Identification and regulation of c-Myb target genes in MCF-7 cells.
BACKGROUND
The c-Myb transcription factor regulates differentiation and proliferation in hematopoietic cells, stem cells and epithelial cells. Although oncogenic versions of c-Myb were first associated with leukemias, over expression or rearrangement of the c-myb gene is common in several types of solid tumors, including breast cancers. Expression of the c-myb gene in human breast cancer cells is dependent on estrogen stimulation, but little is known about the activities of the c-Myb protein or what genes it regulates in estrogen-stimulated cells.


METHODS
We used chromatin immunoprecipitation coupled with whole genome promoter tiling microarrays to identify endogenous c-Myb target genes in human MCF-7 breast cancer cells and characterized the activity of c-Myb at a panel of target genes during different stages of estrogen deprivation and stimulation.


RESULTS
By using different antibodies and different growth conditions, the c-Myb protein was found associated with over 10,000 promoters in MCF-7 cells, including many genes that encode cell cycle regulators or transcription factors and more than 60 genes that encode microRNAs. Several previously identified c-Myb target genes were identified, including CCNB1, MYC and CXCR4 and novel targets such as JUN, KLF4, NANOG and SND1. By studying a panel of these targets to validate the results, we found that estradiol stimulation triggered the association of c-Myb with promoters and that association correlated with increased target gene expression. We studied one target gene, CXCR4, in detail, showing that c-Myb associated with the CXCR4 gene promoter and activated a CXCR4 reporter gene in transfection assays.


CONCLUSIONS
Our results show that c-Myb associates with a surprisingly large number of promoters in human cells. The results also suggest that estradiol stimulation leads to large-scale, genome-wide changes in c-Myb activity and subsequent changes in gene expression in human breast cancer cells.
Department of Molecular Genetics and Microbiology, University of New Mexico Health Sciences Center, Albuquerque, NM 87131-0001, USA.
Anita M
AM
Quintana
Fan
F
Liu
John P
JP
O'Rourke
Scott A
SA
Ness
D015153:Blotting, Western; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D004958:Estradiol / Q000494:pharmacology; D004967:Estrogens / Q000494:pharmacology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D000090062:Kruppel-Like Factor 4; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding / Q000187:drug effects; D020598:Proto-Oncogene Proteins c-myb / Q000235:genetics* / Q000378:metabolism*; D034622:RNA Interference; D019718:Receptors, CXCR4 / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
C527276:CXCR4 protein, human; D004967:Estrogens; C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D020598:Proto-Oncogene Proteins c-myb; D019718:Receptors, CXCR4; D004958:Estradiol
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