Journal articles on the topic 'Alpha4 integrins'
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1
Manie, SN, A. Astier, D. Wang, JS Phifer, J. Chen, AI Lazarovits, C. Morimoto, and AS Freedman. "Stimulation of tyrosine phosphorylation after ligation of beta7 and beta1 integrins on human B cells." Blood 87, no. 5 (March 1, 1996): 1855–61. http://dx.doi.org/10.1182/blood.v87.5.1855.1855.
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Abstract B lymphocytes express several members of the integrin family of adhesion molecules that mediate cell-cell and cell-extracellular matrix interactions. In addition to beta1 integrins, predominantly alpha4 beta1, mature B cells also express alpha4 beta7, which is a receptor for vascular cell adhesion molecule-1 and fibronectin, and is also involved in the homing of B cells to mucosal sites through binding to a third ligand, mucosal address in cell adhesion molecule-1. Here we describe that crosslinking of alpha4 beta7 integrins on B cell lines and normal tonsillar B cells, induces tyrosine phosphorylation of multiple substrates of 105–130 kD, indicating that beta7 integrin plays a role as signaling molecule in B cells. This pattern of phosphorylated proteins was very similar to that induced following ligation of alpha4 beta1. Interestingly, ligation of alpha5 beta1 or alpha6 beta1 also stimulated the 105–125 kD group of phosphorylated proteins, whereas ligation of beta2 integrins did not. The focal adhesion tyrosine kinase p125FAK was identified as one of these substrates. Beta1 or beta7 mediated tyrosine phosphorylations were markedly decreased when the microfilament assembly was inhibited by cytochalasin B. These results suggest that intracellular signals initiated by different integrins in B cells may converge, to similar cytoskeleton-dependent tyrosine phosphorylated proteins.
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Manie, SN, A. Astier, D. Wang, JS Phifer, J. Chen, AI Lazarovits, C. Morimoto, and AS Freedman. "Stimulation of tyrosine phosphorylation after ligation of beta7 and beta1 integrins on human B cells." Blood 87, no. 5 (March 1, 1996): 1855–61. http://dx.doi.org/10.1182/blood.v87.5.1855.bloodjournal8751855.
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B lymphocytes express several members of the integrin family of adhesion molecules that mediate cell-cell and cell-extracellular matrix interactions. In addition to beta1 integrins, predominantly alpha4 beta1, mature B cells also express alpha4 beta7, which is a receptor for vascular cell adhesion molecule-1 and fibronectin, and is also involved in the homing of B cells to mucosal sites through binding to a third ligand, mucosal address in cell adhesion molecule-1. Here we describe that crosslinking of alpha4 beta7 integrins on B cell lines and normal tonsillar B cells, induces tyrosine phosphorylation of multiple substrates of 105–130 kD, indicating that beta7 integrin plays a role as signaling molecule in B cells. This pattern of phosphorylated proteins was very similar to that induced following ligation of alpha4 beta1. Interestingly, ligation of alpha5 beta1 or alpha6 beta1 also stimulated the 105–125 kD group of phosphorylated proteins, whereas ligation of beta2 integrins did not. The focal adhesion tyrosine kinase p125FAK was identified as one of these substrates. Beta1 or beta7 mediated tyrosine phosphorylations were markedly decreased when the microfilament assembly was inhibited by cytochalasin B. These results suggest that intracellular signals initiated by different integrins in B cells may converge, to similar cytoskeleton-dependent tyrosine phosphorylated proteins.
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Kawamoto, Eiji, Susumu Nakahashi, Takayuki Okamoto, Hiroshi Imai, and Motomu Shimaoka. "Anti-Integrin Therapy for Multiple Sclerosis." Autoimmune Diseases 2012 (2012): 1–6. http://dx.doi.org/10.1155/2012/357101.
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Integrins are the foremost family of cell adhesion molecules that regulate immune cell trafficking in health and diseases. Integrin alpha4 mediates organ-specific migration of immune cells to the inflamed brain, thereby playing the critical role in the pathogenesis of multiple sclerosis. Anti-alpha4 integrin therapy aiming to block infiltration of autoreactive lymphocytes to the inflamed brain has been validated in several clinical trials for the treatment of multiple sclerosis. This paper provides readers with an overview of the molecular and structural bases of integrin activation as well as rationale for using anti-alpha4 integrin therapy for multiple sclerosis and then chronicles the rise and fall of this treatment strategy using natalizumab, a humanized anti-alpha4 integrin.
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Rose, David M., Jaewon Han, and Mark H. Ginsberg. "alpha4 integrins and the immune response." Immunological Reviews 186, no. 1 (August 2002): 118–24. http://dx.doi.org/10.1034/j.1600-065x.2002.18611.x.
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Escudero, E., M. Nieto, A. Martín, A. Molina, R. R. Lobb, F. Sanchez-Madrid, and F. Mampaso. "Differential effects of antibodies to vascular cell adhesion molecule-1 and distinct epitopes of the alpha4 integrin in HgCl2-induced nephritis in Brown Norway rats." Journal of the American Society of Nephrology 9, no. 10 (October 1998): 1881–91. http://dx.doi.org/10.1681/asn.v9101881.
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Four distinct epitopes (A, B1, B2, and C) have been functionally defined on the human alpha4 integrin. In this study, two cross-reactive antihuman alpha4 monoclonal antibodies (mAb) (HP2/1 and HP2/4 specific for epitopes B1 and B2, respectively) were used to functionally characterize the rat VLA-4 subunit and to define similar functional epitopes in this rodent species. It was found that B1 and B2 anti-alpha4 mAb completely block adhesion to fibronectin, but the inhibition of adhesion to vascular cell adhesion molecule-1 (VCAM-1) with HP2/1 mAb was lower than with HP2/4 mAb. It was also observed that epitope B2 HP2/4 mAb induced homotypic aggregation in rat lymphocytes, whereas epitope B1 HP2/1 mAb did not. Using the HgCl2 model of nephritis, this study shows the protective effect of both anti-alpha4 mAb against infiltration of the renal interstitium by leukocytes. Nevertheless, HP2/1 mAb, but not HP2/4 mAb, virtually abolished the anti-glomerular basement membrane antibody synthesis and glomerular deposits. These findings indicate the dual but independent role played by alpha4 integrins in both extravasation of leukocytes and in the production of antibodies. Finally, this study demonstrates that anti-rat VCAM-1 mAb showed a positive reactivity of the renal vascular endothelium and, most importantly, that administration of anti-VCAM-1 antibodies completely abrogated the interstitial cell infiltrates without affecting anti-glomerular basement membrane antibody production. These results confirm the important role played by VLA-4/VCAM-1 pathway in leukocyte infiltration, and further support the dual and independent role of alpha4 integrins in both renal infiltration and autoantibody synthesis in this model of renal disease.
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Hauzenberger, D., J. Klominek, J. Holgersson, S. E. Bergström, and K. G. Sundqvist. "Triggering of motile behavior in T lymphocytes via cross-linking of alpha 4 beta 1 and alpha L beta 2." Journal of Immunology 158, no. 1 (January 1, 1997): 76–84. http://dx.doi.org/10.4049/jimmunol.158.1.76.
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Abstract The mechanisms by which T lymphocytes are transformed from passively transported cells during circulation in the vascular system to actively migrating cells during extravasation are unknown. Therefore, the possibility that lymphocyte receptors are capable of inducing motility was investigated using a modified Boyden chamber assay. Cross-linking of alphaL beta2 and alpha4 beta1 on human T lymphocytes (T cell line and peripheral blood T cells) with immobilized mAbs induced motile behavior on fibronectin, laminin, collagen type IV, and poly-L-lysine. This induction of T cell migration was very potent and in most cases more efficient than pretreatment of the cells with phorbol esters. In contrast, control Abs to several other integrin- and non-integrin molecules present on T lymphocytes did not induce T cell migration. Anti-CD3 Abs themselves did not trigger motile behavior. However, anti-CD3 promoted T cell migration in the Boyden chamber system if present simultaneously with 40-kDa alpha4 beta1 binding fibronectin fragments or alphaL beta2 binding intercellular adhesion molecule-1/hIgG1Fc fusion proteins on the upper side of the filter. Abs to other surface components on T cells did not trigger motility when presented together with the 40-kDa fibronectin fragments or the intercellular adhesion molecule-1/hIgG1Fc fusion proteins. The induction of motile behavior could be blocked if the T cells were pretreated with Genistein and Calphostin C, indicating the involvement of a protein tyrosine kinase and protein kinase C-dependent signaling pathway in triggering of T cell motility via integrins. These results indicate that alphaL beta2 and alpha4 beta1 on T lymphocytes can selectively trigger motile behavior when cross-linked by their endothelial or extracellular matrix ligands. Furthermore, these data indicate that cross-linking of CD3 facilitates ligand binding and subsequent triggering of a motile phenotype by alphaL beta2 and alpha4 beta1.
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Ulyanova, Tatiana, Ena R. Banerjee, Gregory V. Priestley, Linda Scott, and Thalia Papayannopoulou. "Migration Defects of Leukocytes from Alpha4 or Beta2 Integrin-Deficient Mice during Aseptic Peritonitis." Blood 104, no. 11 (November 16, 2004): 2387. http://dx.doi.org/10.1182/blood.v104.11.2387.2387.
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Abstract Leukocyte migration to the inflammatory sites involves a dynamic and coordinated sequence of adhesion/ migration events, in which the α4 and β2 integrins in leukocytes play critical roles. In a model of aseptic peritonitis, normal mice respond with an initial influx of neutrophils followed by later recruitment of mononuclear cells. Previous studies used antibodies to inhibit α4 integrins in this model and provided only early observations (up to 24 hrs). Events at later stages were not addressed. To clarify the role of α4 integrins in this process, we induced aseptic peritonitis in α4-ablated (α4−/ −) mice and compared the results to similarly treated wild type (WT), and β2 integrin-deficient (β2−/ −) animals. A 4, 16 and 96 hrs post thioglycollate injection, leukocytes, harvested from the peritoneal cavity, were counted and evaluated by FACS and by differentials in smears. 3-5 animals per time point per genotype were analyzed. Early neutrophil accumulation in the peritoneum was similar in WT and α4−/ − mice (Table 1). β2−/ − mice were slow to accumulate neutrophils, but at 16 hrs the peaks were not different from α4−/ − or WT animals. However at later times in both α4−/ − and β2−/ − mice neutrophils persisted longer, suggesting either increased survival or prolonged recruitment. At 16 hrs large numbers of monocyte/ macrophages accumulated in the peritoneum and they were similar in all groups of mice up to 96 hours. The cumulative number of leukocytes in the peritoneum was compared to the concurrent circulating pool of leukocytes in each mouse group. At the peak time (16 hrs) the fraction recovered in the WT represented 394±148%, while in the α4−/ − mice it was 98±33% and in the β2−/ − mice it was 40±19%. Thus, although sufficient numbers of leukocytes accumulated in the peritoneum of both integrin-deficient mice, migration in the WT mice was significantly higher than that observed in the other two genetic models, suggesting impairment in animals with either integrin deficiency. Furthermore, lymphocyte accumulation was dramatically decreased at all time points in the α4−/ − mice (Table 2). Preliminary data in a model of combined, α4- and β2-integrin deficiency, generated in our laboratory, showed that only 3±2% of circulating leukocytes migrated into the peritoneum. Taken together, our results indicate that (1) for migration into peritoneum, neutrophils and monocyte/ macrophages can use α4 and β2 integrins interchangeably, (2) for optimal migration both integrins are needed, and (3) lymphocytes have an absolute requirement of α4 integrins for migration. These data extend previous knowledge on kinetic changes of leukocytes accumulating in the aseptic peritonitis model. Whether different kinetic changes are seen in other inflammatory models using our integrin-deficient animals requires further studies. Accumulation of myeloid cells in peritoneum Neutrophils Monocyte/macrophages 4 hrs 16 hrs 96 hrs 4 hrs 16 hrs 96 hrs WT 5.5±0.5 9.8±7.4 0.2±0.04 3.1±0.6 17.7±3.9 11.0±5.2 β2−/ − 2.5±0.8 10.5±5.3 3.7±1.6 4.7±1.3 12.8±5.2 11.0±4.0 α4−/ − 5.2±2.1 9.8±7.4 0.8±0.4 4.6±1.1 11.0±5.2 12.6±5.1 Lymphocyte accumulation in peritoneum 4 hrs 16 hrs 96 hrs Cell number (x106) in peritoneum at various time points; in bold are the numbers that significantly differ from the WT for each time point (p<0.05) WT 2.7±1.2 2.5±0.3 3.1±0.5 β2−/ − 6.7±1.7 4.7±1.2 3.3±1.0 α4−/ − 0.5±0.1 0.4±0.1 0.6±0.1
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Rabinowich, H., M. Manciulea, R. B. Herberman, and T. L. Whiteside. "Beta1 integrin-mediated activation of focal adhesion kinase and its association with Fyn and Zap-70 in human NK cells." Journal of Immunology 157, no. 9 (November 1, 1996): 3860–68. http://dx.doi.org/10.4049/jimmunol.157.9.3860.
