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Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3β (GSK-3β) in vitro

Cell adhesion to extracellular matrix (ECM) is thought to confer resistance against cell-damaging agents, that is, drugs and radiation, in tumour and normal cells in vitro. The dependence of cell survival on β1-integrin-linked kinase (ILK), protein kinase Bα/Akt (PKBα/Akt) and glycogen synthase kina...

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Autores principales: Cordes, N, van Beuningen, D
Formato: Texto
Lenguaje:English
Publicado: Nature Publishing Group 2003
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2741045/
https://www.ncbi.nlm.nih.gov/pubmed/12778079
http://dx.doi.org/10.1038/sj.bjc.6600912
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author Cordes, N
van Beuningen, D
author_facet Cordes, N
van Beuningen, D
author_sort Cordes, N
collection PubMed
description Cell adhesion to extracellular matrix (ECM) is thought to confer resistance against cell-damaging agents, that is, drugs and radiation, in tumour and normal cells in vitro. The dependence of cell survival on β1-integrin-linked kinase (ILK), protein kinase Bα/Akt (PKBα/Akt) and glycogen synthase kinase-3β (GSK-3β) activity, which participate in β1-integrin signalling and cell cycle progression was investigated as a function of radiation exposure. Colony-formation assays on polystyrene, fibronectin (FN), laminin (LA), bovine serum albumin (BSA) or poly-L-lysine (poly-L) (0–8 Gy), kinase assays, flow cytometric DNA and annexin-V analysis and immunoblotting were performed in nonirradiated and irradiated (2 or 6 Gy) A549 human lung cancer cells and CCD32 normal human lung fibroblasts. Cell contact to FN in contrast to polystyrene elevated basal ILK, PKBα/Akt and GSK-3β kinase activities in A549 and CCD32 cells, as well as the basal amount of A549 G2 phase cells. Irradiation on FN or LA as compared to polystyrene, BSA or poly-L significantly improved cell survival. Following irradiation, kinase activities were stimulated strongly on polystyrene but showed to be less prominent on FN, which was because of the FN-related basal induction. Following irradiation, FN compared to polystyrene enlarged and prolonged G2 arrest in both the cell lines. For the analysis of phosphatidylinositol-3 kinase (PI3-K) dependence of protein kinases and cell cycle transition, the PI3-K inhibitors LY294002 and wortmannin were used showing decreased kinase activities, antiproliferative and radiation-dependent G2 accumulation-abrogating effects accompanied by downregulation of cyclin D1 and phospho-pRb in cells attached to polystyrene. Fibronectin partly abrogated these effects PI3-K-independently. These findings suggest a novel pathway that makes direct phosphorylation of GSK-3β by ILK feasible after irradiation. Conclusively, the data indicate that ILK, PKBα/Akt and GSK-3β are involved in modulations of the cell cycle after irradiation. These interactions are strictly dependent on ECM components in a cell line-specific manner. Our findings provide molecular insights into mechanisms likely to be important for ECM-dependent cell survival and cellular radioresistance as well as tumour growth.
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spelling pubmed-27410452009-09-10 Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3β (GSK-3β) in vitro Cordes, N van Beuningen, D Br J Cancer Experimental Therapeutics Cell adhesion to extracellular matrix (ECM) is thought to confer resistance against cell-damaging agents, that is, drugs and radiation, in tumour and normal cells in vitro. The dependence of cell survival on β1-integrin-linked kinase (ILK), protein kinase Bα/Akt (PKBα/Akt) and glycogen synthase kinase-3β (GSK-3β) activity, which participate in β1-integrin signalling and cell cycle progression was investigated as a function of radiation exposure. Colony-formation assays on polystyrene, fibronectin (FN), laminin (LA), bovine serum albumin (BSA) or poly-L-lysine (poly-L) (0–8 Gy), kinase assays, flow cytometric DNA and annexin-V analysis and immunoblotting were performed in nonirradiated and irradiated (2 or 6 Gy) A549 human lung cancer cells and CCD32 normal human lung fibroblasts. Cell contact to FN in contrast to polystyrene elevated basal ILK, PKBα/Akt and GSK-3β kinase activities in A549 and CCD32 cells, as well as the basal amount of A549 G2 phase cells. Irradiation on FN or LA as compared to polystyrene, BSA or poly-L significantly improved cell survival. Following irradiation, kinase activities were stimulated strongly on polystyrene but showed to be less prominent on FN, which was because of the FN-related basal induction. Following irradiation, FN compared to polystyrene enlarged and prolonged G2 arrest in both the cell lines. For the analysis of phosphatidylinositol-3 kinase (PI3-K) dependence of protein kinases and cell cycle transition, the PI3-K inhibitors LY294002 and wortmannin were used showing decreased kinase activities, antiproliferative and radiation-dependent G2 accumulation-abrogating effects accompanied by downregulation of cyclin D1 and phospho-pRb in cells attached to polystyrene. Fibronectin partly abrogated these effects PI3-K-independently. These findings suggest a novel pathway that makes direct phosphorylation of GSK-3β by ILK feasible after irradiation. Conclusively, the data indicate that ILK, PKBα/Akt and GSK-3β are involved in modulations of the cell cycle after irradiation. These interactions are strictly dependent on ECM components in a cell line-specific manner. Our findings provide molecular insights into mechanisms likely to be important for ECM-dependent cell survival and cellular radioresistance as well as tumour growth. Nature Publishing Group 2003-05-06 2003-04-29 /pmc/articles/PMC2741045/ /pubmed/12778079 http://dx.doi.org/10.1038/sj.bjc.6600912 Text en Copyright © 2003 Cancer Research UK https://creativecommons.org/licenses/by/4.0/This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material.If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit https://creativecommons.org/licenses/by/4.0/.
spellingShingle Experimental Therapeutics
Cordes, N
van Beuningen, D
Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3β (GSK-3β) in vitro
title Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3β (GSK-3β) in vitro
title_full Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3β (GSK-3β) in vitro
title_fullStr Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3β (GSK-3β) in vitro
title_full_unstemmed Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3β (GSK-3β) in vitro
title_short Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3β (GSK-3β) in vitro
title_sort cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent g2 phase arrest involving integrin-linked kinase (ilk) and glycogen synthase kinase-3β (gsk-3β) in vitro
topic Experimental Therapeutics
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2741045/
https://www.ncbi.nlm.nih.gov/pubmed/12778079
http://dx.doi.org/10.1038/sj.bjc.6600912
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