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Results for “Integrin alpha5beta1”

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PubMed2006

Integrin receptor activation triggers converging regulation of Cav1.2 calcium channels by c-Src and protein kinase A pathways.

L-type, voltage-gated Ca2+ channels (CaL) play critical roles in brain and muscle cell excitability. Here we show that currents through heterologously expressed neuronal and smooth muscle CaL channel isoforms are acutely potentiated following alpha5beta1 integrin activation. Only the alpha1C pore-forming channel subunit is critical for this process. Truncation and site-directed mutagenesis strategies reveal that regulation of Cav1.2 by alpha5beta1 integrin requires phosphorylation of alpha1C C-terminal residues Ser1901 and Tyr2122. These sites are known to be phosphorylated by protein kinase A (PKA) and c-Src, respectively, and are conserved between rat neuronal (Cav1.2c) and smooth muscle (Cav1.2b) isoforms. Kinase assays are consistent with phosphorylation of these two residues by PKA and c-Src. Following alpha5beta1 integrin activation, native CaL channels in rat arteriolar smooth muscle exhibit potentiation that is completely blocked by combined PKA and Src inhibition. Our results demonstrate that integrin-ECM interactions are a common mechanism for the acute regulation of CaL channels in brain and muscle. These findings are consistent with the growing recognition of the importance of integrin-channel interactions in cellular responses to injury and the acute control of synaptic and blood vessel function.

Animals↗

PubMed2001

Cell-surface transglutaminase promotes fibronectin assembly via interaction with the gelatin-binding domain of fibronectin: a role in TGFbeta-dependent matrix deposition.

Assembly of fibronectin into a fibrillar matrix is critical for regulation of cell growth and migration, embryogenesis and wound healing. We have previously shown that cell-surface tissue transglutaminase serves as an integrin-binding adhesion coreceptor for fibronectin. Here we report that transglutaminase strongly promotes fibronectin assembly mediated by alpha5beta1 integrin. This effect is independent from transglutaminase-mediated enzymatic crosslinking of fibronectin and separate from the ability of transglutaminase to stimulate cell spreading. Surface transglutaminase increases the binding of fibronectin to cells via interaction with its gelatin-binding domain that contains modules I6II1,2I7-9 and lacks integrin-binding motifs. The gelatin-binding fragment of fibronectin binds to surface transglutaminase on cells in suspension but does not interact with cell monolayers where surface transglutaminase is occupied by fibronectin. Surface transglutaminase colocalizes with growing fibronectin fibrils at early timepoints of matrix formation and remains codistributed with fibronectin matrices thereafter. The observed stimulation of matrix assembly by transglutaminase is blocked by the gelatin-binding fragment of fibronectin, but is not strongly perturbed by its N-terminal fragment consisting of modules I1-5. These results implicate an interaction between transglutaminase and the gelatin-binding domain of fibronectin in matrix assembly and suggest its role in initiation of fibrillogenesis. However, blocking antibodies against alpha5beta1 integrin or the cell-binding fragment of fibronectin that contains modules III2-11 most strongly suppress matrix formation and abolish the effects of transglutaminase. Hence, transglutaminase cooperates with but can not substitute for alpha5beta1 integrin in fibronectin assembly. Treatment of fibroblasts with transforming growth factor beta (TGFbeta) significantly increases surface expression of transglutaminase and its association with beta1 integrins, but not with alphaVbeta3 integrin. TGFbeta enhances the binding of fibronectin to the cell surface and elevates matrix formation, whereas antibody against transglutaminase or the gelatin-binding fragment of fibronectin suppresses these effects, indicating an involvement of transglutaminase in TGFbeta-dependent fibronectin assembly. Therefore, TGFbeta-induced fibronectin matrix deposition during normal wound healing or fibrotic disorders may depend on upregulation of integrin-associated surface transglutaminase.

Animals↗

PubMed2006

Microbial pattern recognition receptors mediate M-cell uptake of a gram-negative bacterium.

