Erythroid differentiation of mouse erythroleukemia cells results in reorganization of protein-DNA complexes in the mouse beta maj globin promoter but not its distal enhancer.

Reddy, P M; Shen, Che Kun James

doi:10.1128/mcb.13.2.1093

Cited by 32 publications

(28 citation statements)

References 70 publications

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“…First, consistent with the previous genomic footprinting analysis [44], while little, if any, EKLF binding could be detected in the β maj promoter in MEL cells, there was already some, although not much, EKLF-binding in HS2. Upon DMSO induction, both regions became highly enriched in EKLF (Figure 2A).…”

Section: Discussionsupporting

confidence: 76%

“…This is consistent with our earlier genomic-footprinting analysis, in which it was shown that the promoter was empty in uninduced MEL, and it became occupied with factor(s) upon DMSO induction [44]. On the other hand, no apparent enrichment of EKLF binding in the εy and βh1 globin promoter could be detected (data not shown).…”

Section: Eklf Binding In the Murine α-Like Globin Locussupporting

confidence: 81%

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Chromatin-binding in vivo of the erythroid kruppel-like factor, EKLF, in the murine globin loci

et al. 2006

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EKLF is an erythroid-specific, zinc finger-containing transcription factor essential for the activation of the mammalian beta globin gene in erythroid cells of definitive lineage. We have prepared a polyclonal anti-mouse EKLF antibody suitable for Western blotting and immunoprecipitation (IP) qualities, and used it to define the expression patterns of the EKLF protein during mouse erythroid development. We have also used this antibody for the chromatin-immunoprecipitation (ChIP) assay. EKLF was found to bind in vivo at both the mouse beta-major-globin promoter and the HS2 site of beta-LCR in the mouse erythroleukemia cells (MEL) in a DMSO-inducible manner. The DMSO-induced bindings of EKLF as well as three other proteins, namely, RNA polymerase II, acetylated histone H3, and methylated histone H3, were not abolished but significantly lowered in CB3, a MEL-derived cell line with null-expression of p45/NF-E2, an erythroid-enriched factor needed for activation of the mammalian globin loci. Interestingly, binding of EKLF in vivo was also detected in the mouse alpha-like globin locus, at the adult alpha globin promoter and its far upstream regulatory element alpha-MRE (HS26). This study provides direct evidence for EKLF-binding in vivo at the major regulatory elements of the mouse beta-like globin gene clusters the data also have interesting implications with respect to the role of EKLF-chromatin interaction in mammalian globin gene regulation.

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Section: Discussionsupporting

confidence: 76%

Section: Eklf Binding In the Murine α-Like Globin Locussupporting

confidence: 81%

Chromatin-binding in vivo of the erythroid kruppel-like factor, EKLF, in the murine globin loci

et al. 2006

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“…To control for any nonspecific effects of RNA injection, a total of 350 pg mRNA was injected into each blastomere, supplemented with PU.1rs or G1fs as required. (Lane 1 DNA sites (Strauss et al 1992;Reddy and Shen 1993) and several cofactors (Osada et al 1995;Tsang et al 1997;Blobel et al 1998) in MEL cells despite the presence of high levels of PU.1. Furthermore, we found that CBP and FOG binding to GATA-1 was not diminished in the presence of excess PU.1 in vitro (N. Rekhtman and A.I.…”

Section: Mechanism Of Pu1-mediated Repression Of Gata-1mentioning

confidence: 99%

Direct interaction of hematopoietic transcription factors PU.1 and GATA-1: functional antagonism in erythroid cells

