Two independent mechanisms for escaping epidermal growth factor-mediated growth inhibition in epidermal growth factor receptor-hyperproducing human tumor cells.
Human squamous cell carcinoma cell lines often possess increased levels of epidermal growth factor (EGF) receptor. The growth of these EGF receptor-hyperproducing cells is usually inhibited by EGF. To investigate the mechanism of EGF-mediated inhibition of cell growth, variants displaying alternate responses to EGF were isolated from two squamous cell carcinoma lines, NA and Ca9-22; these cell lines possess high numbers of the EGF receptor and an amplified EGF receptor (EGFR) gene. The variants were isolated from NA cells after several cycles of EGF treatment and they have acquired EGF-dependent growth. Scatchard plot analysis revealed a decreased level of EGF receptor in these ER variants as compared with parental NA cells. Southern blot analysis and RNA dot blot analysis demonstrated that the ER variants had lost the amplified EGFR gene. One variant isolated from Ca9-22 cells, CER-1, grew without being affected by EGF. CER-1 cells had higher numbers of EGF receptor than parental Ca9-22 but similar EGFR gene copy number. Flow cytometric analysis indicated an increase in ploidy and cell volume which may give rise to the increase in receptor number per cell. The EGF receptors on both Ca9-22 and CER-1 cells were autophosphorylated upon EGF exposure in a similar manner suggesting no obvious alteration in receptor tyrosine kinase. However, very efficient down-regulation of the EGF receptor occurred in CER-1 cells. These data suggest two independent mechanisms by which EGF receptor-hyperproducing cells escape EGF-mediated growth inhibition: one mechanism is common and involves the loss of the amplified EGFR genes, and another is novel and involves the efficient down-regulation of the cell-surface receptor.
Hirai
M
M
Department of Molecular Biology, Keio University School of Medicine, Tokyo, Japan.
Gamou
S
S
Minoshima
S
S
Shimizu
N
N
D002294:Carcinoma, Squamous Cell; D002455:Cell Division / Q000187:drug effects; D004247:DNA / Q000032:analysis; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D005434:Flow Cytometry; D005784:Gene Amplification; D005786:Gene Expression Regulation; D006801:Humans; D007621:Karyotyping; D009693:Nucleic Acid Hybridization; D010766:Phosphorylation; D011003:Ploidies; D012313:RNA / Q000032:analysis; D014407:Tumor Cells, Cultured
D004815:Epidermal Growth Factor; D012313:RNA; D004247:DNA; D066246:ErbB Receptors