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Abstract 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.
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Palecanda, A., M. J. Briskin, and T. B. Issekutz. "Rat mast cell lines bind to the vascular cell adhesion molecule-1 (VCAM-1) and the mucosal addressin cell adhesion molecule-1 (MAdCAM-1)." Journal of Immunology 158, no. 6 (March 15, 1997): 2904–10. http://dx.doi.org/10.4049/jimmunol.158.6.2904.
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Abstract Mast cells are key mediators of allergy and inflammation. Increased mast cell numbers are observed in the gut during helminth infestation and at many sites of inflammation. To determine whether mast cells express functional receptors for endothelial cell adhesion molecules, we studied the adhesion of two rat mucosal-type mast cell lines RBL-1 and RCMC-1 to transfected mucosal addressin cell adhesion molecule-1 (MAdCAM-1) and VCAM-1. Both mast cell lines expressed high levels of alpha4 integrins on their surface and bound to CHO cells transfected with VCAM-1. Anti-alpha4 mAbs, TA-2 and L25, inhibited the specific adhesion of the mast cells to VCAM-1 by about 92 and 63%, respectively. Both of the mast cell lines also demonstrated an increased adhesion to CHO cells transfected with MAdCAM-1. The adhesion of RBL-1 to MAdCAM-1 was also significantly inhibited by the anti-alpha4 mAbs TA-2, L25, and HP2/1 by 39, 76, and 42%, respectively. In addition, RBL-1 cells adhered to both VCAM-1 and MAdCAM-1 under both static and nonstatic (shear) conditions, and this was also inhibited by the anti-alpha4 mAb TA-2. Thus, mucosal-type mast cell lines express functional alpha4 integrins that can mediate adhesion to VCAM-1 and MAdCAM-1. These results suggest a mechanism for mast cell accumulation at sites of inflammation and in the gut.
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McIntyre, B. W., D. G. Woodside, D. A. Caruso, D. K. Wooten, S. I. Simon, S. Neelamegham, J. K. Revelle, and P. Vanderslice. "Regulation of human T lymphocyte coactivation with an alpha4 integrin antagonist peptide." Journal of Immunology 158, no. 9 (May 1, 1997): 4180–86. http://dx.doi.org/10.4049/jimmunol.158.9.4180.
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Abstract The cyclic hexapeptide CWLDVC (TBC 772) is an antagonist of alpha4 integrins and a potent inhibitor of lymphocyte interactions with fibronectin, vascular cell adhesion molecule-1, and muscosal vascular addressin cell adhesion molecule-1 (MAdCAM-1). As such, peptide TBC 772 effectively inhibits the activation of freshly isolated human T lymphocytes stimulated with purified vascular cell adhesion molecule-1 coimmobilized with anti-CD3 mAb. The influence of peptide binding on distinct sites of the alpha4beta1 complex was determined by flow cytometry and cellular adhesion assays employing a panel of mAbs. Binding of the alpha4-specific mAb L25 and the beta1-specific mAb 33B6 was not altered by the peptide; however, binding of mAb 19H8, which is specific for a combinatorial epitope of alpha4beta1, was dramatically inhibited. Treatment of lymphocytes with the peptide caused an increase in a ligand-induced epitope on beta1 integrin defined by mAb 15/7. In T cell activation studies using coimmobilized anti-CD3 mAb and the anti-integrin mAbs, the peptide had broader inhibitory activity, suppressing costimulation induced by all the integrin mAbs. The peptide was not generally toxic and was integrin selective in its suppressive activity, as coactivation by ligation of CD3 in conjunction with CD28 or CD26 was not affected. These results suggest that the antagonist peptide CWLDVC can effectively neutralize integrin coactivation systems by a mechanism independent of competitive binding.
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Chin, J. E., C. A. Hatfield, G. E. Winterrowd, J. R. Brashler, S. L. Vonderfecht, S. F. Fidler, R. L. Griffin, et al. "Airway recruitment of leukocytes in mice is dependent on alpha4-integrins and vascular cell adhesion molecule-1." American Journal of Physiology-Lung Cellular and Molecular Physiology 272, no. 2 (February 1, 1997): L219—L229. http://dx.doi.org/10.1152/ajplung.1997.272.2.l219.
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The involvement of the alpha4-integrin very late activation antigen 4 and vascular cell adhesion molecule-1 (VCAM-1) in leukocyte trafficking into the airways of ovalbumin (OA)-sensitized and OA-challenged mice was investigated using in vivo administration of anti-alpha4 monoclonal antibodies (mAb) PS/2, R1-2, and M/K-2.7 (MK2), specific for VCAM-1. VCAM-1 was upregulated on endothelial cells in lung tissue after OA inhalation. PS/2, R1-2, or MK2 significantly inhibited the recruitment of eosinophils and lymphocytes into the bronchoalveolar lavage (BAL) fluid and decreased inflammation in the lung tissues. Escalating in vivo doses of PS/2 or MK2 increased circulating levels of rat immunoglobulin G in the plasma. The binding of phycoerytherin-labeled anti-alpha4 mAb to blood T cells from PS/2-treated mice was reduced, implying that alpha4 sites were already occupied. T cells and eosinophils in BAL fluid from mice treated with PS/2 or MK2 were phenotypically different from controls. Selective decreases of alpha4+ T cells in the BAL fluid after PS/2 or MK2 treatment were coupled with changes in CD8+, CD11a, and CD62L expression. The alpha4-integrin and VCAM-1 may have important roles in the antigen-induced recruitment of T cells and eosinophils during OA-induced airway inflammation. The data suggest that these adhesion molecules may be suitable targets for therapeutic intervention in certain conditions of pulmonary inflammation.
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Matsunaga, Takuya, Fumio Fukai, Takuro Kameda, Kotaro Shide, Haruko Shimoda, Eri Torii, Yasuhiro Taniguchi, et al. "Potentiated Activation of VLA-4 and VLA-5 Accelerates Proplatelet-Like Formation In Megakaryocytes." Blood 116, no. 21 (November 19, 2010): 2585. http://dx.doi.org/10.1182/blood.v116.21.2585.2585.
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Abstract Abstract 2585 Several lines of reports have suggested that mature magakaryocytes (MKs) form long cytoplasmic processes containing platelets (PLT) organelles from which PLT break off due to blood flow pressures in bone marrow (BM). These cytoplasmic processes were termed ‘proplatelet'. MKs differentiated from hematopoietic stem cells by in vitro culture also develop similar processes, referred to as ‘proplatelet-like formation (PPF)'. It has been already reported that fibronectin (FN) and phorbol 12-myristate 13-acetate (PMA) are essential for inducing PPF in MKs using CHRF-288 human megakaryoblastic cell line (Jiang F et al. Blood 99, 2002). FN plays important roles in megakaryocytopoiesis through the FN-receptors. The role of adhesive interactions with FN in BM stroma and FN-receptor beta1-integrins has been reported in proliferation, differentiation and maintenance of megakaryocytic lineage cells. However, the substantial role of these FN-receptors and their functional assignment in PPF are not yet fully understood. We first investigated the effects of beta1-integrins on PPF using CHRF-288 cells, which express alpha4beta1-integrin (VLA-4) and alpha5beta1-integrin (VLA-5) as FN-receptors. When the cells were cultured on FN for 3 days, PMA prompted PPF in a dose-dependent manner. While nearly 15% of the cells displayed PPF with PMA (100 ng/mL), no cells cultured with FN alone or PMA alone exhibited PPF. PPF induced by FN plus PMA combination (FN/PMA) was abrogated by addition of anti-alpha4-integrin monoclonal antibodies (mAb) plus anti-alpha5-integrin mAb combination, but not by the addition of anti-alpha4-integrin mAb alone or anti-alpha5-integrin mAb alone. Thus, the adhesive interaction with FN via VLA-4 and VLA-5 were responsible for PPF. We next investigated the effect of TNIIIA2, which enhances the adhesive interaction between FN and beta1-integrins, in PPF induced by FN/PMA. TNIIIA2 (RSTDLPGLKAATHYTITIRGVC) is a 22-mer peptide derived from the 14th FN type III-like (FNIII) repeat in tenascin (TN)-C molecule which we found recently, and it induces the conformational change necessary for functional activation of beta1-integrins (Fukai F et al. J Biol Chem 282, 2007; J Biol Chem 284, 2009). The PPF induced by FN/PMA was highly accelerated when CHRF-288 cells were enforced adhering to FN by treatment with TNIIIA2 (25 microg/mL). More than 45% of the cells displayed PPF with FN/PMA plus TNIIIA2 combination (FN/PMA/TNIIIA2). Blocking experiments using anti-beta1-integrin mAbs indicated that adhesive interaction with FN via VLA-4 and VLA-5 was also responsible for acceleration of PPF induced by FN/PMA/TNIIIA2. On the other hand, control peptide, TNIIIA2mutant (RSTDLPGLKAATHYTATARGVC) did not accelerate PPF induced by PMA/FN. The calculated yield of the cells with PPF induced by FN/PMA/TNIIIA2 was 2.5-fold more than that induced by FN/PMA. We have previously established ‘a three-phase serum-free culture system' to generate large amount of PLT from human cord blood CD34+ cells (Matsunaga T et al. Stem cells 24, 2006). A study on the effect of TNIIIA2 on our ‘three-phase serum-free culture system' is now underway. Finally, we investigated signal transduction pathways responsible for PPF induced by FN/PMA. While FN/PMA induced activation of extracellular signal-regulated protein kinase 1 (ERK1/2), FN alone or PMA alone did not induce ERK1/2 activation. The results was in accordance with the data previously reported by Jiang et at (Blood 99, 2002). TNIIIA2 strongly enhanced activation of ERK1/2 by FN/PMA. However, c-Jun amino-terminal kinase 1 (JNK1), p38 and phosphoinositide-3 kinase (PI3K)/Akt were not stimulated by FN/PMA even in the presence of TNIIIA2. Thus, enhanced activation of ERK1/2 by FN/PMA/TNIIIA2 was responsible for acceleration of PPF by FN/PMA. Disclosures: No relevant conflicts of interest to declare.
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Jacques, T. S., J. B. Relvas, S. Nishimura, R. Pytela, G. M. Edwards, C. H. Streuli, and C. ffrench-Constant. "Neural precursor cell chain migration and division are regulated through different beta1 integrins." Development 125, no. 16 (August 15, 1998): 3167–77. http://dx.doi.org/10.1242/dev.125.16.3167.