The receptors involved in the sampling of particulate microbial antigens by the gut are largely unknown. Here we demonstrate for the first time in an in vitro M-cell model and in situ in isolated murine intestinal segments that the receptors TLR-4, PAF-R, and alpha5beta1 integrin are all involved in mediating bacterial uptake associated with transcytosis. The pattern of expression of TLR-4 and alpha5beta1 integrin differed between M cells and enterocytes. There was increased apical expression of TLR-4 in M-cell cultures, and it was present on the apical surface of murine M cells but not enterocytes in situ. In contrast, PAF-R was expressed equally by both cell types in vitro and was abundantly expressed throughout the intestinal epithelium. Inhibition of TLR-4 and PAF-R, but not TLR-2, reduced gram-negative bacterial uptake by both cell types, whereas inhibition of the apically expressed alpha5beta1 integrin significantly reduced the ability of M cells to translocate bacteria. Hence, the involvement of each receptor was dependent not only on differences in the level of receptor expression but the cellular localization. Using bacteria that had mutations that affected the bacterial lipooligosaccharide structure indicated that the oligosaccharide moiety was important in bacterial uptake. Taken together, the data suggest that pathogen-associated molecular pattern interactions with pattern recognition receptors are key factors in M-cell recognition of intestinal antigens for mucosal immune priming.

Animals↗

PubMed2005

In vitro attachment of bovine hatched blastocysts on fibronectin is mediated by integrin in a RGD dependent manner.

We investigated the effect of extracellular matrix protein on in vitro attachment and outgrowth of bovine hatched blastocysts. In vitro produced bovine hatched blastocysts were cultured on a fibronectin- or laminin-coated Petri dishes. Hatched blastocysts adhered and outgrew on the fibronectin-coated dish whereas no attachment was observed on the laminin-coated dish. The attachment and outgrowth on fibronectin were significantly inhibited in the presence of synthetic peptides containing the Arg-Gly-Asp (RGD) sequence, which interacts with the fibronectin receptor (integrin alpha5beta1), but were not inhibited by the control peptides containing the Arg-Gly-Glu (RGE) sequence. Addition of anti-fibronectin receptor (integrin alpha5beta1) antibody to the culture medium also inhibited the attachment and outgrowth on fibronectin-coated Petri dishes. Subsequently we examined mRNA expression and protein expression of alpha5 and beta1 integrin subunit in the hatched blastocyst by reverse transcription-polymerase chain reaction (RT-PCR) and immunostaining, respectively. Expression of both mRNA and protein were detected in blastocysts. These results indicate that trophectoderm cells of bovine hatched blastocysts have already acquired the ability to adhere and outgrow on fibronectin in vitro by an integrin- mediated manner.

Animals↗

PubMed2005

Inhibitory effects of Porphyromonas gingivalis fimbriae on interactions between extracellular matrix proteins and cellular integrins.

Porphyromonas gingivalis is a predominant periodontal pathogen, whose fimbriae are considered to be a major virulence factor, especially for bacterial adherence and invasion of host cells. In the present study, we investigated the influence of fimbriae on the interactions between alphavbeta3- and alpha5beta1-integrins and their ligand extracellular matrix (ECM) proteins (vitronectin and fibronectin), using human alphavbeta3- and alpha5beta1-integrin-overexpressing CHO cell lines (CHOalphavbeta3 and CHOalpha5beta1, respectively). P. gingivalis was found to have significantly greater binding to CHOalphavbeta3 and CHOalpha5beta1 than to control cells, whereas a fimbria-deficient mutant showed negligible binding to any of the tested cell lines. CHOalphavbeta3 and CHOalpha5beta1 cells attached to the polystyrene culture dishes in the presence of their ligand ECM proteins, while fimbriae markedly inhibited those attachments in a dose-dependent manner, with the highest dose of fimbriae achieving complete inhibition. In addition, the binding of vitronectin and fibronectin to CHOalphavbeta3 and CHOalpha5beta1 was inhibited by P. gingivalis cells. These results suggest that P. gingivalis fimbriae compete with ECM proteins for alphavbeta3- and alpha5beta1-integrins, and inhibit integrin/ECM protein-related cellular functions.

Animals↗

PubMed1999

Targeted disruption of fibronectin-integrin interactions in human gingival fibroblasts by the RI protease of Porphyromonas gingivalis W50.

