Abstract
The c-Myb transcription factor is a major regulator that controls differentiation and proliferation of hematopoietic progenitor cells, which is frequently deregulated in hematological diseases, such as lymphoma and leukemia. Understanding of the mechanisms regulating the transcription of c-myb gene is challenging as it lacks a typical promoter and multiple factors are involved. Our previous studies identified some distal regulatory elements in the upstream regions of c-myb gene in murine myeloid progenitor M1 cells, but the detailed mechanisms still remain unclear. In the present study, we found that a cell differentiation-related DNase1 hypersensitive site is located at a −28k region upstream of c-myb gene and that transcription factors Hoxa9, Meis1 and PU.1 bind to the −28k region. Circular chromosome conformation capture (4C) assay confirmed the interaction between the −28k region and the c-myb promoter, which is supported by the enrichment of CTCF and Cohesin. Our analysis also points to a critical role for Hoxa9 and PU.1 in distal regulation of c-myb expression in murine myeloid cells and cell differentiation. Overexpression of Hoxa9 disrupted the IL-6-induced differentiation of M1 cells and upregulated c-myb expression through binding of the −28k region. Taken together, our results provide an evidence for critical role of the −28k region in distal regulatory mechanism for c-myb gene expression during differentiation of myeloid progenitor M1 cells.
Highlights
Additional studies demonstrate that overexpression or mutations in c-myb can release its oncogenic potential, especially in myeloid cells.[8,9] c-myb transcription is regulated through several mechanisms. c-myb mRNA elongation can be blocked in intron I in a cell-type-dependent manner.[10,11,12] c-Myb can negatively regulate its own expression.[13]
Our previous data indicated the existence of regulatory elements in the c-myb upstream region in M1 cells, based on the enrichment of active histone modifications H3K4me[1] and H3K4me3.26 Here sequencing of DNase I hypersensitive sites (DHS-seq) was used to identify the open sites of the upstream region of c-myb gene in M1 cells
As c-myb expression goes down during differentiation of M1 cells induced by IL-6 treatment, DHS-seq was performed using IL-6-treated M1 cells
Summary
Additional studies demonstrate that overexpression or mutations in c-myb can release its oncogenic potential, especially in myeloid cells.[8,9] c-myb transcription is regulated through several mechanisms. c-myb mRNA elongation can be blocked in intron I in a cell-type-dependent manner.[10,11,12] c-Myb can negatively regulate its own expression.[13]. In our previous work,[26] enrichment of active histone modifications H3K4me[3] and H3K4me[1] (hallmarks of enhancers) was found in the Mml[1], Mml[2] and Mml[3] regions upstream of c-myb gene. These regions are proximal to the 5′ regulatory region of c-myb gene through DNA looping, indicating that these regulatory elements are involved in c-myb expression regulation. The present work provides more details about the long-range regulatory mechanism of c-myb during differentiation of M1 cells
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