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Stefan Rosewicz - One of the best experts on this subject based on the ideXlab platform.
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galectin 1 sensitizes carcinoma cells to anoikis via the Fibronectin Receptor α5β1 integrin
Cell Death & Differentiation, 2011Co-Authors: H Sanchezruderisch, Katharina M. Detjen, Martina Welzel, Sabine André, Christian Fischer, Hans-joachim Gabius, Stefan RosewiczAbstract:Anoikis resistance is a hallmark of transformed epithelial cells. Here, we show that treatment of anoikis-resistant carcinoma cell lines with the endogenous lectin galectin-1 (Gal-1) promoted apoptosis via interaction with the unligated Fibronectin Receptor α5β1-integrin. Gal-1 efficiency correlated with expression of α5β1-integrin, and transfection of the α5-subunit into deficient cell lines conferred Gal-1 binding and anoikis stimulation. Furthermore, Gal-1 and the α5- and β1-integrin subunits co-precipitated in Gal-1-stimulated cells undergoing anoikis. Other members of the galectin family failed to be active. The functional interaction between Gal-1 and α5β1-integrin was glycan dependent with α2,6-sialylation representing a switch-off signal. Desialylation of cell surface glycans resulted in increased electrophoretic mobility of α5β1-integrin and facilitated Gal-1 binding and anoikis stimulation. On the level of signaling, Gal-1-stimulated anoikis was prevented by filipin, which impaired the internalization of α5β1-integrin via cholesterol-enriched microdomains, and by pretreatment with a caspase-8 inhibitor. We propose that Gal-1/α5β1-integrin interaction participates in the control of epithelial integrity and integrin sialylation may enable carcinoma cells to evade this Gal-1-dependent control mechanism.
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Galectin-1 sensitizes carcinoma cells to anoikis via the α5β1 Fibronectin Receptor
Cell Death and Differentiation, 2010Co-Authors: Katharina M. Detjen, Hugo Sanchez-ruderisch, Martina Welzel, Sabine André, Christian Fischer, Hans-joachim Gabius, Stefan RosewiczAbstract:Anoikis resistance is a hallmark of transformed epithelial cells. Here, we show that treatment of anoikis-resistant carcinoma cell lines with the endogenous lectin galectin-1 (Gal-1) promoted apoptosis via interaction with the unligated Fibronectin Receptor α5β1-integrin. Gal-1 efficiency correlated with expression of α5β1-integrin, and transfection of the α5-subunit into deficient cell lines conferred Gal-1 binding and anoikis stimulation. Furthermore, Gal-1 and the α5-and β1-integrin subunits coprecipitated in Gal-1-stimulated cells undergoing anoikis. Other members of the galectin family failed to be active. The functional interaction between Gal-1 and α5β1-integrin was glycan-dependent with α2,6-sialylation representing a switch-off signal. Desialylation of cell surface glycans resulted in increased electrophoretic mobility of α5β1-integrin and facilitated Gal-1 binding and anoikis stimulation. On the level of signaling, Gal-1-stimulated anoikis was prevented by filipin, which impaired the internalization of α5β1-integrin via cholesterol-enriched microdomains, and by pretreatment with a caspase-8 inhibitor. We propose that Gal-1/α5β1-integrin interaction participates in the control of epithelial integrity and integrin sialylation may enable carcinoma cells to evade this Gal 1 dependent control mechanism.
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a novel function for the tumor suppressor p16ink4a induction of anoikis via upregulation of the α5β1 Fibronectin Receptor
Journal of Cell Biology, 2000Co-Authors: Thomas Plath, Katharina M. Detjen, M Welzel, Zofia Von Marschall, Derek Murphy, Michael Schirner, B Wiedenmann, Stefan RosewiczAbstract:The tumor suppressor gene p16INK4a inhibits the kinase activity of the cyclin-dependent kinase 4–6/cyclin D complexes and subsequent phosphorylation of critical substrates necessary for transit through the G1 phase of the cell cycle. Recent studies suggested that control of the G1/S boundary might not be the sole biological function of p16INK4a. We hypothesized that p16INK4a might influence hitherto unknown critical features of a malignant epithelial phenotype, such as anchorage dependence. Here we provide evidence that stable transfection of p16INK4a restitutes apoptosis induction upon loss of anchorage (anoikis) in a variety of human cancer cells. Anoikis in p16INK4a-transfected cells was evidenced by DNA fragmentation and poly(ADP-ribose) polymerase cleavage upon cultivation on polyhydroxyethylmethacrylate-coated dishes and was associated with suppression of anchorage-independent growth as well as complete loss of tumorigenicity. p16INK4a-mediated anoikis was due to selective transcriptional upregulation of the α5 integrin chain of the α5β1 Fibronectin Receptor as detected by FACS® analysis, immunoprecipitation, Northern blotting, and nuclear run-on assays. Addition of soluble Fibronectin and inhibitory α5 antibodies to nonadherent cells completely abolished p16INK4a-mediated anoikis, whereas laminin was ineffective. Furthermore, antisense-induced downregulation of the α5 integrin chain in p16INK4a-transfected cells restored resistance to anoikis. These data suggest a novel functional interference between a cell cycle–regulating tumor suppressor gene and membrane-bound integrins, thus regulating a hallmark feature of an epithelial transformed phenotype: susceptibility to anoikis.
