Erythropoietin (Epo) is the major regulator of differentiation, proliferation and survival of erythroid progenitors, but the Epo-induced changes in gene expression that lead to these effects are not fully understood. The aim of this study was to examine how Epo, via activation of phosphatidylinositol 3-kinase (PI3K)/Akt, exerts its role in the development of erythroid progenitors from CD34+ cells, and to identify early Epo target genes in human erythroid progenitors. In CD34+ progenitor cells, Epo alone was able to induce cell cycle progression as demonstrated by upregulation of cyclin D3, E and A leading to hyperphosphorylation of the retinoblastoma protein (RB). These effects were completely counteracted by the PI3K inhibitor LY294002. Furthermore, enforced expression of an activated form of Akt kinase highly augmented Epo-induced erythropoiesis. Fluorescent-activated cell sorting (FACS)-sorted CD34+CD71+CD45RA-GPA- erythroid progenitors stimulated with Epo in the presence or absence of LY294002 were subjected to gene expression profiling. Several novel target genes of Epo were identified, and the majority were regulated in a PI3K-dependent manner, including KIT (CD117) and CDH1 (E-cadherin). FACS analysis of Epo-stimulated erythroid progenitors showed that the increased mRNA expression of KIT and CDH1 was accompanied by an induction of the corresponding proteins CD117 and E-cadherin.