Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 5 de 5
Filter
Add more filters










Database
Language
Publication year range
1.
Cell Rep ; 2(3): 470-7, 2012 Sep 27.
Article in English | MEDLINE | ID: mdl-22999934

ABSTRACT

The JAK2 tyrosine kinase is a critical mediator of cytokine-induced signaling. It plays a role in the nucleus, where it regulates transcription by phosphorylating histone H3 at tyrosine 41 (H3Y41ph). We used chromatin immunoprecipitation coupled to massively parallel DNA sequencing (ChIP-seq) to define the genome-wide pattern of H3Y41ph in human erythroid leukemia cells. Our results indicate that H3Y41ph is located at three distinct sites: (1) at a subset of active promoters, where it overlaps with H3K4me3, (2) at distal cis-regulatory elements, where it coincides with the binding of STAT5, and (3) throughout the transcribed regions of active, tissue-specific hematopoietic genes. Together, these data extend our understanding of this conserved and essential signaling pathway and provide insight into the mechanisms by which extracellular stimuli may lead to the coordinated regulation of transcription.


Subject(s)
Histones/metabolism , Janus Kinase 2/metabolism , Promoter Regions, Genetic/physiology , STAT5 Transcription Factor/metabolism , Signal Transduction/physiology , Transcription, Genetic/physiology , Cell Line, Tumor , Histones/genetics , Humans , Janus Kinase 2/genetics , Phosphorylation/physiology , STAT5 Transcription Factor/genetics
2.
Nucleic Acids Res ; 38(19): 6363-74, 2010 Oct.
Article in English | MEDLINE | ID: mdl-20525788

ABSTRACT

Ets transcription factors play important roles during the development and maintenance of the haematopoietic system. One such factor, Elf-1 (E74-like factor 1) controls the expression of multiple essential haematopoietic regulators including Scl/Tal1, Lmo2 and PU.1. However, to integrate Elf-1 into the wider regulatory hierarchies controlling haematopoietic development and differentiation, regulatory elements as well as upstream regulators of Elf-1 need to be identified. Here, we have used locus-wide comparative genomic analysis coupled with chromatin immunoprecipitation (ChIP-chip) assays which resulted in the identification of five distinct regulatory regions directing expression of Elf-1. Further, ChIP-chip assays followed by functional validation demonstrated that the key haematopoietic transcription factor PU.1 is a major upstream regulator of Elf-1. Finally, overexpression studies in a well-characterized erythroid differentiation assay from primary murine fetal liver cells demonstrated that Elf-1 downregulation is necessary for terminal erythroid differentiation. Given the known activation of PU.1 by Elf-1 and our newly identified reciprocal activation of Elf-1 by PU.1, identification of an inhibitory role for Elf-1 has significant implications for our understanding of how PU.1 controls myeloid-erythroid differentiation. Our findings therefore not only represent the first report of Elf-1 regulation but also enhance our understanding of the wider regulatory networks that control haematopoiesis.


Subject(s)
DNA-Binding Proteins/genetics , Enhancer Elements, Genetic , Erythroid Cells/cytology , Gene Expression Regulation , Nuclear Proteins/genetics , Promoter Regions, Genetic , Proto-Oncogene Proteins/metabolism , Trans-Activators/metabolism , Transcription Factors/genetics , Acetylation , Animals , Base Sequence , Cell Differentiation , Cell Line , Conserved Sequence , DNA-Binding Proteins/metabolism , Down-Regulation , Erythroid Cells/metabolism , Genetic Loci , Genomics , Histones/metabolism , Mice , Nuclear Proteins/metabolism , Transcription Factors/metabolism , Transcription, Genetic
3.
Blood ; 114(9): 1820-30, 2009 Aug 27.
Article in English | MEDLINE | ID: mdl-19571317

ABSTRACT

The discovery of JAK2V617F as an acquired mutation in the majority of patients with myeloproliferative disorders (MPDs) and the key role of the JAK2-STAT5 signaling cascade in normal hematopoiesis has focused attention on the downstream transcriptional targets of STAT5. Despite evidence of its vital role in normal erythropoiesis and its ability to recapitulate many of the features of myeloid malignancies, including the MPDs, few functionally validated targets of STAT5 have been described. Here we used a combination of comparative genomics and chromatin immunoprecipitation assays to identify ID1 as a novel target of the JAK2-STAT5 signaling axis in erythroid cells. STAT5 binds and transactivates a downstream enhancer of ID1, and ID1 expression levels correlate with the JAK2V617F mutation in both retrovirally transfected fetal liver cells and polycythemia vera patients. Knockdown and overexpression studies in a well-characterized erythroid differentiation assay from primary murine fetal liver cells demonstrated a survival-promoting action of ID1. This hitherto unrecognized function implicates ID1 in the expansion of erythroblasts during terminal differentiation and suggests that ID1 plays an important role in the pathogenesis of polycythemia vera. Furthermore, our findings contribute to an increasing body of evidence implicating ID proteins in a wider range of cellular functions than initially appreciated.


