HUMAN ERYTHROPOIETIN RECEPTOR INCREASES GATA-2 AND BCL-X(L) BY A PROTEIN-KINASE-C DEPENDENT PATHWAY IN HUMAN ERYTHROPOIETIN-DEPENDENT CELL-LINE AS-E2

Erythropoietin (Epo) is a cytokine known to stimulate proliferation an d differentiation of erythroid cells. However, recent gene disruption experiments demonstrated that Epo receptor signaling is not an obligat ory step in erythroid differentiation. Here, we describe the role of E po in proliferation, terminal differentiation, and apoptosis in a nove l human Epo-dependent cell line, AS-EP. Upon withdrawal of Epo, the ce lls ceased to proliferate and underwent apoptotic death. Accompanying this cell death, an increase in the number of hemoglobin-positive cell s of approximately 2-fold was observed. This was associated with immed iate up-regulation of the GATA-1:GATA-2 ratio and down-regulation of B cl-x(L). Treatment with Epo or 12-O-tetradecanoyl-phorbol-13-acetate ( TPA) up-regulated expression of GATA-2 and Bcl-x(L), and these elevati ons were inhibited by inhibitors of protein kinase C (PKC), H7 and H8. HA1004, a structural analogue of H7 but a poor inhibitor of PKC, had no inhibitory effect. Therefore, in AS-E2 cells, it is likely that Epo plays a role in (a) proliferation, (b) inhibition of differentiation, and (c) survival, by maintaining GATA-2 and Bcl-x(L) expression throu gh activation of PKC.