Abstract
The c-myb gene is primarily expressed in hematopoietic cells, and it is overexpressed in many leukemias. The regulation of its expression is of critical importance in hematopoietic cells. We identified the major positive regulatory sites in the 5'-flanking sequence of the human c-myb gene, and we found that the positive regulators differed in cells of different lineages. In the Molt-4 T-cell line, two Ets-like binding sites were required for the expression of c-myb. The 5' site played a minor role in the regulation of c-myb expression, and we demonstrated that a protein of 67 kDa bound to this site. Antibodies against Ets proteins showed no cross-reactivity with this protein. We showed that Ets-1 bound to the 3'-regulatory site in the c-myb promoter by electrophoretic mobility shift assay and antibody studies. Both of these Ets-like binding sites were nonfunctional in the DHL-9 B-cell line and the K562 myeloid cell line. We identified a novel transcription factor of 50.5 kDa that was required for expression of c-myb in these cell lines.
Highlights
C-Myb plays a central role in the regulation of hematopoietic cell development, and the control of its expression is critically important
Since c-myb is expressed in all hematopoietic cells, we wished to determine whether the same promoter regions were involved in positive regulation in B- and myeloid cell lines
We have identified the major positive regulatory regions of the c-myb promoter in T, B, and myeloid cell lines
Summary
C-Myb plays a central role in the regulation of hematopoietic cell development, and the control of its expression is critically important. An important mechanism for regulation of murine c-myb expression is a block to transcription elongation within the first intron of the c-myb locus, recognized as a pause site [13,14,15]. In cotransfection studies c-Myb is involved in positive autoregulation of the c-myb gene in hamster fibroblasts [17]. We have shown that two c-Myb binding sites function as negative regulators of c-myb expression in T-cell lines [18]. Further studies of the regulation of expression of c-myb have shown that c-Jun and JunD are positive regulators of the c-myb promoter in hamster fibroblasts. Mutation of the two guanines in the core binding area results in a loss of 95% of the promoter activity of the T-cell receptor ␣ gene [25]. In the T-cell line, Molt-4, we find that two Ets-like binding sites are required for c-myb expression. These Ets binding sites are nonfunctional in both a B- and myeloid cell line, and we find that a novel transcription factor is required for c-myb expression in these cell lines
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