Magnus Höök - One of the best experts on this subject based on the ideXlab platform.
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Fibronectin Receptors from gram-positive bacteria: Comparison of active sites
Biochemistry, 1994Co-Authors: Hyeon J. Joh, Sivashankarappa Gurusiddappa, Karen House-pompeo, Joseph M. Patti, Magnus HöökAbstract:Many parasitic bacteria express Fibronectin binding proteins that are located on the cell surface. These proteins may act as adhesins and mediate the adherence of the microorganisms to Fibronectin-containing host tissues. The ligand binding sites in the Fibronectin Receptor proteins from Gram-positive bacteria are composed of unique 37-48 amino acid long motifs that are repeated 3-4 times. We have now expressed the ligand binding sites of Fibronectin Receptors from Staphylococcus aureus, Streptococcus dysgalactiae (two Receptors), and Streptococcus pyogenes as recombinant proteins. The purified recombinant proteins have the expected molecular weights as indicated by electrospray mass spectroscopy although they migrate abnormally on SDS-PAGE. Each recombinant protein effectively inhibited the binding of 125I-labeled intact Fibronectin or the N-terminal Fibronectin domain to Staphylococcus aureus, Streptococcus dysgalactiae, and Streptococcus pyogenes. The relative inhibitory potency of the different recombinant proteins was similar for all target bacteria and is reflected in their relative affinities for Fibronectin. Synthetic peptides corresponding to the repeat units of the ligand binding site of the Fibronectin Receptor proteins were shown to inhibit the binding of the N-terminal Fibronectin fragment to Streptococcus pyogenes cells. Together with amino acid sequence comparison, these data demonstrate that the repeat motif of the Fibronectin Receptor of Streptococcus pyogenes conforms to the consensus sequence previously reported for the Staphylococcus aureus Receptor and to one of the Streptococcus dysgalactiae Receptors (McGavin et al., 1993).
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Fibronectin binding determinants of the Staphylococcus aureus Fibronectin Receptor.
The Journal of biological chemistry, 1991Co-Authors: Martin J. Mcgavin, Giuseppe Raucci, Sivashankarappa Gurusiddappa, Magnus HöökAbstract:Abstract Synthetic peptide analogs mimicking a repeated motif within the Staphylococcus aureus Fibronectin Receptor inhibit binding of the bacteria to Fibronectin (Signas, C., Raucci, G., Jonsson, K., Lindgren, P. E., Anantharamaiah, G. M., Hook, M., and Lindberg, M. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 699-703). In this study, we have further localized the Fibronectin-binding determinant within the 37 amino acid D3 peptide. Chemical modification of the carboxyl side chains of the glutamic and aspartic residues in D3 abolished Fibronectin-binding activity, whereas modifications of lysine or tyrosine residues had little effect. An active peptide encompassing residues 15-36 was isolated from a trypsin digest of D3, and a synthetic peptide S16-36 had activity comparable with that of intact D3. Scrambling the amino acid sequence of S16-36 or replacing the aspartic and glutamic residues with asparagine and glutamine resulted in loss of activity. Therefore, one or more of the acidic residues are essential for activity. However, additional sequence is required. Reduction in the size of S16-36 from either the N- or C-terminal end resulted in peptides with greatly diminished activity. These data suggest that the amino acids essential for binding Fibronectin are contained within residues 21-33 of the D3 peptide and that the flanking N- and C-terminal amino acids are necessary for the peptide to acquire a conformation that is favorable for Fibronectin binding.
