EGF effects on p53 in MDA-468 human breast cancer cells: implications for G1 arrest.
EGF, in pharmacological concentrations, inhibits cell proliferation of the MDA-468 human breast cancer cell line. Previously, we have demonstrated that this was characterized by a reversible cell cycle arrest at the G1-S boundary, concomitant with downregulation of mRNA levels for p53 (a point mutant, p53(273.His)). Since p53(273.His) is regarded as a gain-of-function mutant and acts to enhance cell proliferation, we hypothesized that the G1 arrest induced by EGF might be mediated by p53(273.His). In this study, we report an EGF-dependent altered conformation as indicated by immunofluorescence, while no significant immediate effects of EGF-treatment on p53(273.His) protein levels and synthesis were observed. These experiments demonstrated a decreased PAb 240 (mutant-specific) reactivity of nuclear p53(273.His) in EGF-treated cells, while that of PAb 1620 (wild-type specific) was enhanced. Staining with PAb 1801 (pan specific), on the other hand, showed little change upon EGF treatment. Further studies indicated a decreased phosphorylation of nuclear p53(273.His) in EGF-treated cells. These EGF-dependent events were detected early enough to be attributed as causative of cell cycle arrest. We suggest that EGF-mediated, phosphorylation-dependent conformational change in nuclear p53(273.His), and in turn altered p53 function, may be responsible for EGF-dependent growth inhibition MDA-468 cells.
Prasad
K A
KA
Terry Fox Cancer Research Laboratories, Division of Basic Medical Sciences, Faculty of Medicine, Memorial University of Newfoundland, St John's, Canada.
Church
J G
JG
D015153:Blotting, Western; D001943:Breast Neoplasms / Q000473:pathology*; D002451:Cell Compartmentation / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D011487:Protein Conformation; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D009687:Nuclear Proteins; D010750:Phosphoproteins; D016159:Tumor Suppressor Protein p53; D004815:Epidermal Growth Factor