The leukemia oncoprotein BCR-ABL complements the erythropoietin receptor

The myeloproliferative disorder CML results from the formation of a fusion P21OBCR-ABL oncoprotein in a primitive hematopoietic stem cell and is characterized by an expansion of the myeloid compartment. Despite the cytokine-independent growth conferred by BCR-ABL expression in myeloid cell lines, myeloid progenitors from CML patients require growth factors to proliferate and differentiate in vitro. In contrast, CML erythroid progenitors exhibit EPO-independent growth in vitro, a property common to erythroid progenitors in myeloproliferative disorders. We used high liter retroviral supernatants to express the P210BCR-ABL protein in the physiological context of EPO-R"'" mouse fetal liver cells, to address the degree of contribution of BCR-ABL (P210) to the EPO-independent erythroid differentiation. EPO-R~'~ mice die from severe anemia, around day 13-15 of embryogenesis, due to the absence of differentiation of CFU-E to mature red cells. However, mature (CFU-E) and more primitive (BFU-E) erythroid progenitors are found in normal numbers in EPO-R"'" fetal liver cells transduced with the EPO-R and cultured in the presence of Erythropoietin (EPO) alone in a CFU-E assay or EPO+Steel Factor (SF) in a BFU-E assay. The expression of P21QBCR-ABL in EPO-R-'' fetal liver cells rescued over 80% of CFU-E progenitors and ~ 25% of BFU-E in the absence of any exogenously added growth factor (n=4). The addition of SF rescued the remaining BCR-ABL transduced BFU-Es. All EPO-independent progenitors contained the BCR-ABL gene as detected by genomic PCR on individual erythroid colonies. Importantly, the colony size, morphology, and degree of hemoglobinization of BCR-ABL transduced erythroid colonies, even in the absence of SF, were comparable to those reconstituted by the EPO-R in the presence of EPO, as determined by benzidine staining and cytospin analysis. Using mutants of BCR-ABL we have demonstrated that rescue of erythroid differentiation requires a functional tyrosine kinase domain but domains implicated in either Ras or Myc activation are dispensable. Our findings demonstrate that the cytoplasmic oncoprotein BCR-ABL can complement for signals typically provided by the EPO-R in erythroid proliferation and differentiation. In addition, our system provides a paradigm for the characterization of genes involved in other myeloproliferative disorders.