K562 cells contain a Bcr-Abl chimeric gene and differentiate into various lineages in response to different inducers. We studied the role of the mitogen-activated protein kinase (MAPK) kinase 1 (MEK1)/extracellular signal-regulated kinase (ERK) pathway during the erythroid differentiation of K562 cells induced by tyrosine kinase inhibitors (herbimycin A or STI571), using genetically modified cells (constitutively MEK1-activated K562: K562/MEK1, and inducible ERK-inactivated K562: K562/CL100). Basal expression of glycophorin A was markedly reduced in K562/MEK1 cells compared with that in parental cells, while it was augmented in K562/CL100 cells. Herbimycin A and STI571 differentiated K562 cells accompanying with the transient down-regulated ERK. Moreover, the erythroid differentiation was markedly suppressed in K562/MEK1 cells, and early down-regulation of ERK activity was not observed in these cells. In contrast, the induction of ERK-specific phosphatase in K562/CL100 cells potentiated erythroid differentiation. Once the phosphatase was induced, the initial ERK activity became repressed and its early down-regulation by the inhibition of Bcr-Abl was marked and prolonged. These results demonstrate that the erythroid differentiation of K562 cells induced by herbimycin A or STI571 requires the down-regulation of MEK1/ ERK pathway.