Regulation of osteosarcoma EGF receptor affinity by phorbol ester and cyclic AMP.
We studied the binding and degradation of 125I-labeled epidermal growth factor (EGF) by UMR-106 osteosarcoma cells and the regulation of EGF receptor affinity for EGF by the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and by treatments that raise intracellular levels of cyclic AMP. Cell surface binding of [125I]EGF to A431 cells reached a plateau after a 30 minute incubation at 37 degrees C but was undetectable in UMR-106 cells. Degradation of [125I]EGF proceeded at a 50-fold higher rate in A431 cells on a per cell basis, but receptor-bound [125I]EGF was internalized and degraded at a 3.5-fold higher rate by UMR-106 cells on a per receptor basis. At 4 degrees C, [125I]EGF labeled a single class of surface binding sites in the UMR-106 cell. Treatment with TPA at 37 degrees C reduced subsequent cell surface binding of [125I]EGF at 4 degrees C a maximum of 80% with an IC50 of 1.25 ng/ml. Maximal TPA reduction of [125I]EGF binding was observed within 5-15 minutes and was due to a reduction in the affinity of cell surface receptors of [125I]EGF without a change in receptor density. Pretreatment of the cells for 4 h with 30 microM forskolin, 1 mM isobutylmethylxanthine (IBMX) plus 30 microM forskolin, or 1 mM IBMX plus 100 ng/ml parathyroid hormone (PTH) attenuated the loss in [125I]EGF binding caused by a subsequent dose of 10 ng/ml of TPA by 17% (p less than 0.0005), 39% (p less than 0.0002), and 35% (p less than 0.002), respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Borst
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Emory University Department of Medicine, Atlanta, GA.
Catherwood
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D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism / Q000502:physiology*; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007457:Iodine Radioisotopes; D012516:Osteosarcoma / Q000503:physiopathology*; D010281:Parathyroid Hormone / Q000502:physiology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D007457:Iodine Radioisotopes; D010281:Parathyroid Hormone; D005576:Colforsin; D004815:Epidermal Growth Factor; D000242:Cyclic AMP; D066246:ErbB Receptors; D013755:Tetradecanoylphorbol Acetate; D015056:1-Methyl-3-isobutylxanthine