c-myc is an erythropoietin early response gene in normal erythroid cells: evidence for a protein kinase C-mediated signal.
The proto-oncogene c-myc has been identified as an early response gene for erythropoietin (Epo) in transformed murine erythroleukemia cells. Epo activation of c-myc in these cells requires protein kinase C. We now show the fidelity of this signaling pathway in normal erythroid cells isolated from the spleens of phenylhydrazine-treated mice. Mouse spleen cells rich in erythroid progenitors were washed free of endogenous Epo and then incubated in the absence of Epo. Subsequent addition of Epo for 1 hour led to a dramatic elevation of c-myc transcript. Addition of the protein synthesis inhibitor cycloheximide did not prevent the c-myc response, thus identifying c-myc as an Epo early response gene in normal cells. We used this c-myc response as a reporter for signals initiated by the Epo receptor. Using a series of inhibitors with known specificities and established rank-orders of potency for different kinases, we determined that the c-myc response to Epo was blocked with the following rank order: staurosporine much greater than H7 greater than sangivamycin greater than H8. This sequence is identical to that obtained using transformed cells and is diagnostic of a protein kinase C-dependent signal. Because direct activation of protein kinase by phorbol esters does not induce terminal differentiation of normal cells, the pathway to c-myc established by these studies must represent one part of a signal transduction mechanism.
Spangler
R
R
Laboratory for Cell and Molecular Biology, New England Deaconess Hospital, Boston, MA 02215.
Sytkowski
A J
AJ
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D015672:Erythroid Precursor Cells / Q000378:metabolism*; D004921:Erythropoietin / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects*; D016259:Genes, myc; D007546:Isoquinolines / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D010659:Phenylhydrazines / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011741:Pyrimidine Nucleosides / Q000494:pharmacology; D015398:Signal Transduction; D013154:Spleen / Q000166:cytology; D019311:Staurosporine; D014158:Transcription, Genetic
D000470:Alkaloids; D007546:Isoquinolines; D010659:Phenylhydrazines; D010879:Piperazines; D011741:Pyrimidine Nucleosides; C030299:phenylhydrazine; D004921:Erythropoietin; C005699:sangivamycin; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; C053177:N-(2-(methylamino)ethyl)-5-isoquinolinesulfonamide; D011493:Protein Kinase C; D019311:Staurosporine