The transcription factor GATA-1 is expressed in early hematopoietic progenitors and specifically down-regulated in myelomonocytic cells during lineage determination. Our earlier observation that the differentiation of Myb-Ets-transformed chicken hematopoietic progenitors into myeloblasts likewise involves a GATA-1 down-regulation, whereas expression is maintained in erythroid, thrombocytic, and eosinophilic derivatives, prompted us to study the effect of forced GATA-1 expression in Myb-Ets-transformed myeloblasts. We found that the factor rapidly suppresses myelomonocytic markers and induces a reprogramming of myeloblasts into cells resembling either transformed eosinophils or thromboblasts. In addition, we observed a correlation between the level of GATA-1 expression and the phenotype of the cell, intermediate levels of the factor being expressed by eosinophils and high levels by thromboblasts, suggesting a dosage effect of the factor. GATA-1 can also induce the formation of erythroblasts when expressed in a myelomonocytic cell line transformed with a Myb-Ets mutant containing a lesion in Ets. These cells mature into erythrocytes following temperature-inactivation of the Ets protein. Finally, the factor can reprogram a v-Myc-transformed macrophage cell line into myeloblasts, eosinophils, and erythroblasts, showing that the effects of GATA-1 are not limited to Myb-Ets-transformed myeloblasts. Our results suggest that GATA-1 is a lineage-determining transcription factor in transformed hematopoietic cells, which not only activates lineage-specific genetic programs but also suppresses myelomonocytic differentiation. They also point to a high degree of plasticity of transformed hematopoietic cells.