Biochemical and genetic characterization of a yeast TFIID mutant that alters transcription in vivo and DNA binding in vitro.
Open Access
- 1 May 1992
- journal article
- Published by Taylor & Francis in Molecular and Cellular Biology
- Vol. 12 (5), 2372-2382
- https://doi.org/10.1128/mcb.12.5.2372
Abstract
A mutation in the gene that encodes Saccharomyces cerevisiae TFIID (SPT15), which was isolated in a selection for mutations that alter transcription in vivo, changes a single amino acid in a highly conserved region of the second direct repeat in TFIID. Among eight independent spt15 mutations, seven cause this same amino acid change, Leu-205 to Phe. The mutant TFIID protein (L205F) binds with greater affinity than that of wild-type TFIID to at least two nonconsensus TATA sites in vitro, showing that the mutant protein has altered DNA binding specificity. Site-directed mutations that change Leu-205 to five different amino acids cause five different phenotypes, demonstrating the importance of this amino acid in vivo. Virtually identical phenotypes were observed when the same amino acid changes were made at the analogous position, Leu-114, in the first repeat of TFIID. Analysis of these mutations and additional mutations in the most conserved regions of the repeats, in conjunction with our DNA binding results, suggests that these regions of the repeats play equivalent roles in TFIID function, possibly in TATA box recognition.Keywords
This publication has 59 references indexed in Scilit:
- Activation domains of stably bound GAL4 derivatives alleviate repression of promoters by nucleosomesCell, 1991
- A novel mediator between activator proteins and the RNA polymerase II transcription apparatusCell, 1990
- Mechanism of transcriptional activation by Sp1: Evidence for coactivatorsCell, 1990
- Analysis of structure-function relationships of yeast TATA box binding factor TFIIDCell, 1990
- An element of symmetry in yeast TATA-box binding protein transcription factor IIDFEBS Letters, 1990
- Cryptic initiation sequence revealedNature, 1990
- A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformaion of Escherichia coliGene, 1987
- Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genesJournal of Molecular Biology, 1986
- Interaction of a gene-specific transcription factor with the adenovirus major late promoter upstream of the TATA box regionCell, 1985
- [12] One-step gene disruption in yeastMethods in Enzymology, 1983