title: Assembly of human small nuclear RNA gene-specific transcription factor IIIB complex de novo on and off promoter.
abstract: In humans, transcription factor IIIB (TFIIIB)-alpha governs basal transcription from small nuclear RNA genes by RNA polymerase III (pol III). One of the components of this complex, BRFU/TFIIIB50, is specific for these promoters, whereas TATA-binding protein (TBP) and hB" are required for pol III transcription from both gene external and internal promoters. We show that hB" is specifically recruited to a promoter-bound TBP.BRFU complex, which we have previously demonstrated as forming on TATA-containing templates. The N-terminal region of BRFU, containing a zinc ribbon domain, acts as a damper of hB" binding. TBP deactivates this negative mechanism through protein-protein contacts with both BRFU and hB", which may then promote their cooperative binding to form TFIIIB-alpha. In addition, we have identified a GC-rich sequence downstream from the TATA box (the BURE) which, depending on the strength of TATA box, can either enhance BRFU binding to the TBP.DNA complex or hB" association with the BRFU.TBP.DNA complex, and subsequently stimulate pol III transcription. Moreover, mutation of the BURE reduces pol III transcription and induces transcription by RNA polymerase II from the U2 gene promoter carrying a minimal TATA box.
authors.affiliation: Chemical Pathology Unit, Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, United Kingdom. pavel.cabart@path.ox.ac.uk
authors.forename: Pavel
authors.initials: P
authors.lastname: Cabart
authors.forename: Shona
authors.initials: S
authors.lastname: Murphy
mesh_terms: D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism; D017353:Gene Deletion; D006367:HeLa Cells; D006801:Humans; D008954:Models, Biological; D009154:Mutation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012342:RNA, Small Nuclear / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D035181:TATA-Box Binding Protein; D035361:Transcription Factor TFIIIB; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D015032:Zinc / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D012342:RNA, Small Nuclear; D011994:Recombinant Proteins; D035181:TATA-Box Binding Protein; D035361:Transcription Factor TFIIIB; D014157:Transcription Factors; D015032:Zinc