Subject(s)
Erythroid Cells/cytology , Inhibitor of Differentiation Protein 1/metabolism , Janus Kinase 2/metabolism , STAT5 Transcription Factor/metabolism , Animals , Cell Separation , Flow Cytometry , Humans , Janus Kinase 2/genetics , Mice , Mice, Inbred C57BL , Mice, Inbred CBA , Retroviridae/genetics , Signal Transduction
4.
Proc Natl Acad Sci U S A ; 104(45): 17692-7, 2007 Nov 06.
Article in English | MEDLINE | ID: mdl-17962413

ABSTRACT

Conservation of the vertebrate body plan has been attributed to the evolutionary stability of gene-regulatory networks (GRNs). We describe a regulatory circuit made up of Gata2, Fli1, and Scl/Tal1 and their enhancers, Gata2-3, Fli1+12, and Scl+19, that operates during specification of hematopoiesis in the mouse embryo. We show that the Fli1+12 enhancer, like the Gata2-3 and Scl+19 enhancers, targets hematopoietic stem cells (HSCs) and relies on a combination of Ets, Gata, and E-Box motifs. We show that the Gata2-3 enhancer also uses a similar cluster of motifs and that Gata2, Fli1, and Scl are expressed in embryonic day-11.5 dorsal aorta where HSCs originate and in fetal liver where they multiply. The three HSC enhancers in these tissues and in ES cell-derived hemangioblast equivalents are bound by each of these transcription factors (TFs) and form a fully connected triad that constitutes a previously undescribed example of both this network motif in mammalian development and a GRN kernel operating during the specification of a mammalian stem cell.


Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , GATA2 Transcription Factor/metabolism , Hematopoiesis/physiology , Hematopoietic Stem Cells/physiology , Proto-Oncogene Protein c-fli-1/metabolism , Proto-Oncogene Proteins/metabolism , Animals , Basic Helix-Loop-Helix Transcription Factors/chemistry , Basic Helix-Loop-Helix Transcription Factors/genetics , Binding Sites , Blood Vessels/embryology , Blood Vessels/physiology , Embryo, Mammalian/physiology , Enhancer Elements, Genetic , GATA2 Transcription Factor/chemistry , GATA2 Transcription Factor/genetics , Gene Expression Regulation , Mice , Proto-Oncogene Protein c-fli-1/chemistry , Proto-Oncogene Protein c-fli-1/genetics , Proto-Oncogene Proteins/chemistry , Proto-Oncogene Proteins/genetics , T-Cell Acute Lymphocytic Leukemia Protein 1
5.
Int J Cancer ; 120(9): 1964-70, 2007 May 01.
Article in English | MEDLINE | ID: mdl-17266040

ABSTRACT

The clinical diagnosis of cervical neoplasia by spectroscopic methods is potentially a reliable, fast and cost-effective alternative to the conventional smear test. However, it is currently limited by significant inter-patient variation in the spectroscopic properties of the cervix. Characterisation of suitable in vitro models of the spectroscopic changes that take place during neoplastic progression may prove to be a significant step towards the successful development of reliable in vivo systems. In this study, we used organotypic epithelial raft culture as an in vitro model of cervical tissue to analyse changes in the fluorescence properties of surface squamous epithelium that are associated with the development of neoplastic disease. Collagen plugs lined by primary human keratinocytes (PHKs) were used to model the normal cervical epithelium, and plugs lined by cells of the SiHa line were used as a model of neoplastic cervical tissue. Fluorescence emission spectra of these rafts were recorded at excitation wavelengths in the 250-330 nm range, complementing previous work published at longer wavelengths. Normalised, truncated emission spectra were analysed using multivariate principal component analysis. We successfully distinguished between in vitro models of normal and neoplastic cervical tissue and demonstrated a differential effect of acetic acid, which enhances the discrimination of normal from neoplastic tissue. Identification of these differences between in vitro organotypic epithelial rafts may ultimately aid the discrimination of cervical lesions in vivo.


Subject(s)
Precancerous Conditions/diagnosis , Spectrometry, Fluorescence/methods , Uterine Cervical Neoplasms/diagnosis , Cell Line, Tumor , Female , Humans , Phenotype
SELECTION OF CITATIONS
SEARCH DETAIL
...