Mutagenesis of yeast TFIIIB70 reveals C-terminal residues critical for interaction with TBP and C34

The yeast TFIIIB transcription factor is composed of three components, TBP, TFIIIB90 or B", and TFIIIB70 or BRF. TFIIIB70 is a pivotal component since it interacts with TBP, TFIIIC and RNA polymerase III (pol III). In order to better understand the role of TFIIIB70, we mutagenized extensively t...

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Bibliographic Details
Published in:Journal of molecular biology 1999-05, Vol.288 (4), p.511-520
Main Authors: Andrau, J.C, Sentenac, A, Werner, M
Format: Article
Language:English
Subjects:
Amino Acid Sequence
amino acid sequences
binding sites
DNA-Binding Proteins - metabolism
DNA-directed RNA polymerase
genes
initiation
Molecular Sequence Data
mutagenesis
Mutagenesis, Site-Directed
Protein Binding
regulator genes
RNA Polymerase III - metabolism
Saccharomyces cerevisiae
Saccharomyces cerevisiae - metabolism
Saccharomyces cerevisiae Proteins
Sequence Homology, Amino Acid
taba-binding protein
TATA-Box Binding Protein
transcription (genetics)
Transcription Factor TFIIIB
transcription factors
Transcription Factors - chemistry
Transcription Factors - genetics
Transcription Factors - metabolism
transfer RNA
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Summary:The yeast TFIIIB transcription factor is composed of three components, TBP, TFIIIB90 or B", and TFIIIB70 or BRF. TFIIIB70 is a pivotal component since it interacts with TBP, TFIIIC and RNA polymerase III (pol III). In order to better understand the role of TFIIIB70, we mutagenized extensively three evolutionary conserved motifs of its pol III-specific C-terminal extension. Conditional mutations lying in conserved regions II and III were obtained, some of which altered the interaction with the C34 subunit of pol III and were co-lethal with rpc34 mutations. Two conditional mutations in region II impaired the interaction with TBP and were suppressed by its overexpression. The pattern of suppression of the strongest mutation by overexpression of various mutant TBP, suggested a contact between TBP-R220 and TFIIIB70-D464 residues in vivo. As expected, this TFIIIB70 mutation impaired the assembly of TFIIIB. TFIIIC.DNA complexes and affected in vitro transcription of the SUP4 tRNA gene. Our results underscore the important role of region II of TFIIIB70 in pre-initiation as well as transcription complex assembly via C34 and TBP binding.
ISSN:0022-2836
1089-8638
DOI:10.1006/jmbi.1999.2724