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Proliferation and tangential migration of neural precursor cells are essential determinants of CNS development. We have established cell culture models of both these processes using neural precursor cells grown as neurospheres. The pattern of migration that we observe in these cells is homotypic and occurs in the absence of a glial or neuronal scaffold, and is therefore equivalent to that previously described as chain migration. To determine the role of integrins in proliferation and migration, we have analysed the expression pattern of integrins on neurosphere cells and then performed blocking peptide and antibody experiments. Neurosphere cells express five major integrins, alpha5 beta1, alpha 6Abeta1, alphav beta1, alphav beta5 and alpha vbeta8 and, in addition, express low levels of alpha 6Bbeta1. Chain migration is inhibited by blocking the alpha 6beta1 integrin. Proliferation, by contrast, is inhibited by blocking the other beta1 integrins, alphav beta1 and alpha5 beta1. These results show that integrins are important regulators of neural precursor cell behaviour, with distinct beta1 integrins regulating proliferation and migration. They also demonstrate a novel role for the alpha6 beta1 integrin in the cell-cell interactions underlying homotypic chain migration.
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Kumar P, Vinod. "Is the New Approach in Inflammatory Bowel Disease Treat-to-Target?" Internationale Pharmaceutica Sciencia 12, no. 1 (April 5, 2021): 1–14. http://dx.doi.org/10.31531/2231-5896.1000111.
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Inflammatory bowel disease is a chronic inflammatory disease of which the etiology is unknown. Ulcerative colitis and Crohn's disease are the two main entities of inflammatory bowel disease that are challenging clinicians. In addition to tumor necrosis factor blockers, this overview summarizes current and future new drugs, in the treatment of inflammatory bowel disease according to their goals. The infiltration of lymphocytes into the intestinal lining is a target for therapeutic purposes in inflammatory bowel disease. The vascular cell adhesion molecule-1 and the mucosal addressin cell adhesion molecule-1 are a family of integrins for the alpha4 that are specifically expressed in the alimentary canal on vascular endothelial cells. In Crohn's disease, the alpha4beta7 integrin, and its endothelial receptor, the mucosal addressin cell adhesion molecule-1, have proven to be a relevant factor in the development of chronic intestinal inflammation. New biological and chemical drugs are emerging, with additional molecules pending approval.
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Sechler, J. L., A. M. Cumiskey, D. M. Gazzola, and J. E. Schwarzbauer. "A novel RGD-independent fibronectin assembly pathway initiated by alpha4beta1 integrin binding to the alternatively spliced V region." Journal of Cell Science 113, no. 8 (April 15, 2000): 1491–98. http://dx.doi.org/10.1242/jcs.113.8.1491.
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Fibronectin (FN) matrix assembly is a multi-step process that involves binding to integrin receptors, FN-FN interactions and connections to the actin cytoskeleton. Ultimately, FN is converted into stable matrix fibrils that are detergent-insoluble. RGD-binding integrins such as alpha5beta1 play a major role in the assembly of fibrillar FN. Here we show that alpha4beta1 binding to the alternatively spliced V (IIICS) region of FN initiates an alternative assembly pathway. Activation of alpha4beta1 with exogenous agents such as Mn(2+) or a beta1-stimulatory antibody TS2/16 was sufficient to induce initiation of FN fibrillogenesis by Ramos B lymphoma cells and by CHO(B2)alpha4 cells. Using recombinant FNs lacking specific sequences, we show that assembly is independent of the RGD sequence but requires the V25/CS-1 segment. Previously, we have characterized an activated recombinant FN (FN III(1–7)) that rapidly forms detergent-insoluble multimers upon binding to alpha5beta1 integrin. Alpha4beta1 also formed FNdeltaIII(1–7) multimers without the aid of exogenous stimulants, suggesting that an activated form of FN can override the need for activation of the integrin. In contrast to assembly by alpha5beta1, actin filaments remained largely cortical and no change in cell growth rate was observed with alpha4beta1-mediated assembly. These results show that binding sites on FN other than the RGD sequence/synergy site and distant from the cell binding domain can promote FN assembly. Thus, there appear to be multiple, integrin-specific mechanisms for assembly of FN matrix.
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Low, Elizabeth N., Lauren Zagieboylo, Benjamin Martino, and Eric Wilson. "IgA ASC accumulation to the lactating mammary gland is dependent on VCAM-1 and alpha4 integrins." Molecular Immunology 47, no. 7-8 (April 2010): 1608–12. http://dx.doi.org/10.1016/j.molimm.2010.01.015.
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Bayless, K. J., G. A. Meininger, J. M. Scholtz, and G. E. Davis. "Osteopontin is a ligand for the alpha4beta1 integrin." Journal of Cell Science 111, no. 9 (May 1, 1998): 1165–74. http://dx.doi.org/10.1242/jcs.111.9.1165.
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Recent work has shown that osteopontin expression is upregulated at sites of cardiovascular injury. It has been hypothesized that osteopontin provides an adhesive matrix for endothelial and smooth muscle cells during remodeling of the vascular wall following injury. Osteopontin has also been found to be synthesized by monocytes and macrophages within injury sites. Here, we present data showing that osteopontin can promote leukocyte adhesion through the alpha4beta1 integrin. In the presence of physiologic concentrations of Mg2+ and Ca2+, osteopontin purified from bovine milk promoted cell-substrate adhesion of HL-60 and Ramos cells, two model leukocyte cell lines. As with other adhesive ligands, adhesion to osteopontin required leukocyte activation. Under these conditions, no adhesion to control substrates such as bovine serum albumin was observed. Leukocyte adhesion was inhibited by anti-integrin antibodies directed at either the alpha4 or beta1 integrin subunits but not by control antibodies directed to other integrins. Further adhesion experiments revealed that leukocyte binding to osteopontin was completely inhibited by an alpha4beta1-binding peptide containing the leucine-aspartate-valine (LDV) sequence, while a control, non-binding peptide containing leucine-glutamate-valine (LEV) had minimal effects. Affinity chromatography using either surface labeled HL-60 or Ramos cell extracts revealed that the alpha4beta1 integrin specifically bound to osteopontin. Immunoprecipitation of eluted fractions from these columns positively identified the alpha4beta1 integrin. In order to localize potential alpha4beta1-binding sites within osteopontin, the protein was proteolytically cleaved with thrombin. A 30 kDa N-terminal osteopontin fragment purified using fast protein liquid chromatography promoted alpha4beta1 dependent leukocyte adhesion in a manner similar to that of the intact protein. These data collectively demonstrate that the alpha4beta1 integrin is a new adhesion receptor for osteopontin and that an alpha4beta1 binding site exists in the NH2-terminal thrombin fragment of osteopontin.
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Díaz-González, F., J. Forsyth, B. Steiner, and M. H. Ginsberg. "Trans-dominant inhibition of integrin function." Molecular Biology of the Cell 7, no. 12 (December 1996): 1939–51. http://dx.doi.org/10.1091/mbc.7.12.1939.
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Occupancy of integrin adhesion receptors can alter the functions of other integrins and cause partition of the ligand-occupied integrin into focal adhesions. Ligand binding also changes the conformation of integrin extracellular domains. To explore the relationship between ligand-induced conformational change and integrin signaling, we examined the effect of ligands specific for integrin alpha IIb beta 3 on the functions of target integrins alpha 5 beta 1 and alpha 2 beta 1. We report that binding of integrin-specific ligands to a suppressive integrin can inhibit the function of other target integrins (trans-dominant inhibition). Trans-dominant inhibition is due to a blockade of integrin signaling. Furthermore, this inhibition involves both a conformational change in the extracellular domain and the presence of the beta cytoplasmic tail in the suppressive integrin. Similarly, ligand-induced recruitment of alpha IIb beta 3 to focal adhesions also involves a conformational rearrangement of its extracellular domain. These findings imply that the ligand-induced conformational changes can propagate from an integrin's extracellular to its intracellular face. Trans-dominant inhibition by integrin ligands may coordinate integrin signaling and can lead to unexpected biological effects of integrin-specific inhibitors.
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Sastry, S. K., M. Lakonishok, D. A. Thomas, J. Muschler, and A. F. Horwitz. "Integrin alpha subunit ratios, cytoplasmic domains, and growth factor synergy regulate muscle proliferation and differentiation." Journal of Cell Biology 133, no. 1 (April 1, 1996): 169–84. http://dx.doi.org/10.1083/jcb.133.1.169.
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The role of integrins in muscle differentiation was addressed by ectopic expression of integrin alpha subunits in primary quail skeletal muscle, a culture system particularly amenable to efficient transfection and expression of exogenous genes. Ectopic expression of either the human alpha5 subunit or the chicken alpha6 subunit produced contrasting phenotypes. The alpha5-transfected myoblasts remain in the proliferative phase and are differentiation inhibited even in confluent cultures. In contrast, myoblasts that overexpress the alpha6 subunit exhibit inhibited proliferation and substantial differentiation. Antisense suppression of endogenous quail alpha6 expression inhibits myoblast differentiation resulting in sustained proliferation. These effects of ectopic alpha subunit expression are mediated, to a large extent, by the cytoplasmic domains. Ectopic expression of chimeric alpha subunits, alpha5ex/6cyto and alpha6ex/5cyto, produced phenotypes opposite to those observed with ectopic alpha5 or alpha6 expression. Myoblasts that express alpha5ex/6cyto show decreased proliferation while differentiation is partially restored. In contrast, the alpha6ex/5cyto transfectants remain in the proliferative phase unless allowed to become confluent for at least 24 h. Furthermore, expression of human alpha5 subunit cytoplasmic domain truncations, before and after the conserved GFFKR motif, shows that this sequence is important in alpha5 regulation of differentiation. Ectopic alpha5 and alpha6 expression also results in contrasting responses to the mitogenic effects of serum growth factors. Myoblasts expressing the human alpha5 subunit differentiate only in the absence of serum while differentiation of untransfected and alpha6-transfected myoblasts is insensitive to serum concentration. Addition of individual, exogenous growth factors to alpha5-transfected myoblasts results in unique responses that differ from their effects on untransfected cells. Both bFGF or TGFbeta inhibit the serum-free differentiation of alpha5-transfected myoblasts, but differ in that bFGF stimulates proliferation whereas TGF-beta inhibits it. Insulin or TGF-alpha promote proliferation and differentiation of alpha5-transfected myoblasts; however, insulin alters myotube morphology. TGF-alpha or PDGF-BB enhance muscle alpha-actinin organization into myofibrils, which is impaired in differentiated alpha5 cultures. With the exception of TGF-alpha, these growth factor effects are not apparent in untransfected myoblasts. Finally, myoblast survival under serum-free conditions is enhanced by ectopic alpha5 expression only in the presence of bFGF and insulin while TGF-alpha and TGF-beta promote survival of untransfected myoblasts. Our observations demonstrate (1) a specificity for integrin alpha subunits in regulating myoblast proliferation and differentiation; (2) that the ratio of integrin expression can affect the decision to proliferate or differentiate; (3) a role for the alpha subunit cytoplasmic domain in mediating proliferative and differentiative signals; and (4) the regulation of proliferation, differentiation, cytoskeletal assembly, and cell survival depend critically on the expression levels of different integrins and the growth factor environment in which the cells reside.
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De Arcangelis, A., M. Mark, J. Kreidberg, L. Sorokin, and E. Georges-Labouesse. "Synergistic activities of alpha3 and alpha6 integrins are required during apical ectodermal ridge formation and organogenesis in the mouse." Development 126, no. 17 (September 1, 1999): 3957–68. http://dx.doi.org/10.1242/dev.126.17.3957.
