Reconstitution of transcription from the human U6 small nuclear RNA promoter with eight recombinant polypeptides and a partially purified RNA polymerase III complex.
The human U6 small nuclear (sn) RNA core promoter consists of a proximal sequence element, which recruits the multisubunit factor SNAP(c), and a TATA box, which recruits the TATA box-binding protein, TBP. In addition to SNAP(c) and TBP, transcription from the human U6 promoter requires two well defined factors. The first is hB", a human homologue of the B" subunit of yeast TFIIIB generally required for transcription of RNA polymerase III genes, and the second is hBRFU, one of two human homologues of the yeast TFIIIB subunit BRF specifically required for transcription of U6-type RNA polymerase III promoters. Here, we have partially purified and characterized a RNA polymerase III complex that can direct transcription from the human U6 promoter when combined with recombinant SNAP(c), recombinant TBP, recombinant hB", and recombinant hBRFU. These results open the way to reconstitution of U6 transcription from entirely defined components.
Department of Microbiology and Graduate Program of Molecular and Cellular Biology, State University of New York at Stony Brook, Stony Brook, New York 11794, USA.
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Chong
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Hu
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Hernandez
D000595:Amino Acid Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D012320:RNA Polymerase III / Q000378:metabolism*; D012342:RNA, Small Nuclear / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D016385:TATA Box; D035181:TATA-Box Binding Protein; D014157:Transcription Factors / Q000378:metabolism
D004268:DNA-Binding Proteins; D012342:RNA, Small Nuclear; D011994:Recombinant Proteins; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; C405486:U6 small nuclear RNA; D012320:RNA Polymerase III