Recent evidence has demonstrated an appreciable expression of metallothionein (MT) in erythrocytes. However, the induction of the MT protein by hematopoietic growth factors and its subsequent functional significance on clonal expansion or differentiation of erythroid progenitor cells remain elusive. We therefore examined the effects of growth factors erythropoietin (EPO), granulocyte-monocyte colony-stimulating factor (GM-CSF), and interleukin-3 (IL-3) on MT gene expression in erythroid progenitor K562 cell line. EPO, but not IL-3 or GM-CSF, induced a 3-fold increase in MT transcripts in K562 cells. MT induction was associated with EPO-induced cellular proliferation, suggesting a specific role for MT induction by EPO in erythroid progenitor cells. However, EPO- or sodium butyrate-induced differentiation as monitored by hemoglobin formation was inhibited in K562 cells stably transfected with an expression vector containing human MT-IIA gene. This inhibition of differentiation was partially reversed in these cells by an MT antisense phosphorothioate oligonucleotide. Furthermore, the MT-induced inhibition of differentiation was associated with downregulation of EPO receptor transcripts in K562 cells. These data suggest that, among growth factors required for erythropoiesis, EPO is a potent inducer of MT, and that MT may plays a significant role in EPO-induced proliferation, but not in the erythroid-specific differentiation of the progenitor cells.