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Integrins alpha6beta1 and alpha6beta4 are cell surface receptors for laminins. Integrin alpha6-null mice die at birth with severe skin blistering and defects in the cerebral cortex and in the retina. Integrin alpha3beta1 can associate with laminins and other ligands. Integrin alpha3-null mice also die at birth, with kidney and lung defects at late stages of development, and moderate skin blistering. To investigate possible overlapping functions between alpha3 and alpha6 integrins, we analyzed the phenotype of compound alpha3−/−/alpha6−/− mutant embryos. Double homozygous mutant embryos were growth-retarded and displayed several developmental defects not observed in the single mutant animals. First, limb abnormalities characterized by an absence of digit separation and the fusion of preskeletal elements were observed. Further analyses indicated a defect in the apical ectodermal ridge, an essential limb organizing center. In the double mutant, the ridge appeared flattened, and ridge cells did not show a columnar morphology. A strong reduction in ridge cell proliferation and alterations of the basal lamina underlying the ectoderm were observed. These results suggest that alpha3 and alpha6 integrins are required for the organization or compaction of presumptive apical ectodermal ridge cells into a distinct differentiated structure. Additional defects were present: an absence of neural tube closure, bilateral lung hypoplasia, and several abnormalities in the urogenital tract. Finally, an aggravation of brain and eye lamination defects was observed. The presence of novel phenotypes in double mutant embryos demonstrates the synergism between alpha3 and alpha6 integrins and their essential roles in multiple processes during embryogenesis.
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Piacentino, Gennj, Vittorina Della Bianca, Elena Zenaro, Enrica Caterina Pietronigro, Tommaso Carlucci, Silvia Dusi, and Gabriela Constantin. "P2-126: Blockade of ALPHA4 Integrins Ameliorates Cognitive Dysfunction and Neuropathological Changes in Transgenic Animals with Alzheimer's-Like Disease." Alzheimer's & Dementia 12 (July 2016): P660. http://dx.doi.org/10.1016/j.jalz.2016.06.1496.
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Laidler, P., D. Gil, A. Pituch-Noworolska, D. Ciołczyk, D. Ksiazek, M. Przybyło, and A. Lityńska. "Expression of beta1-integrins and N-cadherin in bladder cancer and melanoma cell lines." Acta Biochimica Polonica 47, no. 4 (December 31, 2000): 1159–70. http://dx.doi.org/10.18388/abp.2000_3968.
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Changes in the expression of integrins and cadherins might contribute to the progression, invasion and metastasis of transitional cell cancer of the bladder and of melanomas. The expression of alpha5 (P < 0.001), alpha2 and beta1 (P < 0.05 - P < 0.001) integrin subunits in melanoma cells from noncutaneous metastatic sites (WM9, A375) were significantly increased as compared to cutaneous primary tumor (WM35) and metastatic (WM239) cell lines. These differences might be ascribed to the invasive character of melanoma cells and their metastasis to the noncutaneous locations. The significantly heterogeneous expression of beta1 integrin subunit in two malignant bladder cancer cell lines (T24 and Hu456) and nonsignificant differences in the expression of alpha2, alpha3, and alpha5 subunits between malignant and non-malignant human bladder cell lines do not allow an unanimous conclusion on the role of these intergrin subunits in the progression of transitional cancer of bladder. The adhesion molecule, expressed in all studied melanoma and bladder cell lines, that reacted with anti-Pan cadherin monoclonal antibodies was identified as N-cadherin except in the HCV29 non-malignant ureter cell line. However, neither this nor any other bladder or melanoma cell line expressed E-cadherin. The obtained results imply that the replacement of E-cadherin by N-cadherin accompanied by a simultaneous increase in expression of alpha2, alpha3 and alpha5 integrin subunits clearly indicates an increase of invasiveness of melanoma and, to a lesser extent, of transitional cell cancer of bladder. High expression of N-cadherin and alpha5 integrin subunit seems to be associated with the most invasive melanoma phenotype.
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Caniggia, I., J. Liu, R. Han, J. Wang, A. K. Tanswell, G. Laurie, and M. Post. "Identification of receptors binding fibronectin and laminin on fetal rat lung cells." American Journal of Physiology-Lung Cellular and Molecular Physiology 270, no. 3 (March 1, 1996): L459—L468. http://dx.doi.org/10.1152/ajplung.1996.270.3.l459.
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Fibronectin and laminin have been implicated in regulating lung morphogenesis. In the present study, the cell surface receptors involved in fetal lung cell binding to laminin and fibronectin were identified. Messages for alpha5- and beta1-integrin subunits were detected in both fetal lung epithelial cells and fibroblasts. The presence of alpha5 beta1 -integrin on both cell types was demonstrated by immunocytochemistry and confirmed by cell adhesion experiments with fibronectin and RGD-containing peptides. Epithelial cells adhered more readily to laminin than fibroblasts. The alpha4 beta1-integrin, and RGD-independent fibronectin receptor, was weakly expressed on either cell type. Both cell types expressed alpha6-integrin subunit mRNA and stained immunopositive for the alpha6-subunit. Although either cell type expressed nonintegrin 67-kDa laminin-elastin receptor mRNA, no positive immunoreactivity for this laminin-elastin binding protein was detected. None of these findings explain the enhanced attachment of distal fetal lung epithelial cells to laminin compared with fibroblasts. Previously, we have reported that epithelial cells were enriched in alpha3-integrin subunit mRNA and protein expression. Herein, we found that epithelial cell attachment to laminin was nearly completely inhibited by alpha3- but only partially by alpha6 -monoclonal antibodies. A peptide near the globular region at the long arm of the laminin A-chain, which contained the IKVAV sequence, and the laminin A-chain amino acid sequence representing the alpha3 beta1 -integrin binding site, inhibited the adherence of epithelial cells to laminin. Fetal lung epithelial cells attached to substrata coated with the alpha3 beta1-integrin binding site peptide and the peptide containing the IKVAV sequence. These data suggest that both fetal lung cell types bind to fibronectin via the fibronectin receptor, alpha5 beta1, and fetal lung epithelial cells interact with laminin via alpha3 beta1 and proteins that recognize the IKVAV-containing sequence on the laminin A-chain.
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Anderson, R., R. Fassler, E. Georges-Labouesse, R. O. Hynes, B. L. Bader, J. A. Kreidberg, K. Schaible, J. Heasman, and C. Wylie. "Mouse primordial germ cells lacking beta1 integrins enter the germline but fail to migrate normally to the gonads." Development 126, no. 8 (April 15, 1999): 1655–64. http://dx.doi.org/10.1242/dev.126.8.1655.
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Primordial germ cells are the founder cells of the gametes. They are set aside at the initial stages of gastrulation in mammals, become embedded in the hind-gut endoderm, then actively migrate to the sites of gonad formation. The molecular basis of this migration is poorly understood. Here we sought to determine if members of the integrin family of cell surface receptors are required for primordial germ cell migration, as integrins have been implicated in the migration of several other motile cell types. We have established a line of mice which express green fluorescent protein in germline cells that has enabled us to efficiently purify primordial germ cells at different stages by flow cytometry. We have catalogued the spectrum of integrin subunit expression by primordial germ cells during and after migration, using flow cytometry, immunocytochemistry and RT-PCR. Through analysis of integrin beta1(−/−)-->wild-type chimeras, we show that embryonic cells lacking beta1 integrins can enter the germline. However, integrin beta1(−/−) primordial germ cells do not colonize the gonad efficiently. Embryos with targeted deletion of integrin subunit alpha3, alpha6, or alphaV show no major defects in primordial germ cell migration. These results demonstrate a role for beta1-containing integrins in the development of the germline, although an equivalent role for * integrin subunit(s) has yet to be established.
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Desban, N., and J. L. Duband. "Avian neural crest cell migration on laminin: interaction of the alpha1beta1 integrin with distinct laminin-1 domains mediates different adhesive responses." Journal of Cell Science 110, no. 21 (November 1, 1997): 2729–44. http://dx.doi.org/10.1242/jcs.110.21.2729.
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In the present study, to further elucidate the molecular events that control neural crest cell migration, we have analyzed in vitro the adhesive and locomotory response of avian trunk neural crest cells to laminin-1 and searched for the integrin receptors involved in this process. Adhesion of crest cells on laminin-1 was comparable to that found on fibronectin or vitronectin. By contrast, migration was significantly greater on laminin-1 than on the other substrate molecules. Interaction of crest cells with laminin-1 involved two major cell-binding domains situated in different portions of the molecule, namely the E1′ and E8 fragments, which elicited different cellular responses. Cells were poorly spread on the E1′ fragment whereas, on E8, they were extremely flattened and cohesive. Either fragment supported cell locomotion, albeit not as efficiently as laminin-1. Immunoprecipitation and immunocytochemistry analyses revealed that crest cells expressed the alpha1beta1, alpha3beta1, alpha6beta1 and alpha vbeta3 integrins, as well as beta8 integrins, as presumptive laminin-1 receptors, but not alpha6beta4 and alpha2beta1. Immunofluorescence labeling of cultured cells showed that the alpha1, alpha v, beta1 and beta3 subunits were diffuse on the cell surface and in focal contacts. In contrast, alpha3 and beta8 were diffuse, while alpha6 was mostly intracytoplasmic and, secondarily, in focal contacts. Inhibition assays of cell adhesion and migration with function-perturbing antibodies demonstrated that alpha1beta1 played a predominant role in both adhesion and migration on laminin-1 and interacted with either binding sites in the E1′ and E8 fragments. Alpha vbeta3 was also implicated in neural crest cell migration. In contrast, alpha3beta1, alpha6beta1 and the beta8 integrins appeared to play only subsidiary roles in cell adhesion and migration. Finally, the ability of neural crest cells to interact with laminin-1 was found to increase with time in culture, possibly in correlation with changes in alpha3 distribution on the cell surface. In conclusion, our study indicates that (1) the preferential migration of neural crest cells along basal laminae can be accounted for by the ability of laminin-1 to promote migration with great efficiency; (2) interaction with laminin-1 involves two major cell binding domains that are both recognized by the alpha1beta1 integrin; (3) alpha1beta1 integrin can elicit different cellular responses depending on the laminin-1 domains with which it interacts; and (4) changes in the repertoire of integrins expressed by neural crest cells are consistent with the modulations of cell-substratum adhesion occurring throughout migration.
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Bradley, L. "Blockade of both L-selectin and alpha4 integrins abrogates naive CD4 cell trafficking and responses in gut-associated lymphoid organs." International Immunology 10, no. 7 (July 1, 1998): 961–68. http://dx.doi.org/10.1093/intimm/10.7.961.
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Simon, E. E., C. H. Liu, M. Das, S. Nigam, T. J. Broekelmann, and J. A. McDonald. "Characterization of integrins in cultured human renal cortical tubule epithelial cells." American Journal of Physiology-Renal Physiology 267, no. 4 (October 1, 1994): F612—F623. http://dx.doi.org/10.1152/ajprenal.1994.267.4.f612.