R. L. Juliano - One of the best experts on this subject based on the ideXlab platform.
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The alpha v beta 1 integrin functions as a Fibronectin Receptor but does not support Fibronectin matrix assembly and cell migration on Fibronectin
The Journal of cell biology, 1993Co-Authors: Zhuohua Zhang, R. L. Juliano, Alex Morla, Kristiina Vuori, Jeffrey S. Bauer, Erkki RuoslahtiAbstract:The Fibronectin Receptor, alpha 5 beta 1, has been shown to be required for Fibronectin matrix assembly and plays an important role in cell migration on Fibronectin. However, it is not clear whether other Fibronectin binding integrins can take the place of alpha 5 beta 1 during matrix assembly and cell migration. To test this, we expressed the human alpha v subunit in the CHO cell line CHO-B2 that lacks the alpha 5 subunit. We found that the human alpha v combined with CHO cell beta 1 to form the integrin alpha v beta 1. Cells that expressed alpha v beta 1 attached to and spread well on Fibronectin-coated dishes, but did so less well on vitronectin-coated dishes. This, along with other data, indicated that alpha v beta 1 functions as a Fibronectin Receptor in CHO-B2 cells. The alpha v beta 1-expressing cells failed to produce a Fibronectin matrix or to migrate on Fibronectin, although the same cells transfected with alpha 5 do produce a matrix and migrate on Fibronectin. The affinity of the alpha v beta 1-expressing cells for Fibronectin was fourfold lower than that of the alpha 5 beta 1-expressing cells. In addition, alpha v beta 1 was distributed diffusely throughout the cell surface, whereas alpha 5 beta 1 was localized to focal adhesions when cells were seeded onto Fibronectin-coated surfaces. Thus, of the two Fibronectin Receptors, alpha v beta 1 and alpha 5 beta 1, only alpha 5 beta 1 supports Fibronectin matrix assembly and promotes cell migration on Fibronectin in the CHO-B2 cells. Possible reasons for this difference in the activities of alpha v beta 1 and alpha 5 beta 1 include the lower affinity of alpha v beta 1 for Fibronectin and the failure of this integrin to localize in adhesion plaques on a Fibronectin substrate. These results show that two integrins with similar ligand specificities and cell attachment functions may be quite different in their ability to support Fibronectin matrix assembly and cell motility on Fibronectin.
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motility of Fibronectin Receptor deficient cells on Fibronectin and vitronectin collaborative interactions among integrins
Journal of Cell Biology, 1992Co-Authors: J S Bauer, Clara Schreiner, Filippo G Giancotti, E Ruoslahti, R. L. JulianoAbstract:Cells are capable of adhering to and migrating on protein components of the extracellular matrix. These cell-matrix interactions are thought to be mediated largely through a family of cell surface Receptors termed integrins. However, the manner in which individual integrins are involved in cell adhesion and motility has not been fully determined. To explore this issue, we previously selected a series of CHO variants that are deficient in expression of the integrin alpha 5 beta 1, the "classical" Fibronectin Receptor. Two sets of subclones of these variants were defined which respectively express approximately 20% or 2% of Fibronectin Receptor on the cell surface when compared to wild-type cells (Schreiner, C. L., J. S. Bauer, Y. N. Danilov, S. Hussein, M. M. Sczekan, and R. L. Juliano. 1989. J. Cell Biol. 109:3157-3167). In the current study, the variant clones were tested for haptotactic motility on substrata coated with Fibronectin or vitronectin. Data from assays using Fibronectin show that cellular motility of the 20% variants was substantially decreased (30-75% of wild type), while the motility of the 2% variants was nearly abolished (2-20% of wild type). Surprisingly, a similar pattern was seen for haptotactic motility of both 2% and 20% variants when vitronectin was used (approximately 20-30% of wild type). The reduced haptotactic motility of the Fibronectin Receptor-deficient variant clones on vitronectin was shown not to be due to reduced vitronectin Receptor (alpha v beta 3) expression nor to a failure of these variants to adhere to vitronectin substrata. Transfection of the deficient variants with a cDNA for the human alpha 5 subunit resulted in normal levels of Fibronectin Receptor expression (as a human alpha 5/hamster beta 1 chimera) and restored the motility of the CHO variants on Fibronectin and vitronectin. This indicates that expression of the alpha 5 subunit is required for normal haptotactic motility on vitronectin substrata and suggests that the Fibronectin Receptor (alpha 5 beta 1) plays a cooperative role with vitronectin Receptors in cell motility.
