Isolation of coactivators associated with the TATA-binding protein that mediate transcriptional activation.
A key step in the regulation of transcription involves interactions between promoter-selective factors and various components of the transcriptional apparatus. Here we report the requirements for transcriptional activation directed by NTF-1, a developmentally regulated transcription factor in Drosophila. Reconstituted transcription with fractionated Drosophila basal factors reveals that activation by NTF-1 requires factors present in the endogenous TFIID fraction that are distinct from the purified TATA-binding protein (TBP). Glycerol gradient sedimentation and immunoprecipitation analyses indicate that TFIID is a multiprotein complex containing TBP and at least six tightly bound TBP-associated factors (TAFs). Preparations of TBP lacking TAFs after fractionation with denaturants no longer support activation by NTF-1 but retain basal level activity. Addition of immunopurified and renatured TAFs to free TBP restores the ability of NTF-1 to activate transcription without influencing basal transcription. These results suggest that one or more of the TAF polypeptides confer coactivator function.
Dynlacht
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Howard Hughes Medical Institute, Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Hoey
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Tjian
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D000818:Animals; D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000502:physiology*; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000235:genetics*; D005786:Gene Expression Regulation; D006358:Hot Temperature; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011994:Recombinant Proteins; D012045:Regulatory Sequences, Nucleic Acid; D035362:Transcription Factor TFIID; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology*; D014158:Transcription, Genetic; D014508:Urea / Q000737:chemistry
D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D046911:Macromolecular Substances; D046912:Multiprotein Complexes; D009841:Oligonucleotides; D011994:Recombinant Proteins; D035362:Transcription Factor TFIID; D014157:Transcription Factors; C062330:grh protein, Drosophila; D014508:Urea