receptors are structurally dynamic proteins that adopt a number of functionally relevant conformations. We have produced a conformation-dependent anti-alpha5 monoclonal antibody (SNAKA51) that converts alpha5beta1 integrin into a ligand-competent form and promotes fibronectin binding. In adherent fibroblasts, SNAKA51 preferentially bound to integrins in fibrillar adhesions. Clustering of integrins expressing this activation epitope induced directional translocation of alpha5beta1, mimicking fibrillar adhesion formation. Priming of alpha5beta1 integrin by SNAKA51 increased the accumulation of detergent-resistant fibronectin in the extracellular matrix, thus identifying an integrin conformation that promotes matrix assembly. The SNAKA51 epitope was mapped to the calf-1/calf-2 domains. We propose that the action of the antibody causes the legs of the integrin to change conformation and thereby primes the integrin to bind ligand. These findings identify SNAKA51 as the first anti-integrin antibody to selectively recognize a subset of adhesion contacts, and they identify an integrin conformation associated with integrin translocation and fibronectin matrix formation.

Antibodies, Monoclonal↗

PubMed2005

Disintegrins: integrin selective ligands which activate integrin-coupled signaling and modulate leukocyte functions.

Extracellular matrix proteins and cell adhesion receptors (integrins) play essential roles in the regulation of cell adhesion and migration. Interactions of integrins with the extracellular matrix proteins lead to phosphorylation of several intracellular proteins such as focal adhesion kinase, activating different signaling pathways responsible for the regulation of a variety of cell functions, including cytoskeleton mobilization. Once leukocytes are guided to sites of infection, inflammation, or antigen presentation, integrins can participate in the initiation, maintenance, or termination of the immune and inflammatory responses. The modulation of neutrophil activation through integrin-mediated pathways is important in the homeostatic control of the resolution of inflammatory states. In addition, during recirculation, T lymphocyte movement through distinct microenvironments is mediated by integrins, which are critical for cell cycle, differentiation and gene expression. Disintegrins are a family of low-molecular weight, cysteine-rich peptides first identified in snake venom, usually containing an RGD (Arg-Gly-Asp) motif, which confers the ability to selectively bind to integrins, inhibiting integrin-related functions in different cell systems. In this review we show that, depending on the cell type and the microenvironment, disintegrins are able to antagonize the effects of integrins or to act agonistically by activating integrin-mediated signaling. Disintegrins have proven useful as tools to improve the understanding of the molecular events regulated by integrin signaling in leukocytes and prototypes in order to design therapies able to interfere with integrin-mediated effects.

Cell Adhesion↗

PubMed2000

Engagement of beta2 integrins induces surface expression of beta1 integrin receptors in human neutrophils.

Induction of beta1 integrin (CD49/CD29) expression in polymorphonuclear leukocytes (PMN) has been shown to be associated with transendothelial migration recently. Yet, beta1 integrin expression is relatively insensitive to cell activation with soluble agonists, such as N-formyl-methionyl-leucyl-phenylalanine (fMLP). We hypothesized that beta2 integrins (CD11/CD18), critically involved in PMN adhesion and extravasation, may play a role in regulating 1 integrin expression in PMN. Antibody cross-linking of CD18, mimicking adhesion-dependent engagement of beta2 integrins, resulted in rapid, tyrosine kinase-dependent upregulation of beta1 integrins. This response was potentiated by simultaneous chemoattractant (fMLP) stimulation of PMN. Moreover, upregulation of beta1 integrins evoked by CD18 cross-linking was found to support adhesion of fMLP-stimulated PMN to matrix proteins and also was critical for the ability of PMN to migrate in collagen gels in response to a gradient of fMLP. Taken together, these data demonstrate that engagement of beta2 integrins in human PMN induces beta1 integrin expression in these cells of significance for their migration in the extravascular tissue. Thus, beta2 integrins may serve the function to regulate PMN locomotion in extravascular tissue via receptor crosstalk with beta1 integrins.

Biological Transport↗

PubMed2002

Enhanced pathological angiogenesis in mice lacking beta3 integrin or beta3 and beta5 integrins.

Inhibition of alphavbeta3 or alphavbeta5 integrin function has been reported to suppress neovascularization and tumor growth, suggesting that these integrins are critical modulators of angiogenesis. Here we report that mice lacking beta3 integrins or both beta3 and beta5 integrins not only support tumorigenesis, but have enhanced tumor growth as well. Moreover, the tumors in these integrin-deficient mice display enhanced angiogenesis, strongly suggesting that neither beta3 nor beta5 integrins are essential for neovascularization. We also observed that angiogenic responses to hypoxia and vascular endothelial growth factor (VEGF) are augmented significantly in the absence of beta3 integrins. We found no evidence that the expression or functions of other integrins were altered as a consequence of the beta3 deficiency, but we did observe elevated levels of VEGF receptor-2 (also called Flk-1) in beta3-null endothelial cells. These data indicate that alphavbeta3 and alphavbeta5 integrins are not essential for vascular development or pathological angiogenesis and highlight the need for further evaluation of the mechanisms of action of alphav-integrin antagonists in anti-angiogenic therapeutics.

