Abstract
Backgroundc-Myc has been proposed as a putative target gene of signal transducer and activator of transcription 5 (STAT5). No functional STAT5 binding site has been identified so far within the c-Myc gene locus, therefore a direct transcriptional regulation by STAT5 remains uncertain. c-Myc super-enhancer, located 1.7 Mb downstream of the c-Myc gene locus, was recently reported as essential for the regulation of c-Myc gene expression by hematopoietic transcription factors and bromodomain and extra-terminal (BET) proteins and for leukemia maintenance. c-Myc super-enhancer is composed of five regulatory regions (E1–E5) which recruit transcription and chromatin-associated factors, mediating chromatin looping and interaction with the c-Myc promoter.ResultsWe now show that STAT5 strongly binds to c-Myc super-enhancer regions E3 and E4, both in normal and transformed Ba/F3 cells. We also found that the BET protein bromodomain-containing protein 2 (BRD2), a co-factor of STAT5, co-localizes with STAT5 at E3/E4 in Ba/F3 cells transformed by the constitutively active STAT5-1*6 mutant, but not in non-transformed Ba/F3 cells. BRD2 binding at E3/E4 coincides with c-Myc transcriptional activation and is lost upon treatment with deacetylase and BET inhibitors, both of which inhibit STAT5 transcriptional activity and c-Myc gene expression.ConclusionsOur data suggest that constitutive STAT5 binding to c-Myc super-enhancer might contribute to BRD2 maintenance and thus allow sustained expression of c-Myc in Ba/F3 cells transformed by STAT5-1*6.Electronic supplementary materialThe online version of this article (doi:10.1186/s12867-016-0063-y) contains supplementary material, which is available to authorized users.
Highlights
Background cMyc is a master regulator of essential biological processes such as cell proliferation, survival, differentiation, metabolism, angiogenesis and pluripotency establishment and maintenance [1, 2]. c-Myc is found overexpressed in most human cancers and is a hallmark of tumor initiation and maintenance [3, 4]
Whereas signal transducer and activator of transcription 5 (STAT5) phosphorylation, binding to DNA and transcriptional activity are induced by IL-3 in mouse pro-B IL-3-dependent cell line (Ba/F3) and Ba/ F3-WT cells, they are constitutive in Ba/F3-1*6 cells in the absence of IL-3 (Fig. 1a, b and [11, 12, 29, 31, 35]), mimicking the situation found in cancer cells
bromodomain-containing protein 2 (BRD2) co‐localizes with constitutively active STAT5 at c‐Myc super‐enhancer Given that bromodomain-containing protein and 4 respectively (BRD4) is recruited at c-Myc super-enhancer in hematopoietic and leukemic cells [5, 6], and since we recently showed the implication of BRD2 in STAT5mediated transcription in Ba/F3 and Ba/F3-1*6 cells [31], we investigated the recruitment of BRD2 along the c-Myc gene and super-enhancer by chromatin immunoprecipitation (Fig. 4)
Summary
Background cMyc is a master regulator of essential biological processes such as cell proliferation, survival, differentiation, metabolism, angiogenesis and pluripotency establishment and maintenance [1, 2]. c-Myc is found overexpressed in most human cancers and is a hallmark of tumor initiation and maintenance [3, 4]. In contrast to most classical STAT5 target genes (e.g., Cis, Osm, Spi2.1, ...) which display STAT5 responsive elements within their promoter region [13], functional STAT5 binding sites have been identified outside the promoter regions of Bclx and Id-1, notably within Bcl-x first intron [36, 38] and within Id-1 enhancer located several kb downstream of the Id-1 gene [37]. These observations raise the possibility that regulation of c-Myc expression by STAT5 might be likewise unconventional, possibly involving distal elements. In support of this proposition, we previously attempted and failed to detect STAT5 binding along the c-Myc gene [13], including at GAS elements present in its promoter and known to mediate transcriptional response to other STAT family members [39,40,41]
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