Clara Schreiner - One of the best experts on this subject based on the ideXlab platform.
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motility of Fibronectin Receptor deficient cells on Fibronectin and vitronectin collaborative interactions among integrins
Journal of Cell Biology, 1992Co-Authors: J S Bauer, Clara Schreiner, Filippo G Giancotti, E Ruoslahti, R. L. JulianoAbstract:Cells are capable of adhering to and migrating on protein components of the extracellular matrix. These cell-matrix interactions are thought to be mediated largely through a family of cell surface Receptors termed integrins. However, the manner in which individual integrins are involved in cell adhesion and motility has not been fully determined. To explore this issue, we previously selected a series of CHO variants that are deficient in expression of the integrin alpha 5 beta 1, the "classical" Fibronectin Receptor. Two sets of subclones of these variants were defined which respectively express approximately 20% or 2% of Fibronectin Receptor on the cell surface when compared to wild-type cells (Schreiner, C. L., J. S. Bauer, Y. N. Danilov, S. Hussein, M. M. Sczekan, and R. L. Juliano. 1989. J. Cell Biol. 109:3157-3167). In the current study, the variant clones were tested for haptotactic motility on substrata coated with Fibronectin or vitronectin. Data from assays using Fibronectin show that cellular motility of the 20% variants was substantially decreased (30-75% of wild type), while the motility of the 2% variants was nearly abolished (2-20% of wild type). Surprisingly, a similar pattern was seen for haptotactic motility of both 2% and 20% variants when vitronectin was used (approximately 20-30% of wild type). The reduced haptotactic motility of the Fibronectin Receptor-deficient variant clones on vitronectin was shown not to be due to reduced vitronectin Receptor (alpha v beta 3) expression nor to a failure of these variants to adhere to vitronectin substrata. Transfection of the deficient variants with a cDNA for the human alpha 5 subunit resulted in normal levels of Fibronectin Receptor expression (as a human alpha 5/hamster beta 1 chimera) and restored the motility of the CHO variants on Fibronectin and vitronectin. This indicates that expression of the alpha 5 subunit is required for normal haptotactic motility on vitronectin substrata and suggests that the Fibronectin Receptor (alpha 5 beta 1) plays a cooperative role with vitronectin Receptors in cell motility.
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Increased tumorigenicity of Fibronectin Receptor deficient Chinese hamster ovary cell variants.
Cancer research, 1991Co-Authors: Clara Schreiner, Michael Fisher, Saber Hussein, Rudy L. JulianoAbstract:Abstract We have developed a series of variant clonal Chinese hamster ovary cell sublines which are deficient in the expression of the α 5 /β 1 integrin Fibronectin Receptor. When these clonal sublines were injected s.c. into nude mice, there were marked differences in the rate of tumor growth. Tumors from clones expressing very low levels of Fibronectin Receptor grew most rapidly, clones expressing levels of Fibronectin Receptor comparable to wild type cells grew at an intermediate rate, while a clone expressing elevated levels of Fibronectin Receptor grew slowly. Cells recovered from the tumors maintained their original phenotypes in terms of levels of Fibronectin Receptor expression. These results suggest that there is an inverse correlation between the level of expression of the α 5 /β 1 Fibronectin Receptor and the rate of tumor growth.
Katharina M. Detjen - One of the best experts on this subject based on the ideXlab platform.
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galectin 1 sensitizes carcinoma cells to anoikis via the Fibronectin Receptor α5β1 integrin
Cell Death & Differentiation, 2011Co-Authors: H Sanchezruderisch, Katharina M. Detjen, Martina Welzel, Sabine André, Christian Fischer, Hans-joachim Gabius, Stefan RosewiczAbstract:Anoikis resistance is a hallmark of transformed epithelial cells. Here, we show that treatment of anoikis-resistant carcinoma cell lines with the endogenous lectin galectin-1 (Gal-1) promoted apoptosis via interaction with the unligated Fibronectin Receptor α5β1-integrin. Gal-1 efficiency correlated with expression of α5β1-integrin, and transfection of the α5-subunit into deficient cell lines conferred Gal-1 binding and anoikis stimulation. Furthermore, Gal-1 and the α5- and β1-integrin subunits co-precipitated in Gal-1-stimulated cells undergoing anoikis. Other members of the galectin family failed to be active. The functional interaction between Gal-1 and α5β1-integrin was glycan dependent with α2,6-sialylation representing a switch-off signal. Desialylation of cell surface glycans resulted in increased electrophoretic mobility of α5β1-integrin and facilitated Gal-1 binding and anoikis stimulation. On the level of signaling, Gal-1-stimulated anoikis was prevented by filipin, which impaired the internalization of α5β1-integrin via cholesterol-enriched microdomains, and by pretreatment with a caspase-8 inhibitor. We propose that Gal-1/α5β1-integrin interaction participates in the control of epithelial integrity and integrin sialylation may enable carcinoma cells to evade this Gal-1-dependent control mechanism.
