Erythropoietin activates two distinct signaling pathways required for the initiation and the elongation of c-myc

Erythropoietin (Epo) stimulation of erythroid cells results in the activati on of several kinases and a rapid induction of c-myc expression. Protein ki nase C is necessary for Epo up-regulation of c-myc by promoting elongation at the 3'-end of exon 1. PKC epsilon mediates this signal. We now show that Epo triggers two signaling pathways to c-myc. Epo rapidly up-regulated Myc protein in BaF3-EpoR cells. The phosphatidylinositol 3-kinase (PI3K) inhib itor LY294002 blocked Myc up-regulation in a concentration-dependent manner but had no effect on the Epo-induced phosphorylation of ERK1 and ERK2. LY2 94002 also had no effect on Epo up-regulation of c-fos. MEK1 inhibitor PD98 059 blocked both the c-myc and the c-fos responses to Epo. PD98059 and the PKC inhibitor H7 also blocked the phosphorylation of ERK1 and ERK2. PD98059 but not LY294002 inhibited Epo induction of ERK1 and ERK2 phosphorylation in normal erythroid cells. LY294002 blocked transcription of c-myc at exon 1. PD98059 had no effect on transcription from exon 1 but, rather, blocked Epo-induced c-myc elongation at the 3'-end of exon 1. These results identif y two Epo signaling pathways to c-myc, one of which is PI3K-dependent opera ting on transcriptional initiation, whereas the other is mitogen-activated protein kinase-dependent operating on elongation.