Cell surface integrins mediate interactions between cells and their extracellular matrix and are frequently exploited by a range of bacterial pathogens to facilitate adherence and/or invasion. In this study we examined the effects of Porphyromonas gingivalis proteases on human gingival fibroblast (HGF) integrins and their fibronectin matrix. Culture supernatant from the virulent strain W50 caused considerably greater loss of the beta1 integrin subunit from HGF in vitro than did that of the beige-pigmented strain W50/BE1. Prior treatment of the W50 culture supernatant with the protease inhibitor Nalpha-p-tosyl-L-lysine chloromethyl ketone (TLCK) blocked its effects on cultured cells, indicating that this process is proteolytically mediated. Purified arginine-specific proteases from P. gingivalis W50 were able to mimic the effects of the whole-culture supernatant on loss of beta1 integrin expression. However purified RI, an alpha/beta heterodimer in which the catalytic chain is associated with an adhesin chain, was 12 times more active than RIA, the catalytic monomer, in causing loss of the alpha5beta1 integrin (fibronectin receptor) from HGF. No effect was observed on the alphaVbeta3 integrin (vitronectin receptor). The sites of action of RI and RIA were investigated in cells exposed to proteases pretreated with TLCK to inactivate the catalytic component. Use of both monoclonal antibody 1A1, which recognizes only the adhesin chain of RI, and a rabbit antibody against P. gingivalis whole cells indicated localization of RI on the fibroblasts in a clear, linear pattern typical of that seen with fibronectin and alpha5beta1 integrin. Exact colocalization of RI with fibronectin and its alpha5beta1 receptor was confirmed by double labeling and multiple-exposure photomicroscopy. In contrast, RIA bound to fibroblasts in a weak, patchy manner, showing only fine linear or granular staining. It is concluded that the adhesin component of RI targets the P. gingivalis arginine-protease to sites of fibronectin deposition on HGF, contributing to the rapid loss of both fibronectin and its main alpha5beta1 integrin receptor. Given the importance of integrin-ligand interactions in fibroblast function, their targeted disruption by RI may represent a novel mechanism of damage in periodontal disease.

Animals↗

PubMed1999

Expression and function of cell adhesion molecules on fetal liver, cord blood and bone marrow hematopoietic progenitors: implications for anatomical localization and developmental stage specific regulation of hematopoiesis.

The mechanism of localization, migration, and regulation of hematopoiesis at different stages of ontogeny is not well understood, but may relate to the specific adhesive interactions between hematopoietic stem cells and their microenvironment at different ontogenic stages. We studied the expression of cell adhesion molecules (CAM) on fetal liver (FL), umbilical cord blood (UCB) and adult bone marrow (ABM) CD34+ cells, and the adhesion of committed progenitors (CFC) from all three sources to ABM stromal layers and purified extracellular matrix proteins. Compared to ABM CFC, significantly more UCB CFC and fewer FL CFC adhered to ABM stroma. Adhesion of FL CFC to fibronectin (FN), the 75 kD RGD containing FN fragment and the 33-66 kD COOH-terminal heparin binding FN fragment was also significantly less than that of ABM CFC. Like ABM CFC, the adhesion of FL CFC was mediated through alpha4beta1 and alpha5beta1 integrins. Of note, more FL CD34+ cells expressed alpha5 integrins and the number of alpha4, alpha5 and beta1 integins per cell (mean channel frequency) was similar or higher for FL CD34+ cells than ABM CD34+ cells. Further, treatment of FL CFC with a beta1 integrin activating antibody (8A2), increased adhesion of FL CFC to FN to the same level as that of 8A2 treated ABM CFC. This suggests that the alpha4beta1 and alpha5beta1 integrins on FL CD34+ cells may be present in a low avidity/affinity state. We also show that unlike ABM, FL CD34+ cells expressed alpha2 and that approximately 20% FL CFC adhered to collagen IV. Further, alpha2beta1 integrin on FL CFC was functional since their engagement, either by adhesion to collagen IV or through blocking alpha2 antibodies, transmitted proliferation inhibitory signals. In contrast to alpha4b and alpha5beta1 integrin dependent adhesion, alpha2beta1 dependent adhesion of FL CFC to collagen IV was not enhanced after treatment with 8A2. The reason for this is not clear but suggests that alpha2 integrins on FL CFC are maximally activated. This novel adhesive interaction with collagen IV, reminiscent of that described for CML progenitors, may have a role in the extramedullary localization of FL hematopoiesis or its developmental stage-specific regulation by its microenvironment. Studies to evaluate these possibilities are underway.

Adult↗

PubMed2002

Inhibition of IL-8-mediated MAPK activation in human neutrophils by beta1 integrin ligands.

