Transgene Insertion into the Proximity of the c-myb Gene Disrupts Erythroid-Megakaryocytic Lineage Bifurcation.

The nuclear proto-oncogene c-myb plays crucial roles in the growth, survival and differentiation of hematopoietic cells. We established three lines of erythropoietin receptor transgenic mice and found that one of them exhibited anemia, thrombocythemia and splenomegaly. These abnormalities were independent of the function of the transgenic erythropoietin receptor, and were observed exclusively in mice harboring the transgene homozygously, suggesting transgenic disruption of a certain gene. The transgene was inserted 77-kb upstream of the c-myb gene and c-Myb expression was markedly decreased in megakaryocyte-erythrocyte lineage-restricted progenitors (MEPs) of the homozygous mutant mice. In the bone marrows and spleens of the mutant mice, megakaryocytes were increased, but erythroid progenitors were decreased. These abnormalities were reproducible in vitro in a co-culture assay of MEPs with OP9 cells, but eliminated by the retroviral expression of c-Myb in MEPs. The erythroid-megakaryocytic abnormalities were reconstituted in mice in vivo through the transplantation of mutant mouse bone marrow cells. To better understand the transcriptional program that accompanies the decline of c-myb gene expression, we performed DNA microarray analysis of MEPs. We identified 74 genes that are upregulated and 36 genes are downregulated in the homozygous mutant mice. Of these genes, expression levels 10 genes are actually changed in bone marrow cells of the homozygous mutant mice, and harbor c-Myb recognition elements in the upstream region. These results thus demonstrate that the transgene insertion into the c-myb gene far upstream regulatory region affects the gene expression at the stage of MEPs, leading to an imbalance between erythroid and megakaryocytic cells, and suggest that c-Myb is an essential regulator of the erythroid-megakaryocytic lineage bifurcation. Elucidation of transcription network based on the c-myb gene in erythroid and megakaryocytic lineages will take a new turn of lineage specific regulation.

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