Purification by DNA affinity precipitation of the cellular factors HEB1-p67 and HEB1-p94 which bind specifically to the human T-cell leukemia virus type-I 21 bp enhancer.
Transcription driven by the proviral promoter of the Human T-cell Leukemia Virus type I (HTLV-I) is tightly regulated by the Tax1 transactivator. This viral protein potently induces the enhancer activity of a 21 bp motif repeated three times in the promoter. We have previously shown that this induction results from the binding of Tax1 to this enhancer sequence and that this association is mediated by the cellular factor HEB1. In this paper we report the purification of this factor by chromatography and DNA affinity precipitation. The latter method allowed a rapid and efficient purification which led to the identification of two polypeptides with molecular masses of 94- and 67-kDa, named HEB1-p94 and HEB1-p67, respectively. DMS methylation interference and UV crosslinking experiments indicated that both proteins formed different nucleo-protein complexes, but had the same DNA specificity. Study of the interaction of these two proteins with Tax1 showed that only HEB1-p67 can specifically interact with Tax1.
Laboratoire de Biologie Moléculaire et Cellulaire, UMR 49, Centre National de la Recherche Scientifique, Ecole Normale Supérieure de Lyon, France.
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Lombard-Platet
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Jalinot
D001483:Base Sequence; D002460:Cell Line; D002846:Chromatography, Affinity; D004279:DNA, Viral / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000302:isolation & purification* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004742:Enhancer Elements, Genetic; D016356:Gene Products, tax / Q000378:metabolism; D015368:Human T-lymphotropic virus 1 / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D016356:Gene Products, tax