Role of Raf-1 and FAK in cell density-dependent regulation of integrin-dependent activation of MAP kinase.
MAP kinase can be activated by integrin-dependent adhesion in a FAK-dependent manner. Cell-cell contact inhibition is continuously active in controlling cell growth and the loss of cell-cell contact inhibition is correlated with the malignant characteristics of cancer cells. In this study we showed that cell adhesion to fibronectin for 1 h activated MAP kinase phosphorylation. However, when non-tumorigenic HSG cells, MCF-10A cells, or 293 cells were plated on fibronectin-coated substrates for 1 h at high cell density (which favors cell-cell contact), MAP kinase phosphorylation was not enhanced. Tumorigenic breast cancer cells, BT474, Cama, MCF-7, MDA-MB-231 and SKBR3, did not show inhibition of MAP kinase phosphorylation but rather enhanced MAP kinase phosphorylation when cultured at high density on fibronectin-coated substrates. Adhesion of HSG cells to fibronectin also increased FAK phosphorylation and this FAK phosphorylation was partially inhibited when cells were cultured at high density. Expression of Raf-1 catalytic domain-GFP in HSG cells could overcome the cell density-dependent inhibition of MAP kinase phosphorylation and FAK phosphorylation. The expression of Raf-1-catalytic domain-GFP also upregulated the expression of alphav integrin and promoted cell-cell adhesion in HSG cells. These results suggest that the active form of Raf-1 may interrupt cell-cell contact inhibition by promoting alphav integrin expression, which has been implicated in cell aggregation.
Division of Tumor Biology, Northeastern Ontario Regional Cancer Center, Sudbury, Ontario P3E 5J1, Canada.
Lianfeng
L
Zhang
Mary
M
Bewick
Robert M
RM
Lafrenie
D015703:Antigens, CD / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002448:Cell Adhesion / Q000502:physiology; D002450:Cell Communication / Q000502:physiology; D002452:Cell Count; D002478:Cells, Cultured; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005353:Fibronectins / Q000378:metabolism*; D005419:Flavonoids / Q000494:pharmacology; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D049452:Green Fluorescent Proteins; D006801:Humans; D015151:Immunoblotting; D039564:Integrin alphaV; D008164:Luminescent Proteins / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009369:Neoplasms / Q000378:metabolism*; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D019908:Proto-Oncogene Proteins c-raf / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology
D015703:Antigens, CD; D004791:Enzyme Inhibitors; D005353:Fibronectins; D005419:Flavonoids; D039564:Integrin alphaV; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; D019908:Proto-Oncogene Proteins c-raf; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one