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Galectin-1 sensitizes carcinoma cells to anoikis via the α5β1 Fibronectin Receptor
Cell Death and Differentiation, 2010Co-Authors: Katharina M. Detjen, Hugo Sanchez-ruderisch, Martina Welzel, Sabine André, Christian Fischer, Hans-joachim Gabius, Stefan RosewiczAbstract:Anoikis resistance is a hallmark of transformed epithelial cells. Here, we show that treatment of anoikis-resistant carcinoma cell lines with the endogenous lectin galectin-1 (Gal-1) promoted apoptosis via interaction with the unligated Fibronectin Receptor α5β1-integrin. Gal-1 efficiency correlated with expression of α5β1-integrin, and transfection of the α5-subunit into deficient cell lines conferred Gal-1 binding and anoikis stimulation. Furthermore, Gal-1 and the α5-and β1-integrin subunits coprecipitated in Gal-1-stimulated cells undergoing anoikis. Other members of the galectin family failed to be active. The functional interaction between Gal-1 and α5β1-integrin was glycan-dependent with α2,6-sialylation representing a switch-off signal. Desialylation of cell surface glycans resulted in increased electrophoretic mobility of α5β1-integrin and facilitated Gal-1 binding and anoikis stimulation. On the level of signaling, Gal-1-stimulated anoikis was prevented by filipin, which impaired the internalization of α5β1-integrin via cholesterol-enriched microdomains, and by pretreatment with a caspase-8 inhibitor. We propose that Gal-1/α5β1-integrin interaction participates in the control of epithelial integrity and integrin sialylation may enable carcinoma cells to evade this Gal 1 dependent control mechanism.
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a novel function for the tumor suppressor p16ink4a induction of anoikis via upregulation of the α5β1 Fibronectin Receptor
Journal of Cell Biology, 2000Co-Authors: Thomas Plath, Katharina M. Detjen, M Welzel, Zofia Von Marschall, Derek Murphy, Michael Schirner, B Wiedenmann, Stefan RosewiczAbstract:The tumor suppressor gene p16INK4a inhibits the kinase activity of the cyclin-dependent kinase 4–6/cyclin D complexes and subsequent phosphorylation of critical substrates necessary for transit through the G1 phase of the cell cycle. Recent studies suggested that control of the G1/S boundary might not be the sole biological function of p16INK4a. We hypothesized that p16INK4a might influence hitherto unknown critical features of a malignant epithelial phenotype, such as anchorage dependence. Here we provide evidence that stable transfection of p16INK4a restitutes apoptosis induction upon loss of anchorage (anoikis) in a variety of human cancer cells. Anoikis in p16INK4a-transfected cells was evidenced by DNA fragmentation and poly(ADP-ribose) polymerase cleavage upon cultivation on polyhydroxyethylmethacrylate-coated dishes and was associated with suppression of anchorage-independent growth as well as complete loss of tumorigenicity. p16INK4a-mediated anoikis was due to selective transcriptional upregulation of the α5 integrin chain of the α5β1 Fibronectin Receptor as detected by FACS® analysis, immunoprecipitation, Northern blotting, and nuclear run-on assays. Addition of soluble Fibronectin and inhibitory α5 antibodies to nonadherent cells completely abolished p16INK4a-mediated anoikis, whereas laminin was ineffective. Furthermore, antisense-induced downregulation of the α5 integrin chain in p16INK4a-transfected cells restored resistance to anoikis. These data suggest a novel functional interference between a cell cycle–regulating tumor suppressor gene and membrane-bound integrins, thus regulating a hallmark feature of an epithelial transformed phenotype: susceptibility to anoikis.