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We have characterized the integrins present on cultured tubule epithelial cells from human renal cortexes, enriched for proximal cells, using fluorescence microscopy, immunoprecipitation, and cell adhesion assays. By immunofluorescence, the alpha 3-integrin subunit stained most intensely and was present on all cells predominantly at cell-cell contacts. The alpha 6-subunit was present on all cells in a pattern consistent with extracellular matrix contacts. The alpha 5-subunit was present on most cells in a cell-matrix contact pattern; alpha V-subunit was weakly positive and occasionally seen in cell-matrix contacts. The alpha 2-subunit was present on clusters of distal tubule cells, predominantly at cell-cell contacts. Immunoprecipitation revealed the predominant integrin to be alpha 3 beta 1 with some alpha 2 beta 1, presumably contributed by distal cells. The alpha 5 beta 1-, alpha 6 beta 1-, alpha 6 beta 4-, and alpha V beta 3-integrins, as well as trace amounts of alpha 1 beta 1-integrins, were also present. The alpha 4 beta 1-integrin was not detected. Initial attachment to fibronectin was mediated by alpha V beta 3- and alpha 5 beta 1-integrins; initial attachment to laminin was mediated by the alpha 6 beta 1- and alpha 3 beta 1- integrins and, in some preparations, by an unidentified integrin; and initial attachment to collagen type IV was mediated by alpha V beta 3-integrin and an unidentified beta 1-integrin. After extensively immunodepleting membrane extracts with anti-alpha 1, -alpha 2, -alpha 3, -alpha 4, -alpha 5, -alpha 6, and -alpha V antibodies, an anti-beta 1 antibody still precipitated an integrin. Its electrophoretic mobility differs from the laminin-binding alpha 7 beta 1-integrin. Thus we have identified many of the integrins on cortical tubule cells and their role in mediating initial attachment to extracellular matrix. However, the cell adhesion assays and immunoprecipitations suggest the presence of an unidentified beta 1-integrin that may mediate renal tubule cell attachment to laminin and collagen.
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Wu, C., A. J. Fields, B. A. Kapteijn, and J. A. McDonald. "The role of alpha 4 beta 1 integrin in cell motility and fibronectin matrix assembly." Journal of Cell Science 108, no. 2 (February 1, 1995): 821–29. http://dx.doi.org/10.1242/jcs.108.2.821.
Full textAbstract:
The alpha 4 beta 1 integrin has been suggested to play important roles in embryogenesis and pathogenesis of many diseases which involve both cell adhesion and cell migration. Previous studies using anti-alpha 4 beta 1 antibodies and fibronectin (Fn) fragments have suggested that alpha 4 beta 1 integrins may be involved in cell motility on Fn and vascular cell adhesion molecule-1 (VCAM-1). However, the cells used in these studies also express other Fn integrin receptors including alpha 5 beta 1 integrin, which is known to function in cell motility on Fn. To test whether alpha 4 beta 1 integrins mediate cell motility on Fn and VCAM-1 in the absence of alpha 5 beta 1 integrin, we expressed human alpha 4 integrin in a Chinese hamster ovary (CHO) cell line that is deficient in alpha 5 beta 1 integrin (CHO B2). The parental alpha 5 deficient CHO B2 cells were unable to adhere, spread or migrate on Fn, nor could they assemble a fibrillar Fn matrix. Expression of alpha 4 beta 1 integrin in the CHO B2 cells enabled the cells to adhere, spread and migrate on Fn and on VCAM-1 but not to assemble a fibrillar Fn matrix. The cellular processes mediated by the interaction of alpha 4 beta 1 with Fn or VCAM-1 were inhibited by the CS1 peptide derived from the major alpha 4 beta 1 binding site on Fn. These findings demonstrate that alpha 4 beta 1 integrins not only function as cell adhesion receptors but also as cell motility receptors for Fn and VCAM-1 independent of alpha 5 beta 1. Moreover, they reveal important functional differences between Fn binding integrins. The alpha 4-positive, alpha 5-negative CHO cells described in this report will be useful tools in studying the mechanism of molecular signalling during integrin mediated cellular processes.
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Milner, R., and C. Ffrench-Constant. "A developmental analysis of oligodendroglial integrins in primary cells: changes in alpha v-associated beta subunits during differentiation." Development 120, no. 12 (December 1, 1994): 3497–506. http://dx.doi.org/10.1242/dev.120.12.3497.
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We have examined the expression of integrins on primary oligodendroglial cells during the differentiation of the proliferative oligodendrocyte precursor (O-2A progenitor) cell to the postmitotic oligodendrocyte. Cells of the oligodendrocyte lineage expressed a limited repertoire of integrins: alpha 6 beta 1 and alpha v integrins including alpha v beta 1, alpha v beta 3 and alpha v beta 5, as well as a potentially novel integrin alpha v beta 80 kDa. Integrin expression was developmentally regulated; during differentiation alpha v beta 1 was reduced and alpha v beta 5 upregulated. These results suggest that laminin and vitronectin are important extracellular matrix ligands for oligodendrocytes, and provide a rational explanation for previous observations that RGD peptides inhibit the expression of myelin-specific genes. They also suggest a simple model by which switching of integrin beta subunits might regulate differentiation. As chimeric beta 1 integrins with a beta 5 cytoplasmic domain support proliferation less well than normal beta 1 integrins (Pasqualini and Hemler (1994), J. Cell Biol. 125, 447–460) the switch from alpha v beta 1 to alpha v beta 5 might play a key instructive role in the cessation of proliferation and subsequent differentiation.
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Martin, P. T., and J. R. Sanes. "Integrins mediate adhesion to agrin and modulate agrin signaling." Development 124, no. 19 (October 1, 1997): 3909–17. http://dx.doi.org/10.1242/dev.124.19.3909.
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Agrin, a basal lamina-associated proteoglycan, is a crucial nerve-derived organizer of postsynaptic differentiation at the skeletal neuromuscular junction. Because integrins serve as cellular receptors for many basal lamina components, we asked whether agrin interacts with integrins. Agrin-induced aggregation of acetylcholine receptors on cultured myotubes was completely blocked by antibodies to the beta1 integrin subunit and partially blocked by antibodies to the alpha(v) integrin subunit. Agrin-induced clustering was also inhibited by antisense oligonucleotides to alpha(v) and a peptide that blocks the alpha(v) binding site. Non-muscle cells that expressed alpha(v) and beta1 integrin subunits adhered to immobilized agrin, and this adhesion was blocked by anti-alpha(v) and anti-beta1 antibodies. Integrin alpha(v)-negative cells that did not adhere to agrin were rendered adherent by introduction of alpha(v). Together, these results implicate integrins, including alpha(v)beta1, as components or modulators of agrin's signal transduction pathway.
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Yang, J. T., H. Rayburn, and R. O. Hynes. "Cell adhesion events mediated by alpha 4 integrins are essential in placental and cardiac development." Development 121, no. 2 (February 1, 1995): 549–60. http://dx.doi.org/10.1242/dev.121.2.549.
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alpha 4 integrins are cell surface receptors that mediate cell-extracellular matrix (ECM) and cell-cell adhesions by interacting with fibronectin (FN) and vascular cell adhesion molecule 1 (VCAM-1), respectively. We have generated a null mutation in the gene for the alpha 4 integrin subunit. Homozygous null embryos express no alpha 4 integrins and show two unexpected defects, both of which lead to embryonic lethality. The first defect is failure of fusion of the allantois with the chorion during placentation. The second is in the development of the epicardium and coronary vessels leading to cardiac hemorrhage. Both processes clearly involve alpha 4 integrin interactions that were previously unsuspected. alpha 4 integrin and VCAM-1 are expressed at the sites of these interactions. These results raise the possibility of abortifacients targeting alpha 4 integrins, and raise serious questions about potential side effects of drugs currently being designed to block alpha 4 integrin functions in inflammation.
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Dalton, S. L., E. Scharf, R. Briesewitz, E. E. Marcantonio, and R. K. Assoian. "Cell adhesion to extracellular matrix regulates the life cycle of integrins." Molecular Biology of the Cell 6, no. 12 (December 1995): 1781–91. http://dx.doi.org/10.1091/mbc.6.12.1781.
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The expression of alpha 5 beta 1 integrin on the surface of fibroblasts requires adhesion to substratum. We have examined the basis for this adhesion-dependent surface expression by comparing the life cycle of integrins in parallel cultures of adherent and nonadherent cells. Results of biosynthetic labeling experiments in NRK fibroblasts showed that the synthesis and biosynthetic processing of the beta 1 integrin subunit proceed in the absence of cell attachment; however, when examining the behavior of preexisting cell surface integrins, we observed that the alpha beta 1 integrins are internalized and degraded when adhesion to substratum is blocked. A kinetic analysis of integrin internalization in cycloheximide-treated NRK cells showed that each of the fibroblast integrins we examined (in both the beta 1 and beta 3 families) are lost from the cell surface after detachment from substratum. Thus, the default integrin life cycle in fibroblasts involves continuous synthesis, processing, transport to the cell surface, and internalization/degradation. Interestingly, studies with NIH-3T3 cells expressing alpha 1 beta 1 integrin showed that the loss of cell-surface alpha 5 beta 1 integrin is blocked by adhesion of cells to dishes coated with type IV collagen (a ligand for alpha 1 beta 1 integrin) as well as fibronectin. Similarly, adhesion of these cells to dishes coated with type IV collagen stabilizes the surface expression of alpha 5 beta 1 as well as alpha 1 beta 1 integrin. We propose that the adhesion of fibroblasts to extracellular matrix protein alters the integrin life cycle and permits retention of these proteins at the cell surface where they can play important roles in transmitting adhesion-dependent signals.
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Wada, J., A. Kumar, Z. Liu, E. Ruoslahti, L. Reichardt, J. Marvaldi, and Y. S. Kanwar. "Cloning of mouse integrin alphaV cDNA and role of the alphaV-related matrix receptors in metanephric development." Journal of Cell Biology 132, no. 6 (March 15, 1996): 1161–76. http://dx.doi.org/10.1083/jcb.132.6.1161.
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Metanephrogenesis has been a long-standing model to study cell-matrix interactions. A number of adhesion molecules, including matrix receptors (i.e., integrins), are believed to be involved in such interactions. The integrins contain alpha and beta s ubunits and are present in various tissues in different heterodimeric forms. In this study, one of the members of the integrin superfamily, alphaV, was characterized, and its relevance in murine nephrogenesis was investigated. Mouse embryonic renal cDNA libraries were prepared and screened for alphaV, and multiple clones were isolated and sequenced. The deduced amino acid sequence of the alpha-v cDNA clones and hydropathic analysis revealed that it has a typical signal sequence and extracellular, transmembrane, and cytoplasmic domains, with multiple Ca2+ binding sites. No A(U)nA mRNA instability motifs were present. Conformational analysis revealed no rigid long-range-ordered structure in murine alphaV. The alphaV was expressed in the embryonic kidney at day 13 of the gestation, with a transcript size of approximately 7 kb. Its expression increased progressively during the later gestational stages and in the neonatal period. It was distributed in the epithelial elements of developing nephrons and was absent in the uninduced mesenchyme. In mature metanephroi, the expression was relatively high in the glomeruli and blood vessels, as compared to the tubules. Various heterodimeric associations of alphaV, i.e., with beta1, beta3, beta5, and beta6, were observed in metanephric tissues. Inclusion of alphaV-antisense-oligodeoxynucleotide or -antibody in metanephric culture induced dysmorphogenesis of the kidney with reduced population of the nephrons, disorganization of the ureteric bud branches, and reduction of mRNA and protein expressions of alphaV. The expressions of integrin beta3, beta5, and beta6 were unaltered. These findings suggest that the integrin alphaV is developmentally regulated, has a distinct spatio-temporal expression, and is relevant in the mammalian organogenesis.
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Wang, Z., J. M. Symons, S. L. Goldstein, A. McDonald, J. H. Miner, and J. A. Kreidberg. "(Alpha)3(beta)1 integrin regulates epithelial cytoskeletal organization." Journal of Cell Science 112, no. 17 (September 1, 1999): 2925–35. http://dx.doi.org/10.1242/jcs.112.17.2925.
