DNA binding specificities of Spi-1/PU.1 and Spi-B transcription factors and identification of a Spi-1/Spi-B binding site in the c-fes/c-fps promoter.
Spi-1/PU.1 and Spi-B encode hematopoietic-specific transcription factors that are the most distantly related members of the Ets family. The Ets proteins share a conserved 85 amino acids DNA binding domain, the Ets domain and recognize various DNA target sites around a common core 5'-GGAA/T-3'. The DNA binding specificities of Spi-1 and Spi-B were investigated by using the method of polymerase chain reaction (PCR)-mediated random site selection. The deduced Spi-1 and Spi-B consensus binding sites are very similar suggesting that the functional activities of Spi-1 and Spi-B cannot be distinguished on the basis of their DNA binding specificities. We identified a putative Spi-1/Spi-B binding site in the promoter region of the c-fes/c-fps protooncogene which encodes a tyrosine kinase expressed predominantly in myeloid cells. In vitro translated Spi-1 and Spi-B proteins were capable to bind this site similarly and to activate the c-fes promoter in HeLa transfected cells. We showed that Spi-1 binds the Spi-1/Spi-B binding site of c-fes in HL-60 cells suggesting that Spi-1 may be involved in the regulation of c-fes transcription in myeloid cells. Intriguingly, we detected only Spi-1 binding to this site in the Raji cell line which express both Spi-1 and Spi-B proteins. This suggests that Spi-1 and Spi-B exhibit different DNA binding activities in vivo although they share similar DNA binding specificities in vitro.
Ray-Gallet
D
D
Unité 248 INSERM, Faculté de Médecine Lariboisière Saint-Louis, Paris, France.
Mao
C
C
Tavitian
A
A
Moreau-Gachelin
F
F
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002051:Burkitt Lymphoma / Q000378:metabolism / Q000473:pathology; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008969:Molecular Sequence Data; D009857:Oncogenes / Q000235:genetics / Q000502:physiology*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011817:Rabbits; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000378:metabolism
D004268:DNA-Binding Proteins; D014157:Transcription Factors; C076877:SPIB protein, human; D004247:DNA