Chemokine and integrin receptors must work in concert when circulating leukocytes mobilize toward a site of tissue inflammation or infection. In a previous study, we reported that ligation of the alpha5beta1 integrin with a 120-kDa cell-binding fibronectin fragment (120-kDa FN) in suspensions of human polymorphonuclear leukocytes (PMNLs) inhibited chemotaxis toward the chemokine called interleukin-8 (IL-8). Binding of chemokines to their receptors on leukocytes leads to the activation of heterotrimeric G proteins that initiate multiple signaling cascades, including p38 and p42/p44 mitogen-activated protein kinase (MAPK) pathways. In the present study, we examine the potential interaction of beta1 integrin ligation on chemokine-mediated MAPK signaling in human PMNLs. We demonstrate that blockade of the p42/p44 MAPK signaling pathway by the inhibitor PD98059 suppresses IL-8-mediated PMNL chemotaxis. Furthermore, when PMNLs are pretreated with 120-kDa FN or an activating antibody to beta1 integrins (TS2/16), IL-8-mediated phosphorylation of p42/p44 MAPK is also inhibited. In contrast, pretreating PMNL with a specific ligand (laminin-1) for the alpha6beta1 integrin does not suppress IL-8-mediated phosphorylation of p42/p44 MAPK. These observations demonstrate a desensitization of IL-8-mediated p42/p44 MAPK signaling in response to ligation of the alpha5beta1 integrin in PMNL. Also, they suggest an interplay between integrin and chemokine signaling during PMNL migration through the extracellular matrix.

Antibodies, Monoclonal↗

PubMed2003

Beta1 integrins are distributed in adhesion structures with fibronectin and caveolin and in coated pits.

Integrins are found in adhesion structures, which link the extracellular matrix to cytoskeletal proteins. Here, we attempt to further define the distribution of beta1 integrins in the context of their association with matrix proteins and other cell surface molecules relevant to the endocytic process. We find that beta1 integrins colocalize with fibronectin in fibrillar adhesion structures. A fraction of caveolin is also organized along these adhesion structures. The extracellular matrix protein laminin is not concentrated in these structures. The alpha4beta1 integrin exhibits a distinct distribution from other beta1 integrins after cells have adhered for 1 h to extracellular matrix proteins but is localized in adhesion structures after 24 h of adhesion. There are differences between the fibronectin receptors: alpha5beta1 integrins colocalize with adaptor protein-2 in coated pits, while alpha4beta1 integrins do not. This parallels our earlier observation that of the two laminin receptors, alpha1beta1 and alpha6beta1, only alpha1beta1 integrins colocalize with adaptor protein-2 in coated pits. Calcium chelation or inhibition of mitogen-activated protein kinase kinase, protein kinase C, or src did not affect localization of alpha1beta1 and alpha5beta1 integrins in coated pits. Likewise, the integrity of coated-pit structures or adhesion structures is not required for integrin and adaptor protein-2 colocalization. This suggests a robust and possibly constitutive interaction between these integrins and coated pits.

Adaptor Protein Complex 2↗

PubMed2003

Immunohistochemical localisation of extracellular matrix proteins in the periodontium during cementogenesis in the rat molar.

OBJECTIVE: The development of the periodontium involves the coordinated expression of numerous extracellular matrix (ECM) macromolecules and their receptors (integrins). The aim of this study was to determine the expression of selected hard and soft tissue matrix molecules and the integrin alpha5beta1 in the periodontal tissues, during cementogenesis in the rat molar. METHODS: Using immunohistochemical methods, the distribution of the extracellular matrix proteins, fibronectin, tenascin, and bone sialoprotein (BSP), as well as the integrin subunits alpha5 and beta1 were studied in rats aged 3, 5 and 8 weeks. RESULTS: Fibronectin was widely distributed in the gingival epithelium, gingival connective tissue and in the periodontal ligament. Tenascin expression was less marked compared with fibronectin, but was more distinctly associated with cells and peri-cellular areas of the epithelial-connective tissue interface, the gingiva and within the periodontal ligament. The fibronectin-receptor alpha5beta1 integrins were expressed by epithelial cells, periodontal ligament cells and gingival fibroblasts. A notable finding was the increased staining intensity of fibronectin, tenascin and alpha5beta1 integrin in all 5-week old molar sections in the periodontal ligament matrix and cells, apical to the cemento-enamel junction (CEJ) along the alveolar crest (AC) ridge height. Bone sialoprotein was distinctly associated with the hard tissues of the periodontium as acellular cementum and alveolar bone matrix expressed bone sialoprotein throughout all sections, in all age groups. CONCLUSIONS: In conclusion, this study has demonstrated the selective distribution of several hard and soft tissue matrix molecules during periodontogenesis. The results highlight the complex nature of interactions of various proteins and molecules during development. The interactions between these molecules and their specific roles in development and regeneration await further investigation.

