An essential role of integrin-linked kinase in the cellular radiosensitivity of normal fibroblasts during the process of cell adhesion and spreading.
OBJECTIVE
In addition to focal adhesion kinase (FAK), Paxillin and p130 Crk-associated substrate (p130Cas), integrin-linked kinase (ILK) mediates signals from beta integrins for controlling, e.g., survival, adhesion and spreading. To evaluate the role of ILK in the cellular radiosensitivity at different stages of cell adhesion and spreading, ILK(floxed/floxed (fl/fl)) and ILK(-/-) mouse fibroblasts were examined.


METHODS
Cells were irradiated (0 - 4 Gy, X-rays) in suspension, after varying time periods on fibronectin (FN) or after 24 h on different matrix proteins. Irradiation was combined with phosphatidylinositol-3 kinase (PI3K) inhibition using Ly294002. Clonogenic radiation survival, cell adhesion, and kinetics of protein expression and phosphorylation during FN adhesion (ILK, v-akt murine thymoma viral oncogene homolog 1 (AKT), FAK, Paxillin, p130Cas) were examined.


RESULTS
In suspension and during the first hour on FN, irradiated ILK(fl/fl) cells survived significantly better than ILK(-/-) cells in a PI3K- and serum-dependent manner. 24-h cell cultures on different matrix proteins showed no difference in radiosensitivity. During FN adhesion, which was slightly impaired in ILK(-/-) cells, protein kinetics uncovered differences in AKT, FAK, Paxillin and p130Cas phosphorylation in the two cell lines. Phosphorylation of FAK, Paxillin and p130Cas was downregulated upon exposure to ionizing radiation in an ILK-independent manner.


CONCLUSIONS
These findings indicate a critical function of ILK in the cellular radiosensitivity during the early stages of adhesion to and spreading on FN. On the basis of the presented data, a precise correlation of adhesion-, serum- and PI3K-mediated changes in PI3K/AKT and FAK/Paxillin/p130Cas signaling cascades was not found. However, identifying the underlying mechanisms of adhesion- and spreading-related changes in the cellular radiosensitivity might be relevant for an optimization of radiotherapeutic strategies specifically targeting cells located at the invasive edge of a malignant tumor.
Hehlgans
Stephanie
S
OncoRay - Center for Radiation Research in Oncology, Medical Faculty Carl Gustav Carus, Dresden University of Technology, Germany.
Eke
Iris
I
Cordes
Nils
N
D000818:Animals; D002448:Cell Adhesion / Q000502:physiology* / Q000528:radiation effects*; D002460:Cell Line; D002465:Cell Movement / Q000502:physiology* / Q000528:radiation effects*; D002470:Cell Survival / Q000502:physiology / Q000528:radiation effects; D002867:Chromones / Q000494:pharmacology; D050719:Crk-Associated Substrate Protein / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects; D005353:Fibronectins / Q000378:metabolism; D051417:Focal Adhesion Kinase 1 / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009025:Morpholines / Q000494:pharmacology; D051419:Paxillin / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D051057:Proto-Oncogene Proteins c-akt; D011836:Radiation Tolerance / Q000502:physiology*; D011853:Radiobiology; D015398:Signal Transduction; D000099200:Scaffold Protein ILK
D002867:Chromones; D050719:Crk-Associated Substrate Protein; D005353:Fibronectins; D051417:Focal Adhesion Kinase 1; D009025:Morpholines; D051419:Paxillin; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D000099200:Scaffold Protein ILK; C496062:Pxn protein, mouse; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C496051:Ptk2 protein, mouse