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PMID: 1461721 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Cofractionation of the TATA-binding protein with the RNA polymerase III transcription factor TFIIIB.

Nucleic acids research ·Vol. 20 ·No. 22 ·1992-11-25 ·Pages 5889-98

Simmen KA, Bernués J, Lewis JD, Mattaj IW

Abstract

We have investigated the requirement for TBP (TATA-binding protein) in transcription mediated by RNA polymerase III (pol III) in fractionated HeLa cell extracts. Two activities, TFIIIB and TFIIIC, found in phosphocellulose fractions PC B and PC C respectively, have been defined as necessary and sufficient, with pol III, for in vitro transcription of tRNA genes. Depletion of TBP from PC B, using antibodies raised against human TBP, is shown to inhibit the pol III transcriptional activity of the fraction. Furthermore, TBP is present in fractions with human TFIIIB activity, and a proportion of TBP cofractionates with TFIIIB over four chromatographic purification steps. TFIIIB fractions are capable of supplying TBP in the form necessary for pol III transcription, and cannot be substituted by fractions containing other TBP complexes or TBP alone. The use of a 5S RNA gene and two tRNA templates supports the general relevance of our findings for pol III gene transcription. Purified TFIIIB activity can also support pol II-mediated transcription, and is found in a complex of approximately 230kD, suggesting that TFIIIB may be the same as the previously characterized B-TFIID complex (1,2). We suggest that transcription by the three RNA polymerases is mediated by distinct TBP-TAF complexes: SL1 and D-TFIID for pol I and pol II respectively, and TFIIIB for pol III.

MeSH Terms
Chemical Fractionation Chromatography, Ion Exchange DNA-Binding Proteins/metabolism HeLa Cells Humans Immunologic Techniques Promoter Regions, Genetic RNA Polymerase III/metabolism TATA Box TATA-Box Binding Protein Transcription Factor TFIID Transcription Factor TFIIIB Transcription Factors/metabolism Transcription, Genetic
Chemicals
DNA-Binding Proteins TATA-Box Binding Protein Transcription Factor TFIID Transcription Factor TFIIIB Transcription Factors RNA Polymerase III
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Simmen K A
European Molecular Biology Laboratory, Heidelberg, Germany.
Bernués J
Lewis J D
Mattaj I W
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/20.22.5889
Published
1992-11-25
Pages
5889-98
Language
English
Region
England
NLM ID
0411011
PMCID
PMC334451
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/20.22.5889 PMC Full Text View journal details
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