Animals↗

PubMed1996

Beta1 integrin-mediated activation of focal adhesion kinase and its association with Fyn and Zap-70 in human NK cells.

Adhesive interactions mediated by cell surface receptors have been shown to induce signal transduction pathways that regulate changes in cellular function. We have reported recently that fibronectin (FN) receptors, alpha4beta1 and alpha5beta1 integrins, on NK cells transduce transmembrane signals leading to tyrosine phosphorylation of 60-, 70-, and 120-kDa proteins. In the current study, we have identified a 120-kDa phosphoprotein as the focal adhesion kinase (p125FAK), a structurally unique nonreceptor protein tyrosine kinase that localizes to focal adhesions. Activity of p125FAK was induced by adhesion of NK cells to plastic-immobilized FN, by cross-linking of cell surface-bound FN or FN fragments, FN120 or FN40, with anti-FN mAb, or by cross-linking of alpha4beta1 or alpha5beta1 integrins with alpha-chain-specific Abs. We also observed that enhanced in vitro kinase activity was associated with immunoprecipitates of alpha4beta1 or alpha5beta1 integrins from lysates of FN-adherent NK cells as compared with BSA-treated NK cells. In addition to p125FAK activity, FN-induced kinase activity was also found to be mediated by Fyn, Lyn, and Zap-70, as assessed by in vitro phosphorylation of the immunoprecipitated kinases in the presence of [gamma-32P]ATP. Clustering of FN receptors on NK cells by agonists such as immobilized FN or alpha4- or alpha5-specific Abs also induced association of Fyn and Zap-70 with p125FAK. Our observations indicate that activation and phosphorylation of p125FAK as well as Zap-70 and certain kinases of the src family play an important role in formation of active signaling complexes in response to triggering via beta1 integrins on NK cells. These results also suggest the existence of cross-talk or points of convergence between the beta1 integrin-mediated and other receptor-signaling pathways.

Cell Adhesion↗

PubMed1999

Endoglin overexpression modulates cellular morphology, migration, and adhesion of mouse fibroblasts.

Endoglin is the gene mutated in hereditary hemorrhagic telangiectasia type 1 (HHT1), a dominantly inherited vascular disorder. Endoglin glycoprotein is a component of the transforming growth factor type beta (TGF-beta) receptor system which is highly expressed by endothelial cells, and at lower levels on fibroblasts and smooth muscle cells, suggesting the involvement of these lineages in the HHT1 vascular dysplasia. Overexpression of endoglin in mouse NCTC929 fibroblasts led to decreased migration in chemotactic and wound healing assays, as well as changes in the cellular morphology. When plated on uncoated surfaces, endoglin transfectants formed intercellular clusters, endoglin being not specifically localized to the cell-cell junctions, but homogenously distributed on the cellular surface. Although the expression of alpha5beta1 integrin and of an activation epitope of beta1 integrin were unchanged, a polyclonal antibody to alpha5beta1 integrin was able to inhibit cluster formation, suggesting the involvement of integrin ligand/s. In fact, coating with fibronectin, laminin, or an RGD-containing 80 kDa fragment of fibronectin were able to prevent the cellular clustering. Furthermore, synthesis of plasminogen activator inhibitor 1 (PAI-1), and to a weak extent that of fibronectin, were inhibited in endoglin transfectants. Thus, the presence of endoglin in mouse NCTC929 fibroblasts is associated with reduced production of certain extracellular matrix (ECM) components, which might explain their altered morphology, migration and intercellular cluster formation.

Animals↗

PubMedDate not supplied

Metastatic properties of prostate cancer cells are controlled by VEGF.

