AbstractThe exact relationship between EGF‐stimulated tyrosine phosphorylation, induction of the cellular proto‐oncogenesc‐mycandc‐fos, and DNA synthesis remains uncertain. Madin‐Darby Canine Kidney (MDCK) cells possess EGF receptor sites with high binding capacity, and in contrast to A431 cells, respond to EGF by increasing DNA synthesis. Following EGF stimulation of intact MDCK cells, there was a rapid and marked increase in the autophosphorylation of the EGF receptor. This was associated with an increase in the tyrosine phosphorylation of a 120 kDa phosphoprotein believed to be an endogenous substrate of this receptor kinase. The ED50for stimulation of phosphorylation of pp120 was −0.05 nM versus 1.0 nM for receptor autophosphorylation, consistent with amplification of signalling at this step in EGF action. Stimulation of DNA synthesis occurred after 12 to 24 hours and revealed even further amplification with an ED50of about 0.1 nM. Intermediate between these events was a time‐dependent activation ofc‐fosandc‐mycgene expression. However, the ED50for these processes was ã10 nM, indicating a relatively lower sensitivity of EGF for stimulation of proto‐oncogene expression. Tyrphostin (RG 50864), a compound reported to inhibit specifically the EGF receptor kinase, completely blocked EGF stimulation of proto‐oncogene induction. Interestingly, under the same experimental conditions, EGF receptor autophosphorylation was decreased only 60. These data, along with the dose‐response studies, indicate that proto‐oncogene induction requires near maximal stimulation of EGF receptor autophosphorylation. They also suggest that, in MDCK cells, the EGF dependent induction of thec‐fosandc‐mycgenes is not strictly correlated to the extent of EGF receptor autophosphorylation or EGF‐stimulated DNA synthesis, and that EGF stimulation of DNA synthesis likely involves additional
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