Characterization of a megakaryocyte-specific enhancer of the key hemopoietic transcription factor GATA1.
Specification and differentiation of the megakaryocyte and erythroid lineages from a common bipotential progenitor provides a well studied model to dissect binary cell fate decisions. To understand how the distinct megakaryocyte- and erythroid-specific gene programs arise, we have examined the transcriptional regulation of the megakaryocyte erythroid transcription factor GATA1. Hemopoietic-specific mouse (m)GATA1 expression requires the mGata1 enhancer mHS-3.5. Within mHS-3.5, the 3' 179 bp of mHS-3.5 are required for megakaryocyte but not red cell expression. Here, we show mHS-3.5 binds key hemopoietic transcription factors in vivo and is required to maintain histone acetylation at the mGata1 locus in primary megakaryocytes. Analysis of GATA1-LacZ reporter gene expression in transgenic mice shows that a 25-bp element within the 3'-179 bp in mHS-3.5 is critical for megakaryocyte expression. In vitro three DNA binding activities A, B, and C bind to the core of the 25-bp element, and these binding sites are conserved through evolution. Activity A is the zinc finger transcription factor ZBP89 that also binds to other cis elements in the mGata1 locus. Activity B is of particular interest as it is present in primary megakaryocytes but not red cells. Furthermore, mutation analysis in transgenic mice reveals activity B is required for megakaryocyte-specific enhancer function. Bioinformatic analysis shows sequence corresponding to the binding site for activity B is a previously unrecognized motif, present in the cis elements of the Fli1 gene, another important megakaryocyte-specific transcription factor. In summary, we have identified a motif and a DNA binding activity likely to be important in directing a megakaryocyte gene expression program that is distinct from that in red cells.
Guyot
Boris
B
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Murai
Kasumi
K
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Fujiwara
Yuko
Y
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Valverde-Garduno
Veronica
V
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Hammett
Michele
M
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Wells
Sara
S
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Dear
Neil
N
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Orkin
Stuart H
SH
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Porcher
Catherine
C
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Vyas
Paresh
P
Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom. Electronic address: paresh.vyas@molecular-medicine.oxford.ac.uk.
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D004912:Erythrocytes / Q000378:metabolism; D050982:GATA1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D007438:Introns; D008533:Megakaryocytes / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009928:Organ Specificity; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D051789:Proto-Oncogene Protein c-fli-1 / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism
D004268:DNA-Binding Proteins; C497426:Fli1 protein, mouse; D050982:GATA1 Transcription Factor; C494689:Gata1 protein, mouse; D051789:Proto-Oncogene Protein c-fli-1; D014157:Transcription Factors; C491741:Zfp148 protein, mouse