Mechanisms of metastasis, the major complication of prostate cancer, are poorly understood. In this study, we define molecular mechanisms that may contribute to the highly invasive potential of prostate cancer cells. Vascular endothelial growth factor (VEGF), its receptors (VEGFRs), and alpha5beta1 integrin were expressed by prostate cancer cells in vitro and by prostate tumors in vivo, and their expression was elevated at sites of bone metastasis compared to original prostate tumor. VEGF, through interaction with its receptors, regulated adhesive and migratory properties of the cancer cells. Specifically, the highly metastatic prostate cancer cell subline LNCaP-C4-2 showed a decreased adhesive but an enhanced migratory response to fibronectin, a ligand for alpha5beta1 integrin, compared to its nonmetastatic counterpart. A similar pattern was also observed when bone sialoprotein was used as a ligand in migration assays. Increased migration of metastatic prostate cancer cells to fibronectin and bone sialoprotein was regulated by VEGF via VEGFR-2. Tumor suppressor PTEN was involved in control of VEGF/VEGFR-2 stimulated prostate cancer cell adhesion as well as proliferation.

Bone Neoplasms↗

PubMed2001

Blockade of alpha 5 beta 1 integrins reverses the inhibitory effect of tenascin on chemotaxis of human monocytes and polymorphonuclear leukocytes through three-dimensional gels of extracellular matrix proteins.

Tenascin is an extracellular matrix protein found in adults in T cell-dependent areas of lymphoid tissues, sites of inflammation, and tumors. We report here that it inhibited chemotaxis of chemoattractant-stimulated human monocytes and chemoattractant-stimulated polymorphonuclear leukocytes (PMN) through three-dimensional gels composed of collagen I or Matrigel, and chemotaxis of leukotriene B4-stimulated PMN through fibrin gels. The inhibitory effect of tenascin on monocyte or PMN chemotaxis through these matrices was reversed by Abs directed against alpha5beta1 integrins or by a peptide (GRGDSP) that binds to beta1 integrins. Tenascin did not affect leukotriene B4- or fMLP-stimulated expression of beta1 or beta2 integrins, but did exert a small inhibitory effect on PMN adhesion and closeness of apposition to fibrin(ogen)-containing surfaces. Thus, alpha5beta1 integrins mediate the inhibitory effect of tenascin on monocyte and PMN chemotaxis, without promoting close apposition between these leukocytes and surfaces coated with tenascin alone or with tenascin bound to other matrix proteins. This contrasts with the role played by alpha5beta1 integrins in promoting close apposition between fMLP-stimulated PMN and fibrin containing surfaces, thereby inhibiting chemotaxis of fMLP-stimulated PMN through fibrin gels. Thus, chemoattractants and matrix proteins regulate chemotaxis of phagocytic leukocytes by at least two different mechanisms: one in which specific chemoattractants promote very tight adhesion of leukocytes to specific matrix proteins and another in which specific matrix proteins signal cessation of migration without markedly affecting strength of leukocyte adhesion.

Animals↗

PubMed2003

Identification of redundant angiogenic sites in laminin alpha1 and gamma1 chains.

The degradation of the extracellular matrix is one of the first steps involved in angiogenesis, the formation of new vessels from preexisting ones. Laminin, a large extracellular matrix protein, has many biological activities, including the promotion of angiogenesis. Screening of the laminin-1 chains identified 20 angiogenic peptides, of which, A13 and C16, from the alpha1 and gamma1 chains, respectively, were the most active. We recently identified the receptors for C16 as the integrins alpha5beta1 and alphavbeta3. Here, we show unexpectedly that A13 is a redundant active site to C16 present in the N-terminal globular domain of the alpha1 chain. The peptides are located in homologous sites present in the last globular domains of their respective chains, and their amino acids are 66% conserved, as compared to the inactive homologous site in the beta1 chain, B19 to B20, which is only 18%-23% conserved. Cell attachment studies demonstrated that both A13 and C16 reciprocally inhibited their adhesion activity, whereas the corresponding laminin beta1 chain peptides were inactive. Chorioallantoic membrane assays showed that the in vivo angiogenic activity of A13 is blocked by a C16 antagonist, C16S, which also binds to the same integrin receptors. A13 affinity chromatography and immunoprecipitation analysis showed that the alphavbeta3 and alpha5beta1 integrin receptors bind to this sequence. We have therefore identified redundant activity on two laminin chains. These highly conserved functional sites are likely important mediators of the biological responses of laminins because either one or both of these chains (active sites) are present in almost all laminin isoforms identified to date.

