Dominant negative mutations in yeast TFIID define a bipartite DNA-binding region

Genetic analysis showed that the conserved C-terminal 180 amino acids of yeast TFIID contain all the essential functions for growth of yeast and response to acidic transcriptional activation signals. A genetic screen was used to identify functionally important residues within this C-terminal region. Five dominant TFIID mutations were isolated that had lost the ability to bind DNA. Four of these mutations were single amino acid substitutions in the most N-terminal of two 66-67 amino acid repeats in TFIID. Analogous mutations made in the most C-terminal repeat all failed to bind DNA and inhibited growth of cells, suggesting that the DNA-binding function of TFIID is partitioned between the two repeated regions. Overproduction of wild-type TFIID rescued the dominance of the TFIID mutants, suggesting that the mutant proteins are dominant because they compete with wild-type TFIID for binding to one or more essential transcription factors.

[1]  Robert Tjian,et al.  Mechanism of transcriptional activation by Sp1: Evidence for coactivators , 1990, Cell.

[2]  M. Horikoshi,et al.  Analysis of structure-function relationships of yeast TATA box binding factor TFIID , 1990, Cell.

[3]  P. Sharp,et al.  Yeast TATA-binding protein TFIID binds to TATA elements with both consensus and nonconsensus DNA sequences. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[4]  R. Roeder,et al.  Multiple factors required for accurate initiation of transcription by purified RNA polymerase II. , 1980, The Journal of biological chemistry.

[5]  L. Guarente,et al.  Each of three "TATA elements" specifies a subset of the transcription initiation sites at the CYC-1 promoter of Saccharomyces cerevisiae. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[6]  P. Chambon,et al.  A general transcription factor forms a stable complex with RNA polymerase B (II) , 1987, Cell.

[7]  L. Guarente,et al.  Functional dissection and sequence of yeast HAP1 activator , 1989, Cell.

[8]  C. Ingles,et al.  Direct and selective binding of an acidic transcriptional activation domain to the TATA-box factor TFIID , 1990, Nature.

[9]  D. Reinberg,et al.  Factors involved in specific transcription in mammalian RNA polymerase II. Functional analysis of initiation factors IIA and IID and identification of a new factor operating at sequences downstream of the initiation site. , 1987, The Journal of biological chemistry.

[10]  L. Guarente,et al.  Distinctly regulated tandem upstream activation sites mediate catabolite repression of the CYC1 gene of S. cerevisiae , 1984, Cell.

[11]  L. Guarente,et al.  Identification and characterization of HAP4: a third component of the CCAAT-bound HAP2/HAP3 heteromer. , 1989, Genes & development.

[12]  R. Sikorski,et al.  A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. , 1989, Genetics.

[13]  R. Tjian,et al.  Functional domains and upstream activation properties of cloned human TATA binding protein. , 1990, Science.

[14]  P. Sharp,et al.  Function of a yeast TATA element-binding protein in a mammalian transcription system , 1988, Nature.

[15]  A. Sentenac,et al.  A yeast activity can substitute for the HeLa cell TATA box factor , 1988, Nature.

[16]  F. Winston,et al.  Striking conservation of TFIID in Schizosaccharomyces pombe and Saccharomyces cerevisiae , 1990, Nature.

[17]  R. Sauer,et al.  The structural stability of a protein is an important determinant of its proteolytic susceptibility in Escherichia coli. , 1989, The Journal of biological chemistry.

[18]  C. Kao,et al.  Yeast TATA-box transcription factor gene. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[19]  H. Feldmann,et al.  An element of symmetry in yeast TATA‐box binding protein transcription factor IID , 1990, FEBS letters.

[20]  Thomas A. Kunkel,et al.  Rapid and efficient site-specific mutagenesis without phenotypic selection. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[21]  P. Lieberman,et al.  Cloning of a transcriptionally active human TATA binding factor. , 1990, Science.

[22]  T. Kunkel,et al.  Rapid and efficient site-specific mutagenesis without phenotypic selection. , 1987, Methods in enzymology.

[23]  Masami Horikoshi,et al.  Highly conserved core domain and unique N terminus with presumptive regulatory motifs in a human TATA factor (TFIID) , 1990, Nature.

[24]  S. Berger,et al.  Selective inhibition of activated but not basal transcription by the acidic activation domain of VP16: Evidence for transcriptional adaptors , 1990, Cell.

[25]  P. Sharp,et al.  Five intermediate complexes in transcription initiation by RNA polymerase II , 1989, Cell.

[26]  P. Sharp,et al.  Separation and characterization of factors mediating accurate transcription by RNA polymerase II. , 1982, The Journal of biological chemistry.

[27]  A. Hoffmann,et al.  Arabidopsis thaliana contains two genes for TFIID , 1990, Nature.

[28]  R. Roeder,et al.  Factors involved in specific transcription by human RNA polymerase II: analysis by a rapid and quantitative in vitro assay. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[29]  R. Roeder,et al.  Physical analysis of transcription preinitiation complex assembly on a class II gene promoter. , 1988, Science.

[30]  Yeast promoters and lacZ fusions designed to study expression of cloned genes in yeast. , 1983, Methods in enzymology.

[31]  JAN H. J. Hoeijumakers.  Cryptic initiation sequence revealed , 1990, Nature.

[32]  J. Greenblatt,et al.  Structure and associated DNA-helicase activity of a general transcription initiation factor that binds to RNA polymerase II , 1989, Nature.

[33]  L. Guarente,et al.  Heme regulates transcription of the CYC1 gene of S. cerevisiae via an upstream activation site , 1983, Cell.

[34]  Masami Horikoshi,et al.  Cloning and structure of a yeast gene encoding a general transcription initiation factor TFIID that binds to the TATA box , 1989, Nature.

[35]  L. Guarente,et al.  Yeast HAP1 activator competes with the factor RC2 for binding to the upstream activation site UAS1 of the CYC1 gene , 1987, Cell.

[36]  G. Natsoulis,et al.  5-Fluoroorotic acid as a selective agent in yeast molecular genetics. , 1987, Methods in enzymology.

[37]  K. Nagai.  Cryptic initiation sequence revealed , 1990, Nature.

[38]  F. Winston,et al.  SPT15, the gene encoding the yeast TATA binding factor TFIID, is required for normal transcription initiation in vivo , 1989, Cell.

[39]  P. Chambon,et al.  Cloning of the gene encoding the yeast protein BTF1Y, which can substitute for the human TATA box-binding factor. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[40]  Steven Hahn,et al.  Isolation of the gene encoding the yeast TATA binding protein TFIID: A gene identical to the SPT15 suppressor of Ty element insertions , 1989, Cell.

[41]  J. Wise,et al.  A simple high-efficiency method for random mutagenesis of cloned genes using forced nucleotide misincorporation. , 1990, Gene.

[42]  M. Horikoshi,et al.  Purification of a yeast TATA box-binding protein that exhibits human transcription factor IID activity. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[43]  L. Guarente.  Yeast promoters and lacZ fusions designed to study expression of cloned genes in yeast. , 1983, Methods in enzymology.

[44]  A. Kingsman,et al.  Efficient expression of the Saccharomyces cerevisiae PGK gene depends on an upstream activation sequence but does not require TATA sequences , 1986, Molecular and cellular biology.

[45]  A. Hoffmann,et al.  Cloning of the Schizosaccharomyces pombe TFIID gene reveals a strong conservation of functional domains present in Saccharomyces cerevisiae TFIID. , 1990, Genes & development.

[46]  Roger D. Kornberg,et al.  A novel mediator between activator proteins and the RNA polymerase II transcription apparatus , 1990, Cell.