Rekhtman

Radparvar

Evans

et al. 1999

Genes & Development

431

406

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Malignant transformation usually inhibits terminal cell differentiation but the precise mechanisms involved are not understood. PU.1 is a hematopoietic-specific Ets family transcription factor that is required for development of some lymphoid and myeloid lineages. PU.1 can also act as an oncoprotein as activation of its expression in erythroid precursors by proviral insertion or transgenesis causes erythroleukemias in mice. Restoration of terminal differentiation in the mouse erythroleukemia (MEL) cells requires a decline in the level of PU.1, indicating that PU.1 can block erythroid differentiation. Here we investigate the mechanism by which PU.1 interferes with erythroid differentiation. We find that PU.1 interacts directly with GATA-1, a zinc finger transcription factor required for erythroid differentiation. Interaction between PU.1 and GATA-1 requires intact DNA-binding domains in both proteins. PU.1 represses GATA-1-mediated transcriptional activation. Both the DNA binding and transactivation domains of PU.1 are required for repression and both domains are also needed to block terminal differentiation in MEL cells. We also show that ectopic expression of PU.1 in Xenopus embryos is sufficient to block erythropoiesis during normal development. Furthermore, introduction of exogenous GATA-1 in both MEL cells and Xenopus embryos and explants relieves the block to erythroid differentiation imposed by PU.1. Our results indicate that the stoichiometry of directly interacting but opposing transcription factors may be a crucial determinant governing processes of normal differentiation and malignant transformation.

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“…However, others did not observe a decrease in GATA-1 DNA binding in such cells (20), and there are several reports indicating that this activity is not increased in MEL cells when PU.1 levels decline as the cells undergo differentiation (1,10,20). Moreover, in vivo footprinting experiments have shown that GATA-1 binding sites in several erythroid cell-specific promoters are occupied in undifferentiated MEL cells that contain high levels of PU.1 and that occupancy does not change as PU.1 levels decline during differentiation (48,56). Chromatin immunoprecipitation (ChIP) experiments also show that GATA-1 is present at erythroid-specific promoters in MEL cells (68).…”

mentioning

confidence: 99%

PU.1 and pRB Interact and Cooperate To Repress GATA-1 and Block Erythroid Differentiation

Rekhtman

Choe

Matushansky

et al. 2003

Molecular and Cellular Biology

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PU.1 and GATA-1 are two hematopoietic specific transcription factors that play key roles in development of the myeloid and erythroid lineages, respectively. The two proteins bind to one another and inhibit each other's function in transcriptional activation and promotion of their respective differentiation programs. This mutual antagonism may be an important aspect of lineage commitment decisions. PU.1 can also act as an oncoprotein since deregulated expression of PU.1 in erythroid precursors causes erythroleukemias in mice. Studies of cultured mouse erythroleukemia cell lines indicate that one aspect of PU.1 function in erythroleukemogenesis is its ability to block erythroid differentiation by repressing GATA-1 (N. Rekhtman, F. Radparvar, T. Evans, and A. I. Skoultchi, Genes Dev. 13:1398-1411, 1999). We have investigated the mechanism of PU.1-mediated repression of GATA-1. We report here that PU.1 binds to GATA-1 on DNA. We localized the repression activity of PU.1 to a small acidic N-terminal domain that interacts with the C pocket of pRB, a well-known transcriptional corepressor. Repression of GATA-1 by PU.1 requires pRB, and pRB colocalizes with PU.1 and GATA-1 at repressed GATA-1 target genes. PU.1 and pRB also cooperate to block erythroid differentiation. Our results suggest that one of the mechanisms by which PU.1 antagonizes GATA-1 is by binding to it at GATA-1 target genes and tethering to these sites a corepressor that blocks transcriptional activity and thereby erythroid differentiation.

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Erythroid differentiation of mouse erythroleukemia cells results in reorganization of protein-DNA complexes in the mouse beta maj globin promoter but not its distal enhancer.

Cited by 32 publications

References 70 publications

Chromatin-binding in vivo of the erythroid kruppel-like factor, EKLF, in the murine globin loci

Chromatin-binding in vivo of the erythroid kruppel-like factor, EKLF, in the murine globin loci

Direct interaction of hematopoietic transcription factors PU.1 and GATA-1: functional antagonism in erythroid cells

PU.1 and pRB Interact and Cooperate To Repress GATA-1 and Block Erythroid Differentiation

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