Amino Acid Sequence↗

PubMed2004

Brain development in mice lacking L1-L1 homophilic adhesion.

A new mouse line has been produced in which the sixth Ig domain of the L1 cell adhesion molecule has been deleted. Despite the rather large deletion, L1 expression is preserved at normal levels. In vitro experiments showed that L1-L1 homophilic binding was lost, along with L1-alpha5beta1 integrin binding. However, L1-neurocan and L1-neuropilin binding were preserved and sema3a responses were intact. Surprisingly, many of the axon guidance defects present in the L1 knockout mice, such as abnormal corticospinal tract and corpus callosum, were not observed. Nonetheless, when backcrossed on the C57BL/6 strain, a severe hydrocephalus was observed and after several generations, became an embryonic lethal. These results imply that L1 binding to L1, TAG-1, or F3, and L1-alpha5beta1 integrin binding are not essential for normal development of a variety of axon pathways, and suggest that L1-L1 homophilic binding is important in the production of X-linked hydrocephalus.

Animals↗

PubMed2005

Galectin-1 interacts with the {alpha}5{beta}1 fibronectin receptor to restrict carcinoma cell growth via induction of p21 and p27.

Surface binding of galectin family members has the potential to link distinct glycan structures to growth regulation. Therefore, we addressed the antiproliferative potential of galectin-1 (Gal-1) in a panel of carcinoma cell lines. We discovered growth inhibition by Gal-1 in epithelial tumor cell lines from different origins and provide evidence that this effect requires functional interaction with the alpha5beta1 integrin. Antiproliferative effects result from inhibition of the Ras-MEK-ERK pathway and consecutive transcriptional induction of p27. We have further identified two Sp1-binding sites in the p27 promoter as crucial for Gal-1 responsiveness. Inhibition of the Ras-MEK-ERK cascade by Gal-1 increased Sp1 transactivation and DNA binding due to reduced threonine phosphorylation of Sp1. Furthermore, Gal-1 induced p21 transcription and selectively increased p27 protein stability. Gal-1-mediated accumulation of p27 and p21 inhibited cyclin-dependent kinase 2 activity and ultimately resulted in G(1) cell cycle arrest and growth inhibition. These data define a novel mechanism whereby Gal-1 regulates epithelial tumor cell homeostasis via carbohydrate-dependent interaction with the alpha5beta1 integrin.

Adenocarcinoma↗

PubMed2002

The role of syndecan-2 in regulation of actin-cytoskeletal organization of Lewis lung carcinoma-derived metastatic clones.

Syndecans, a family of transmembrane heparan sulphate proteoglycans, contribute to various biological processes, including adhesion, motility, proliferation, differentiation and morphogenesis. We document here the involvement of syndecan-2 acting alone or co-operatively with integrin alpha5beta1, for regulation of actin-cytoskeletal organization on cell adhesion to fibronectin, using fibronectin-recombinant polypeptides containing the ligands for either or both of these receptors as substrata. Lewis lung carcinoma-derived low-metastatic P29 cells binding to the substrata by both receptors formed actin stress fibres, whereas those binding by syndecan-2 or integrin alpha5beta1 alone formed filopodia or cortex actin. In contrast, higher metastatic LM66-H11 cells formed cortex actin even on substrata containing both ligands. Northern-blot and flow-cytometric analyses revealed that syndecan-2 expression in LM66-H11 cells was significantly lower (1/4.5 in mRNA and 1/8 in cell-surface expression) than in P29 cells, whereas expression levels of integrin alpha5beta1 and other syndecans were similar in both cell types. These results suggest that the failure of LM66-H11 to form stress fibres is due to a lower expression of syndecan-2 than that due to a threshold for its function. This was confirmed by the finding that overexpression of syndecan-2 by transfection of its cDNA into LM66-H11 cells caused the formation of stress fibres on the fibronectin substratum. These in vitro cellular responses of the two clones might reflect their in vivo situation in primary tumours in which P29 cells with a stroma-inducing capacity were immediately surrounded by fibronectin-rich matrix formed by the induced stromal cells, whereas LM66-H11 cells without such capacity were not surrounded by a similar matrix.

Actins↗