p45 NF-E2 regulates expression of thromboxane synthase in megakaryocytes.
Transcription factor p45 NF-E2 is highly expressed in the erythroid and megakaryocytic lineages. Although p45 recognizes regulatory regions of several erythroid genes, mice deficient for this protein display only mild dyserythropoiesis but have abnormal megakaryocytes and lack circulating platelets. A number of megakaryocytic marker genes have been extensively studied, but none of them is regulated by NF-E2. To find target genes for p45 NF-E2 in megakaryopoiesis, we used an in vivo immunoselection assay: genomic fragments bound to p45 NF-E2 in the chromatin of a megakaryocytic cell line were immunoprecipitated with an anti-p45 antiserum and cloned. One of these fragments belongs to the second intron of the thromboxane synthase gene (TXS). We demonstrate that the TXS gene, which is mainly expressed in megakaryocytes, is indeed directly regulated by p45 NF-E2. First, its promoter contains a functional NF-E2 binding site; second, the intronic NF-E2 binding site is located within a chromatin-dependent enhancer element; third, p45-null murine megakaryocytes do not express detectable TXS mRNA, although TXS expression can be detected in other cells. These data, and the structure of the TXS promoter and enhancer, suggest that TXS belongs to a distinct subgroup of genes involved in platelet formation and function.
Deveaux
S
S
INSERM U.91, Hôpital Henri Mondor, 51 Avenue du Maréchal de Lattre, 94010 Créteil, France.
Cohen-Kaminsky
S
S
Shivdasani
R A
RA
Andrews
N C
NC
Filipe
A
A
Kuzniak
I
I
Orkin
S H
SH
Roméo
P H
PH
Mignotte
V
V
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002843:Chromatin / Q000502:physiology; D003001:Cloning, Molecular; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004742:Enhancer Elements, Genetic; D050981:Erythroid-Specific DNA-Binding Factors; D015971:Gene Expression Regulation, Enzymologic; D006410:Hematopoiesis; D006801:Humans; D007438:Introns; D008533:Megakaryocytes / Q000201:enzymology*; D051379:Mice; D008969:Molecular Sequence Data; D050983:NF-E2 Transcription Factor; D050984:NF-E2 Transcription Factor, p45 Subunit; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D013930:Thromboxane-A Synthase / Q000096:biosynthesis* / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D016335:Zinc Fingers
D002843:Chromatin; D017931:DNA Primers; D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D050983:NF-E2 Transcription Factor; D050984:NF-E2 Transcription Factor, p45 Subunit; C494692:NFE2 protein, human; C494693:Nfe2 protein, mouse; D011994:Recombinant Proteins; D014157:Transcription Factors; D013930:Thromboxane-A Synthase