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Epithelial cell morphology and cytoskeletal organization are determined by interactions, with both adjacent cells and the extracellular matrix, which are mediated by integrins and cadherins. Little is known, however, of the relative contributions of integrins and cadherins to maintaining the sub-cortical cytoskeleton characteristic of epithelial cells. Since most studies that utilize integrin-blocking antibodies result in a loss of both cell-cell adhesion and sub-cortical cytoskeletal organization, it has been difficult to distinguish whether integrins and cadherins both mediate cytoskeletal assembly in epithelial cells. Therefore, cells derived from kidney collecting ducts of (alpha)3(beta)1 integrin-deficient mice were used to examine the role of integrins in epithelial cell morphology and cytoskeletal organization. In primary cell culture, (alpha)3(beta)1 integrin-deficient kidney collecting duct cells maintain cadherin-mediated cell-cell adhesions but fail to form the sub-cortical cytoskeleton that is characteristic of epithelial cells, and instead assemble actin stress fibers. Moreover, the cell-cell junctions in mutant cells were irregular, rather than being uniformly oriented perpendicular to the culture substrate. These results demonstrated that integrins have an primary and essential function in establishing and maintaining the sub-cortical cytoskeleton that is characteristic of epithelial cells. To further study the role of (alpha)3(beta)1 integrin in establishing and maintaining cytoskeletal organization in tubular epithelial cells, we derived immortalized cell lines from wild-type and (alpha)3(beta)1 integrin-deficient kidney collecting ducts that duplicated the cytoskeletal and cadherin organization observed in primary cells. E-cadherin and (alpha)- and (beta)-catenin were complexed together in equal amounts in membranes of wild-type and (alpha)3(beta)1 integrin-deficient cells. However, association of the cadherin:catenin complex with (alpha)-actinin was greatly decreased in mutant cells, indicating that integrin-mediated assembly of the sub-cortical cytoskeleton is essential for subsequent association of the cytoskeleton with the cadherin:catenin complex. These results present direct evidence for integrin:cadherin cross-regulation in which cadherin function is dependent on the presence of an integrin.
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Testaz, S., M. Delannet, and J. Duband. "Adhesion and migration of avian neural crest cells on fibronectin require the cooperating activities of multiple integrins of the (beta)1 and (beta)3 families." Journal of Cell Science 112, no. 24 (December 15, 1999): 4715–28. http://dx.doi.org/10.1242/jcs.112.24.4715.
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Based on genetic, functional and histological studies, the extracellular matrix molecule fibronectin has been proposed to play a key role in the migration of neural crest cells in the vertebrate embryo. In the present study, we have analyzed in vitro the repertoire and function of integrin receptors involved in the adhesive and locomotory responses of avian truncal neural crest cells to fibronectin. Immunoprecipitation experiments showed that neural crest cells express multiple integrins, namely (alpha)3(beta)1, (alpha)4(beta)1, (alpha)5(beta)1, (alpha)8(beta)1, (alpha)v(beta)1, (alpha)v(beta)3 and a (beta)8 integrin, as potential fibronectin receptors, and flow cytometry analyses revealed no major heterogeneity among the cell population for expression of integrin subunits. In addition, the integrin repertoire expressed by neural crest cells was found not to change dramatically during migration. At the cellular level, only (alpha)v(beta)1 and (alpha)v(beta)3 were concentrated in focal adhesion sites in connection with the actin microfilaments, whereas the other integrins were predominantly diffuse over the cell surface. In inhibition assays with function-perturbing antibodies, it appeared that complete abolition of cell spreading and migration could be achieved only by blocking multiple integrins of the (beta)1 and (beta)3 families, suggesting possible functional compensations between different integrins. In addition, these studies provided evidence for functional partitioning of integrins in cell adhesion and migration. While spreading was essentially mediated by (alpha)v(beta)1 and (alpha)8(beta)1, migration involved primarily (alpha)4(beta)1, (alpha)v(beta)3 and (alpha)8(beta)1 and, more indirectly, (alpha)3(beta)1. (alpha)5(beta)1 and the (beta)8 integrin were not found to play any major role in either adhesion or migration. Finally, consistent with the results of inhibition experiments, recruitment of (alpha)4(beta)1 and (alpha)v(beta)3, individually or in combination using antibodies or recombinant VCAM-1 and PECAM-1 molecules as a substratum, was required for migration but was not sufficient to produce migration of the cell population as efficiently as with fibronectin. In conclusion, our study indicates that neural crest cells express a multiplicity of fibronectin-binding integrins and suggests that dispersion of the cell population requires cooperation between distinct integrins regulating different events of cell adhesion, locomotion and, possibly, proliferation and survival.
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Kikkawa, Y., N. Sanzen, H. Fujiwara, A. Sonnenberg, and K. Sekiguchi. "Integrin binding specificity of laminin-10/11: laminin-10/11 are recognized by alpha 3 beta 1, alpha 6 beta 1 and alpha 6 beta 4 integrins." Journal of Cell Science 113, no. 5 (March 1, 2000): 869–76. http://dx.doi.org/10.1242/jcs.113.5.869.
Full textAbstract:
Laminin-10/11, the laminin isoforms containing the alpha 5 chain, are major components of basement membranes of many fetal and adult tissues. Laminin-10/11 purified from the conditioned medium of human lung carcinoma cells were potent in mediating adhesion of the carcinoma cells in an integrin alpha 3 beta 1-dependent manner. To further define the type(s) of integrins involved in cell adhesion to laminin-10/11, we examined the effects of a panel of function-blocking anti-integrin antibodies on the adhesion of different cell types to laminin-10/11. Although anti-integrin beta 1 antibody inhibited the adhesion of all cell types tested, anti-alpha 3 antibody inhibited the adhesion of carcinoma and glioma cells but not fibroblastic cells. Adhesion of fibroblastic cells was inhibited, however, by a combination of anti-alpha 3 and anti-alpha 6 antibodies, suggesting that both alpha 3 beta 1 and alpha 6 beta 1 integrins function as laminin-10/11 receptors in these cells. To explore this possibility, we examined the adhesion of K562 leukemic cells transfected with integrin alpha 3 or alpha 6 subunit to laminin-10/11 or other laminin isoforms. Laminin-10/11 were potent adhesive ligands for both the alpha 3 beta 11 and alpha 6 beta 1 transfectants, whereas laminin-5 was the preferred ligand for the alpha 3 beta 1 transfectants. Upon stimulation with the activating anti-integrin beta 1 antibody, both transfectants became more adherent to the substratum regardless of the type of laminins coated, although their preference for laminin isoforms remained unaltered. K562 cells transfected with alpha 6 and beta 4 subunits were also capable of adhering to laminin-10/11, indicating that integrin alpha 6 beta 4 is another receptor for laminin-10/11. Even with lung carcinoma cells, the alpha 6-containing integrins partly contributed to adhesion to laminin-10/11 at higher coating concentrations, although non-integrin receptor(s) might also be involved under such conditions. These results indicated that laminin-10/11 are potent and versatile adhesive ligands in basement membranes capable of binding to both alpha 3 beta 1 and alpha 6 beta 1 integrins with high avidity and also to alpha 6 beta 4 integrin.
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Hara, M., M. Yaar, A. Tang, M. S. Eller, W. Reenstra, and B. A. Gilchrest. "Role of integrins in melanocyte attachment and dendricity." Journal of Cell Science 107, no. 10 (October 1, 1994): 2739–48. http://dx.doi.org/10.1242/jcs.107.10.2739.
Full textAbstract:
Integrins are a family of proteins known to mediate attachment of cells to extracellular matrix materials. The substratum specificity and cation dependence of specific integrin heterodimers have been extensively characterized, and to a lesser degree specialized roles in cell attachment versus dendricity have been defined in some cell types. In the past decade, melanocyte attachment rate and morphology have been found to have strong substratum dependence, suggesting a major role for integrins in these processes. In order to investigate this aspect of pigment cell biology, human newborn melanocytes were subjected to flow cytometry analysis and plated on a variety of substrata under conditions known to promote or block the binding of specific integrin pairs. Melanocyte attachment to laminin and type IV collagen was promoted by Mg2+ and Mn2+ but not by Ca2+, in the range of concentrations examined. However, dendrite outgrowth from melanocytes already attached on laminin or type IV collagen was promoted by Ca2+ to a far greater degree than by Mg2+, and Mn2+ had no effect on dendrite outgrowth. Flow cytometry analysis revealed that melanocytes expressed beta 1, alpha 2, alpha 3, alpha 5, alpha 6 and alpha v integrin subunits as well as the alpha v beta 3 heterodimer. The influence of substratum on the profile of integrin expression was minimal, but alpha 6 and beta 1 integrins were observed by confocal microscopy to be expressed over the entire cell surface, while alpha 2, alpha 5 and alpha v beta 3 integrins localized along dendritic processes or at their tips. In accordance with the implications of these distribution patterns, anti-beta 1 and anti-alpha 6 integrin monoclonal antibodies blocked melanocyte attachment to laminin, while anti-alpha 2, anti-alpha 5 and anti-alpha v beta 3 inhibited dendrite outgrowth but did not block substratum attachment on either laminin or type IV collagen. On the basis of these data and the known characteristics of integrin molecules, we conclude that melanocyte attachment to laminin is mediated primarily by alpha 6 beta 1 integrin in a Ca(2+)-independent, Mg(2+)- and/or Mn(2+)-dependent manner, while dendrite outgrowth on laminin and type IV collagen requires extracellular Ca2+ and is mediated by alpha v beta 3 as well as alpha 2 and alpha 5 integrins.
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Adams, J. C., and F. M. Watt. "Expression of beta 1, beta 3, beta 4, and beta 5 integrins by human epidermal keratinocytes and non-differentiating keratinocytes." Journal of Cell Biology 115, no. 3 (November 1, 1991): 829–41. http://dx.doi.org/10.1083/jcb.115.3.829.
Full textAbstract:
We have compared the adhesive properties and integrin expression profiles of cultured human epidermal keratinocytes and a strain of nondifferentiating keratinocytes (ndk). Both cell types adhered to fibronectin, laminin, and collagen types I and IV, but ndk adhered more rapidly and at lower coating concentrations of the proteins. Antibody blocking experiments showed that adhesion of both cell types to fibronectin was mediated by the alpha 5 beta 1 integrin and to laminin by alpha 3 beta 1 in synergy with alpha 2 beta 1. Keratinocytes adhered to collagen with alpha 2 beta 1, but an antibody to alpha 2 did not inhibit adhesion of ndk to collagen. Both cell types adhered to vitronectin by alpha v-containing integrins. Immunoprecipitation of surface-iodinated and metabolically labeled cells showed that in addition to alpha 2 beta 1, alpha 3 beta 1, and alpha 5 beta 1, both keratinocytes and ndk expressed alpha 6 beta 4 and alpha v beta 5. ndk expressed all these integrins at higher levels than normal keratinocytes. ndk, but not normal keratinocytes, expressed alpha v beta 1 and alpha v beta 3; they also expressed alpha 1 beta 1, an integrin that was not consistently detected on normal keratinocytes. Immunofluorescence experiments showed that in stratified cultures of normal keratinocytes integrin expression was confined to cells in the basal layer; terminally differentiating cells were unstained. In contrast, all cells in the ndk population were integrin positive. Our observations showed that the adhesive properties of ndk differ from normal keratinocytes and reflect differences in the type of integrins expressed, the level of expression and the distribution of integrins on the cell surface. ndk thus have a number of characteristics that distinguish them from normal basal keratinocytes.
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Schoenenberger, C. A., A. Zuk, G. M. Zinkl, D. Kendall, and K. S. Matlin. "Integrin expression and localization in normal MDCK cells and transformed MDCK cells lacking apical polarity." Journal of Cell Science 107, no. 2 (February 1, 1994): 527–41. http://dx.doi.org/10.1242/jcs.107.2.527.
