Regulation by ceramide of epidermal growth factor signal transduction and mitogenesis in cell lines overexpressing the growth factor receptor.
Ceramide has emerged as a pleiotropic signal mediator of cellular responses including differentiation, proliferation, cell cycle arrest and apoptosis. In the present study we evaluated the effect of cell permeant ceramide analogues on ligand-induced tyrosine phosphorylation of the EGF receptor (EGFR), phospholipase Cy (PLCgamma) activity and cell proliferation. Treatment with N-acetylsphingosine (C2-cer) and N-hexanoylceramide (C6-cer) prevented EGF-induced tyrosine trans-phosphorylation of the receptor in two different cell lines overexpressing the human EGFR (A431 and EGF-T17 cells). In contrast, treatment of A431 and EGFR-T17 cells with C2-cer or C6-cer did not affect the ligand binding capacity of the receptor, an effect that was however observed after TPA-induced activation of PKC. In addition EGF-stimulated PLCgamma activity was transiently decreased in A431 cells treated with C6-cer and only a modest, albeit significant reduction on ligand-induced 3H-InsP3 generation was observed in EGFR-T17 cells pretreated with ceramide. We also examined the effect of C2-cer on serum (A431)- or EGF (EGFR-T 17)-induced cell proliferation. Treatment of EGFR-TI7 cells with C2-cer (0.1-10 microM) did not affect cell viability, but prevented EGF-induced 3H-thymidine incorporation in a dose-dependent manner. In contrast, 3H-thymidine incorporation in serum-stimulated A431 cells decreased only at the higher doses of C2-cer used (1-10 microM), being this effect accompanied by a slight, albeit significant (20-25%), reduction in cell viability.
Gallardo
G
G
Departamento de Bioquímica, Biología Molecular y Fisiología, Universidad de Las Palmas de Gran Canaria, Spain. gallardo@cicei.ulpgc.es
Tabraue
C
C
Quintana
J
J
López-Blanco
F
F
Cabrera
J
J
Díaz
R
R
Estévez
F
F
Ruiz de Galarreta
C M
CM
Fanjul
L F
LF
Santana
P
P
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002463:Cell Membrane Permeability; D002470:Cell Survival / Q000187:drug effects; D002518:Ceramides / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000378:metabolism / Q000494:pharmacology*; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D051379:Mice; D008934:Mitogens / Q000378:metabolism / Q000494:pharmacology; D051966:Phospholipase C gamma; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism
D002518:Ceramides; D007295:Inositol Phosphates; D007527:Isoenzymes; D008934:Mitogens; D019000:Phosphotyrosine; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D013755:Tetradecanoylphorbol Acetate