Characterization of a myeloid-specific enhancer of the chicken lysozyme gene. Major role for an Ets transcription factor-binding site.
Expression of the lysozyme gene in chicken macrophages is regulated by an enhancer located 2.7 kilobases upstream of the transcription start site. The organization of this enhancer was analyzed by in vitro assays (DNase I footprinting, dimethyl sulfate methylation protection, and band shift assays) and in vivo footprinting experiments. The results show that the enhancer contains four regions (I-IV) of protein-DNA interactions. First, transient gene transfer experiments demonstrate that region II contributes most to the enhancer function. This region contains a recognition site for the macrophage- and B cell-specific transcription factor PU.1, a member of the Ets transcription factor family. This site plays a major role in the enhancer since a mutation of this site abolishes enhancer activity. Second, in competition band shift experiments, we show that the only myeloid-specific complexes in regions I and II with nuclear factors are formed on the PU.1 recognition site by a member of the Ets transcription factor family. Third in in vivo footprinting experiments, the only sign of a protein-DNA interaction is a dimethyl sulfate-hyperreactive guanine 3'-adjacent to the PU.1 recognition site.
Institut für Physiologische Chemie, Universitäts-Krankenhaus Eppendorf, Hamburg, Federal Republic of Germany.
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D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D002460:Cell Line; D002645:Chickens; D004247:DNA; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I; D004742:Enhancer Elements, Genetic; D008264:Macrophages / Q000201:enzymology*; D008745:Methylation; D008969:Molecular Sequence Data; D009113:Muramidase / Q000235:genetics*; D015682:Retroviridae Proteins, Oncogenic; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection
D004268:DNA-Binding Proteins; D015682:Retroviridae Proteins, Oncogenic; D014157:Transcription Factors; C062022:v-Spi-1 protein, Friend spleen focus-forming virus; D004247:DNA; D003850:Deoxyribonuclease I; D009113:Muramidase