Full textAbstract:
Epithelial cells polarize in response to contacts with the extracellular matrix and with neighboring cells. Interactions of cells with the extracellular matrix are mediated mainly by the integrin family of receptors. To begin to understand the role of integrins in polarization, we have investigated the expression and localization of three integrin families in the polarized Madin-Darby canine kidney (MDCK) epithelial cell line and in transformed MDCK cells lacking apical polarity. We find that MDCK cells express several beta 1 integrins, including alpha 2 beta 1, alpha 3 beta 1, and an unidentified integrin designated alpha × beta 1. The beta 1 integrins are the major receptors for collagens I and IV and laminin in MDCK cells, since a blocking anti-beta 1 antibody almost totally abolishes adhesion to these proteins. They also express a vitronectin receptor tentatively identified as alpha v beta 3, and the epithelial-specific integrin alpha 6 beta 4. The latter is not a laminin receptor in MDCK cells because a function blocking anti-alpha 6 antibody has no effect on cell adhesion to laminin. All three integrin families are expressed exclusively on both the basal and lateral surfaces, as determined by immunofluorescence microscopy and surface biotinylation. Transformed MDCK cells express beta 1 integrins as well as alpha v beta 3 and alpha 6 beta 4, but show alterations in the beta 1 family. Expression of alpha × is lacking, and the relative amount of the beta 1 subunit is diminished, resulting in the accumulation of Endo-H-sensitive alpha 3. In addition, surface biotinylation and immunofluorescence indicate that significant amounts of both alpha 2 beta 1 and alpha 3 beta 1 appear on not only the basolateral but also the apical plasma membrane. These results indicate that integrins are the major receptors for the extracellular matrix in MDCK cells, and that they may affect epithelial cell polarization by mediating not only cell-substratum but also cell-cell contacts.
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Leivo, I., T. Tani, L. Laitinen, R. Bruns, E. Kivilaakso, V. P. Lehto, R. E. Burgeson, and I. Virtanen. "Anchoring complex components laminin-5 and type VII collagen in intestine: association with migrating and differentiating enterocytes." Journal of Histochemistry & Cytochemistry 44, no. 11 (November 1996): 1267–77. http://dx.doi.org/10.1177/44.11.8918902.
Full textAbstract:
Anchoring complex component laminin-5 and its subunits laminin (Ln)-alpha3 and Ln-beta3 chains, Type VII collagen, and integrin chains alpha3, alpha6, and beta4 were studied in developing and adult human intestine and compared with findings on Ln-alpha1 and Ln-alpha2 chains. In adult human duodenum, jejunum, and ileum, Ln-5 detected with a polyclonal antiserum and Ln-alpha3 and Ln-beta3 chains, detected with monoclonal antibodies (MAbs), were restricted to the epithelial basement membranes (BMs) of villi, whereas Ln-alpha2 chain was seen only focally in crypt bottoms. In double labeling experiments, the stretch of crypt BM corresponding to the proliferative cell compartment was found to be devoid of both Ln-alpha3 and Ln-alpha2 chains. Double labeling for Ln-5 and proliferating cell nuclear antigen also showed an abrupt onset of Ln-5 expression exactly at the upper edge of the proliferative cell compartment. Type VII collagen was negligible in duodenum and showed a rising duodenal-ileal gradient localizing to villar BMs. Double labeling for Ln-5 and Type VII collagen, however, indicated only partial co-distribution in the intestine. Electron microscopy of ileum revealed both anchoring filaments and anchoring fibrils but no hemidesmosomal plaques. Our results demonstrate the expression of Ln-5 in BMs outside of stratified epithelia and indicate that Ln-5 in the intestine is associated with the compartment of migrating and differentiating enterocytes. Absence of hemidesmosomes and the presence of other anchoring complex components, such as Ln-5, Type VII collagen, and integrin chains alpha3, alpha6, and beta4, suggests unique properties for epithelial cell attachment in the intestine.
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Caniggia, I., R. Han, J. Liu, J. Wang, A. K. Tanswell, and M. Post. "Differential expression of collagen-binding receptors in fetal rat lung cells." American Journal of Physiology-Lung Cellular and Molecular Physiology 268, no. 1 (January 1, 1995): L136—L143. http://dx.doi.org/10.1152/ajplung.1995.268.1.l136.
Full textAbstract:
Interactions of cells with molecules of the extracellular matrix (ECM) are mediated via specific cell surface receptors. Because collagen is an important ECM component in the developing lung, we investigated the expression of collagen receptors by distal fetal lung epithelial cells and fibroblasts. Cell attachment experiments revealed that fibroblasts but not epithelial cells adhered to various types of collagen. With the use of subunit-specific antibodies, we demonstrated the presence of the collagen integrins alpha 1 beta 1, alpha 2 beta 1, and alpha 3 beta 1 at the fibroblast surface but only the integrin alpha 3 beta 1 on epithelial cells. Affinity chromatography on collagen-Sepharose identified only alpha 1 beta 1 and alpha 2 beta 1 as collagen-binding integrins in extracts of 125I surface-labeled fibroblasts. No collagen-binding receptors were detected in extracts of surface-iodinated epithelial cells. Message for alpha 1- and alpha 2-integrin was readily demonstrated for fibroblasts, but both mRNAs were hardly detectable in epithelial cells. In contrast, epithelial cells expressed significantly greater alpha 3 mRNA levels than fibroblasts. These data demonstrate that alpha 1 beta 1- and alpha 2 beta 1-integrins function as collagen-binding receptors in fetal lung fibroblasts. Distal fetal lung epithelial cells do not express the alpha 1 beta 1- and alpha 2 beta 1-integrins and do not adhere to collagen. The alpha 3 beta 1-integrin, which is expressed by both cell types, does not function as a collagen receptor.
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Watt, F. M., M. D. Kubler, N. A. Hotchin, L. J. Nicholson, and J. C. Adams. "Regulation of keratinocyte terminal differentiation by integrin-extracellular matrix interactions." Journal of Cell Science 106, no. 1 (September 1, 1993): 175–82. http://dx.doi.org/10.1242/jcs.106.1.175.
Full textAbstract:
Suspension-induced terminal differentiation of human epidermal keratinocytes can be inhibited by fibronectin through binding to the alpha 5 beta 1 integrin. We have investigated the effect of fibronectin on expression of integrins and proteins of the actin cytoskeleton and have explored the nature of the differentiation stimulus by testing different combinations of anti-integrin monoclonal antibodies or extracellular matrix proteins in the suspension assay. Fibronectin prolonged cell surface expression of beta 1 integrins but did not overcome the inhibition of intracellular transport of integrins that occurs when keratinocytes are placed in suspension. Fibronectin did not prevent the suspension-induced decline in the level of mRNAs encoding the beta 1 integrin subunit, actin, filamin and alpha-actinin; furthermore, the inhibition of terminal differentiation did not depend on the state of assembly of microfilaments or microtubules. Terminal differentiation could be partially inhibited by an adhesion-blocking monoclonal antibody to the beta 1 integrin subunit or by a combination of adhesion blocking antibodies recognising the alpha subunits that associate with beta 1 (alpha 2, alpha 3 and alpha 5). Although laminin and type IV collagen do not inhibit terminal differentiation individually, they were inhibitory when added to cells in combination with a low concentration of fibronectin. We conclude that the proportion of keratinocyte beta 1 integrins occupied by ligand can regulate the initiation of terminal differentiation independently of the state of assembly of the actin cytoskeleton.
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Wu, C., A. E. Chung, and J. A. McDonald. "A novel role for alpha 3 beta 1 integrins in extracellular matrix assembly." Journal of Cell Science 108, no. 6 (June 1, 1995): 2511–23. http://dx.doi.org/10.1242/jcs.108.6.2511.
Full textAbstract:
To study the biological role of alpha 3 beta 1 integrins in cell adhesion, migration, and in the deposition of extracellular matrix, we stably expressed the human alpha 3 integrin subunit in the alpha 4, alpha 5 integrin deficient CHO cell line B2. The expression of alpha 3 beta 1 integrins enhanced cell adhesion on entactin (also known as nidogen), but not on fibronectin. Using recombinant GST-fusion proteins that span the entire length of the entactin molecule, we located cell adhesive activity to the G2 domain of entactin. These results suggest that the alpha 3 beta 1 integrin functions as an adhesion receptor interacting with the G2 domain of entactin. On the other hand, the expression of alpha 3 beta 1 integrins did not confer the ability to migrate on entactin. Strikingly, the expression of alpha 3 beta 1 dramatically increased the deposition of entactin and fibronectin into the pericellular matrix. This was accompanied by increased binding activity of the 29 kDa amino-terminal domain of fibronectin. Thus, similar to alpha 5 beta 1 integrins, alpha 3 beta 1 integrins can play an important role in modulating the assembly of pericellular matrices. However, unlike fibronectin deposition supported by alpha 5 beta 1, alpha 3 beta 1 supported fibronectin deposition into pericellular matrix was not inhibited by antibodies binding to the RGD containing cell adhesion domain of fibronectin, demonstrating that the two processes are mechanistically distinct. The role of alpha 3 beta 1 in pericellular matrix assembly potentially implicates this receptor in the assembly and/or recognition of entactin-containing pericellular matrices, an observation consistent with its apparent role in the renal glomerulus of the mammalian kidney.
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Martel, V., L. Vignoud, S. Dupe, P. Frachet, M. R. Block, and C. Albiges-Rizo. "Talin controls the exit of the integrin alpha 5 beta 1 from an early compartment of the secretory pathway." Journal of Cell Science 113, no. 11 (June 1, 2000): 1951–61. http://dx.doi.org/10.1242/jcs.113.11.1951.
Full textAbstract:
Talin is a major cytosolic protein that links the intracellular domains of beta1 and beta3 integrins to the cytoskeleton. It is required for focal adhesion assembly. However, its downregulation not only slows down cell spreading and organization of focal adhesions but also impairs the maturation of some beta1 integrins, including the fibronectin receptor alpha5beta1. To investigate this, we characterized the beta1 integrin synthesized in cells expressing talin anti-sense RNA (AT22 cells). We identified a large intracellular pool of beta1 integrins that is abnormally accumulated in an earlier compartment of the secretory pathway. In this report, we show that in talin-deficient AT22 cells, the aberrant glycosylation of integrin receptors is accompanied by a delay in the export of the integrin alpha5beta1. In normal cells, talin was found associated with beta1 integrins in an enriched membrane fraction containing Golgi and endoplasmic reticulum. Finally, microinjection of anti-talin antibodies resulted in accumulation of the integrins within the cells. These data strongly suggest that talin plays a specific role in the export of newly synthesized integrins. We propose that talin binding to the integrin may disclose a diphenylalanine export signal, which is present in the membrane-proximal GFFKR motif conserved in all integrin alpha chains.
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Tomaselli, K. J., C. H. Damsky, and L. F. Reichardt. "Purification and characterization of mammalian integrins expressed by a rat neuronal cell line (PC12): evidence that they function as alpha/beta heterodimeric receptors for laminin and type IV collagen." Journal of Cell Biology 107, no. 3 (September 1, 1988): 1241–52. http://dx.doi.org/10.1083/jcb.107.3.1241.