Animals↗

PubMed1999

cDNA cloning and chromosomal localization of human alpha(11) integrin. A collagen-binding, I domain-containing, beta(1)-associated integrin alpha-chain present in muscle tissues.

We previously identified a novel integrin alpha-chain in human fetal muscle cells (Gullberg, D., Velling, T., Sjöberg, G., and Sejersen, T. (1995) Dev. Dyn. 204, 57-65). We have now isolated the full-length cDNA for this integrin subunit, alpha(11). The open reading frame of the cDNA encodes a precursor of 1188 amino acids. The predicted mature protein of 1166 amino acids contains seven conserved FG-GAP repeats, an I domain with a metal ion-dependent adhesion site motif, a short transmembrane region, and a unique cytoplasmic domain of 24 amino acids containing the sequence GFFRS. alpha(11), like other I domain integrins, lacks a dibasic cleavage site for generation of a heavy chain and a light chain, and it contains three potential divalent cation binding sites in repeats 5-7. The presence of 22 inserted amino acids in the extracellular stalk portion (amino acids 804-826) distinguishes the alpha(11) integrin sequence from other integrin alpha-chains. Amino acid sequence comparisons reveal the highest identity of 42% with the alpha(10) integrin chain. Immunoprecipitation with antibodies to alpha(11) integrin captures a 145-kDa protein distinctly larger than the 140-kDa alpha(2) integrin chain when analyzed by SDS-polyacrylamide gel electrophoresis under nonreducing conditions. Fluorescence in situ hybridization maps the integrin alpha(11) gene to chromosome 15q23, in the vicinity of an identified locus for Bardet-Biedl syndrome. Based on Northern blotting, integrin alpha(11) mRNA levels are high in the adult human uterus and in the heart and intermediate in skeletal muscle and some other tissues tested. During in vitro myogenic differentiation, alpha(11) mRNA and protein are up-regulated. Studies of ligand binding properties show that alpha(11)beta(1) binds collagen type I-Sepharose, and cultured muscle cells localize alpha(11)beta(1) into focal contacts on collagen type I. Future studies will reveal the importance of alpha(11)beta(1) for muscle development and integrity in adult muscle and other tissues.

Adult↗

PubMed2006

Contribution of alpha6 integrins to hematopoietic stem and progenitor cell homing to bone marrow and collaboration with alpha4 integrins.

The laminin receptor integrin alpha6 chain is ubiquitously expressed in human and mouse hematopoietic stem and progenitor cells. We have studied its role for homing of stem and progenitor cells to mouse hematopoietic tissues in vivo. A function-blocking anti-integrin alpha6 antibody significantly reduced progenitor cell homing to bone marrow (BM) of lethally irradiated mice, with a corresponding retention of progenitors in blood. Remarkably, the anti-integrin alpha6 antibody profoundly inhibited BM homing of long-term multilineage engrafting stem cells, studied by competitive repopulation assay and analysis of donor-derived lymphocytes and myeloid cells in blood 16 weeks after transplantation. A similar profound inhibition of long-term stem cell homing was obtained by using a function-blocking antibody against alpha4 integrin, studied in parallel. Furthermore, the anti-integrin alpha6 and alpha4 antibodies synergistically inhibited homing of short-term repopulating stem cells. Intravenous injection of anti-integrin alpha6 antibodies, in contrast to antibodies against alpha4 integrin, did not mobilize progenitors or enhance cytokine-induced mobilization by G-CSF. Our results provide the first evidence for a distinct functional role of integrin alpha6 receptor during hematopoietic stem and progenitor cell homing and collaboration of alpha6 integrin with alpha4 integrin receptors during homing of short-term stem cells.

Animals↗

PubMed1994

Disruption of integrin function and induction of tyrosine phosphorylation by the autonomously expressed beta 1 integrin cytoplasmic domain.

The cytoplasmic domains of integrin beta subunits are essential for the function of integrins in cell adhesion and signaling. A chimera combining the transmembrane and cytoplasmic domains of the beta 1 integrin subunit with an irrelevant extracellular domain derived from L3T4 (murine CD4) was tested for its ability to interfere with integrin function. Expression of this construct in cultured human embryonic kidney cells under the control of the inducible metallothionein promoter resulted in cell rounding and detachment, and blocked cell adhesion mediated by the beta 1 and alpha v beta 5 integrins. Expression of the beta 1 chimera at basal levels interfered with the tyrosine phosphorylation of a 125-kDa protein induced by antibody-induced clustering of integrins. Induced expression of the chimera resulted in sustained tyrosine phosphorylation of this protein, which could be enhanced by clustering of the chimera but was insensitive to clustering of integrins. These results demonstrate that the autonomously expressed beta 1 integrin cytoplasmic domain can act as a trans-dominant inhibitor of integrin function, presumably via competitive interactions with cytoplasmic components that are required for integrin-mediated cell adhesion and tyrosine phosphorylation.

Amino Acid Sequence↗