Full textAbstract:
Cells of the rat neuronal line, PC12, adhere well to substrates coated with laminin and type IV collagen, but attach poorly to fibronectin. Adhesion and neurite extension in response to these extracellular matrix proteins are inhibited by Fab fragments of an antiserum (anti-ECMR) that recognizes PC12 cell surface integrin subunits of Mr 120,000, 140,000, and 180,000 (Tomaselli, K. J., C. H. Damsky, and L. F. Reichardt. 1987. J. Cell Biol. 105:2347-2358). Here we extend our study of integrin structure and function in PC12 cells using integrin subunit-specific antibodies prepared against synthetic peptides corresponding to the cytoplasmic domains of the human integrin beta 1 and the fibronectin receptor alpha (alpha FN) subunits. Anti-integrin beta 1 immunoprecipitated a 120-kD beta 1 subunit and two noncovalently associated integrin alpha subunits of 140 and 180 kD from detergent extracts of surface-labeled PC12 cells. Immunodepletion studies using anti-integrin beta 1 demonstrated that these two putative alpha/beta heterodimers are identical to those recognized by the adhesion-perturbing ECMR antiserum. Anti-alpha FN immunoprecipitated fibronectin receptor heterodimers in human and rat fibroblastic cells, but not in PC12 cells. Thus, low levels of expression of the integrin alpha FN subunit can explain the poor attachment of PC12 cells to FN. The PC12 cell integrins were purified using a combination of lectin and ECMR antibody affinity chromatography. The purified integrins: (a) completely neutralize the ability of the anti-ECMR serum to inhibit PC12 cell adhesion to laminin and collagen IV; (b) have hydrodynamic properties that are very similar to those of previously characterized integrin alpha/beta heterodimeric receptors for ECM proteins; and (c) can be incorporated into phosphatidylcholine vesicles that then bind specifically to substrates coated with laminin or collagen IV but not fibronectin. Thus, the ligand-binding specificity of the liposomes containing the purified PC12 integrins closely parallels the substrate-binding preference of intact PC12 cells. These results demonstrate that mammalian integrins purified from a neuronal cell line can, when incorporated into lipid vesicles, function as receptors for laminin and type IV collagen.
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Ylänne, J., Y. Chen, TE O'Toole, JC Loftus, Y. Takada, and MH Ginsberg. "Distinct functions of integrin alpha and beta subunit cytoplasmic domains in cell spreading and formation of focal adhesions." Journal of Cell Biology 122, no. 1 (July 1, 1993): 223–33. http://dx.doi.org/10.1083/jcb.122.1.223.
Full textAbstract:
Integrin-mediated cell adhesion often results in cell spreading and the formation of focal adhesions. We exploited the capacity of recombinant human alpha IIb beta 3 integrin to endow heterologous cells with the ability to adhere and spread on fibrinogen to study the role of integrin cytoplasmic domains in initiation of cell spreading and focal adhesions. The same constructs were also used to analyze the role of the cytoplasmic domains in maintenance of the fidelity of the integrin repertoire at focal adhesions. Truncation mutants of the cytoplasmic domain of alpha IIb did not interfere with the ability of alpha IIb beta 3 to initiate cell spreading and form focal adhesions. Nevertheless, deletion of the alpha IIb cytoplasmic domain allowed indiscriminate recruitment of alpha IIb beta 3 to focal adhesions formed by other integrins. Truncation of the beta 3 subunit cytoplasmic domain abolished cell spreading mediated by alpha IIb beta 3 and also abrogated recruitment of alpha IIb beta 3 to focal adhesions. This truncation also dramatically impaired the ability of alpha IIb beta 3 to mediate the contraction of fibrin gels. In contrast, the beta 3 subunit cytoplasmic truncation did not reduce the fibrinogen binding affinity of alpha IIb beta 3. Thus, the integrin beta 3 subunit cytoplasmic domain is necessary and sufficient for initiation of cell spreading and focal adhesion formation. Further, the beta 3 cytoplasmic domain is required for the transmission of intracellular contractile forces to fibrin gels. The alpha subunit cytoplasmic domain maintains the fidelity of recruitment of the integrins to focal adhesions and thus regulates their repertoire of integrins.
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Bhatia, R., JB McCarthy, and CM Verfaillie. "Interferon-alpha restores normal beta 1 integrin-mediated inhibition of hematopoietic progenitor proliferation by the marrow microenvironment in chronic myelogenous leukemia." Blood 87, no. 9 (May 1, 1996): 3883–91. http://dx.doi.org/10.1182/blood.v87.9.3883.bloodjournal8793883.
Full textAbstract:
Chronic myelogenous leukemia (CML) progenitors show decreased adhesion to stroma and fibronectin (FN) through beta 1 integrin receptors. We have previously shown that interferon-alpha (IFN-alpha) restores beta 1 integrin-mediated adhesion of CML progenitors to stroma. Because beta1 integrins transmit proliferation inhibitory signals from the microenvironment to normal hematopoietic progenitors, we hypothesized that decreased integrin-mediated adhesion of CML progenitors contributes to their continuous proliferation when in contact with stroma and that IFN-alpha treatment, by restoring integrin-mediated adhesion, also restores integrin-mediated microenvironmental inhibition of CML progenitor proliferation. We show here that, in contrast to normal colony-forming cells (CFC), the percentage of malignant CML CFC in S-phase was not significantly reduced following coculture with stromal layers. However, IFN-alpha treatment resulted in a significant reduction in the proliferation of CML CFC on coculture with stroma. This effect was not because of a direct antiproliferative effect of IFN- alpha on CML CFC because the proliferation of IFN-alpha treated CML CFC kept in suspension culture was not reduced. We examined the role of restored signaling through beta 1 integrin receptors in IFN-alpha induced inhibition of CML progenitors in two sets of experiment. In the first set of experiments, we demonstrated that proliferation of IFN- alpha-treated CML CFC, but not untreated CML CFC, was significantly reduced following coculture with 33/66-kD and 75-kD FN fragments, recognized by alpha 4 beta 1 and alpha 5 beta 1 integrins respectively. In a second set of experiments, we demonstrate that direct stimulation of integrin receptors by crosslinking with blocking antibodies to alpha 4, alpha 5, and beta 1 integrins and secondary goat antimouse antibodies resulted in significant reduction in proliferation of normal and IFN-alpha treated CML progenitors but not untreated CML CFC. These studies indicate that CML hematopoietic progenitors are unresponsive to beta 1-integrin mediated proliferation inhibition and that IFN-alpha not only restores beta 1 integrin-mediated adhesion but also beta1- mediated microenvironmental inhibition of CML progenitor proliferation. These observations may explain, at least in part, the therapeutic efficacy of IFN-alpha in CML.
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Wang, RN, and L. Rosenberg. "Maintenance of beta-cell function and survival following islet isolation requires re-establishment of the islet-matrix relationship." Journal of Endocrinology 163, no. 2 (November 1, 1999): 181–90. http://dx.doi.org/10.1677/joe.0.1630181.
Full textAbstract:
Islet transplantation is associated with a high rate of early graft failure, a problem that remains poorly understood. It is probable that the destruction of the islet microenvironment and loss of tropic support that occur during isolation lead to compromised survival. The purpose of this study was to determine the role of matrix-integrin interactions on beta-cell survival and function following islet isolation. Canine islets were obtained by conventional methods. Immediately after isolation, the peri-insular basement membrane (BM) was absent. The ability of islets maintained in suspension culture to attach to a collagen matrix declined progressively over 6 days. Attachment could be blocked by an arginine-glycine-aspartate (RGD) motif-presenting synthetic peptide, thereby implicating an integrin-mediated process. Characterization of cell surface integrins by immunocytochemistry (ICC) demonstrated that the expression of integrins alpha3, alpha5 and alphaV diminished during the culture period. This change was coincident with both a decrease in beta-cell function (proinsulin gene expression, islet insulin content and stimulated insulin release) and a rise in beta-cell death from apoptosis, as determined by in situ cell death detection (TUNEL) assay. These adverse events were prevented or delayed by exposure of islets to matrix proteins. In conclusion, routine islet isolation disrupts the cell-matrix relationship leading to a variety of structural and functional abnormalities, including apoptotic cell death. These alterations can be diminished by restoration of a culture microenvironment that includes matrix proteins.
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Hadari, Y. R., R. Arbel-Goren, Y. Levy, A. Amsterdam, R. Alon, R. Zakut, and Y. Zick. "Galectin-8 binding to integrins inhibits cell adhesion and induces apoptosis." Journal of Cell Science 113, no. 13 (July 1, 2000): 2385–97. http://dx.doi.org/10.1242/jcs.113.13.2385.
Full textAbstract:
The interaction of cells with the extracellular matrix regulates cell adhesion, motility, growth, survival and differentiation through integrin-mediated signal transduction. Here we demonstrate that galectin-8, a secreted mammalian (beta)-galactoside binding protein, inhibits adhesion of human carcinoma (1299) cells to plates coated with integrin ligands, and induces cell apoptosis. Pretreatment of the cells with Mn(2+), which increases the affinity of integrins for their ligands, abolished the inhibitory effects of galectin-8. The inhibitory effects of galectin-8 were specific and were not mimicked by plant lectins or other galectins (galectin-1 and galectin-3). In accordance with its anti-adhesive effects, transfection of galectin-8 cDNA into 1299 cells significantly reduced (by 75%) colony formation, when compared to the number of colonies formed by cells transfected with an empty vector. Affinity chromatography over immobilized galectin-8 indicated that few membrane proteins interacted with galectin-8 in a sugar-dependent manner. Microsequencing and western immunoblotting revealed that (alpha)(3)(beta)(1)integrin derived from 1299 as well as other cells (e.g. HeLa and human endothelial cells) is a major galectin-8 binding-protein. Furthermore, immunoprecipitation and immunohistochemical studies suggested that endogenous galectin-8, secreted from 1299 cells, forms complexes with (alpha)(3)(beta)(1) integrins expressed on the surface of 1299 cells. Galectin-8 also interacts with other members of the integrin family, like (alpha)(6)(beta)(1)integrins. In contrast, galectin-8 only minimally interacts with (alpha)(4)or (beta)(3)integrins. We propose that galectin-8 is an integrin binding-protein that interacts to a different extent with several, but not all members of the integrin family. Binding of galectin-8 modulates integrin interactions with the extracellular matrix and thus regulates cell adhesion and cell survival.
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Malek-Hedayat, S., and LH Rome. "Expression of a beta 1-related integrin by oligodendroglia in primary culture: evidence for a functional role in myelination." Journal of Cell Biology 124, no. 6 (March 15, 1994): 1039–46. http://dx.doi.org/10.1083/jcb.124.6.1039.
Full textAbstract:
We have investigated the expression of integrins by rat oligodendroglia grown in primary culture and the functional role of these proteins in myelinogenesis. Immunochemical analysis, using antibodies to a number of alpha and beta integrin subunits, revealed that oligodendrocytes express only one detectable integrin receptor complex (alpha OL beta OL). This complex is immunoprecipitated by a polyclonal anti-human beta 1 integrin subunit antibody. In contrast, astrocytes, the other major glial cell type in brain, express multiple integrins including alpha 1 beta 1, alpha 3 beta 1, and alpha 5 beta 1 complexes that are immunologically and electrophoretically indistinguishable from integrins expressed by rat fibroblasts. The beta subunit of the oligodendrocyte integrin (beta OL) and rat fibroblast beta 1 have different electrophoretic mobilities in SDS-PAGE. However, the two beta subunits appear to be highly related based on immunological cross-reactivity and one-dimensional peptide mapping. After removal of N-linked carbohydrate chains, beta OL and beta 1 comigrated in SDS-PAGE and peptide maps of the two deglycosylated subunits were identical, suggesting differential glycosylation of beta 1 and beta OL accounts entirely for their size differences. The oligodendrocyte alpha subunit, alpha OL, was not immunoprecipitated by antibodies against well characterized alpha chains which are known to associate with beta 1 (alpha 3, alpha 4, and alpha 5). However, an antibody to alpha 8, a more recently identified integrin subunit, did precipitate two integrin subunits with electrophoretic mobilities in SDS-PAGE identical to alpha OL and beta OL. Functional studies indicated that disruption of oligodendrocyte adhesion to a glial-derived matrix by an RGD-containing synthetic peptide resulted in a substantial decrease in the level of mRNAs for several myelin components including myelin basic protein (MBP), proteolipid protein (PLP), and cyclic nucleotide phosphodiesterase (CNP). These results suggest that integrin-mediated adhesion of oligodendrocytes may trigger signal(s) that induce the expression of myelin genes and thus influence oligodendrocyte differentiation.