scholarly journals Interdependent Interactions between TFIIB, TATA Binding Protein, and DNA

2002 ◽  
Vol 22 (24) ◽  
pp. 8735-8743 ◽  
Author(s):  
Robin M. Buratowski ◽  
Jessica Downs ◽  
Stephen Buratowski
Keyword(s):  
Biochemical Characterization ◽  
Binding Protein ◽  
Tata Binding Protein ◽  
Temperature Sensitive ◽  
Hydrophobic Residues ◽  
Step Model ◽  
Tata Element ◽  
Binding Surface ◽  
Tbp Gene ◽  
Dna Complex

ABSTRACT Temperature-sensitive mutants of TFIIB that are defective for essential interactions were isolated. One mutation (G204D) results in disruption of a protein-protein contact between TFIIB and TATA binding protein (TBP), while the other (K272I) disrupts an interaction between TFIIB and DNA. The TBP gene was mutagenized, and alleles that suppress the slow-growth phenotypes of the TFIIB mutants were isolated. TFIIB with the G204D mutation [TFIIB(G204D)] was suppressed by hydrophobic substitutions at lysine 239 of TBP. These changes led to increased affinity between TBP and TFIIB. TFIIB(K272I) was weakly suppressed by TBP mutants in which K239 was changed to hydrophobic residues. However, this mutant TFIIB was strongly suppressed by conservative substitutions in the DNA binding surface of TBP. Biochemical characterization showed that these TBP mutants had increased affinity for a TATA element. The TBPs with increased affinity could not suppress TFIIB(G204D), leading us to propose a two-step model for the interaction between TFIIB and the TBP-DNA complex.

scholarly journals Contribution of sequences downstream of the TATA element to a protein-DNA complex containing the TATA-binding protein.

1993 ◽  
Vol 13 (4) ◽  
pp. 2593-2603 ◽  
Author(s):  
B A Purnell ◽  
D S Gilmour
Keyword(s):  
Binding Protein ◽  
Core Promoter ◽  
Tata Binding Protein ◽  
Start Site ◽  
Histone H4 ◽  
Normal Core ◽  
Tata Element ◽  
Gel Shift Assay ◽  
Dna Complex ◽  
Made In

A TATA complex that forms on the hsp70 promoter has been found to depend on sequence-specific interactions that occur at the transcription start and regions further downstream. The complex was detected with a gel shift assay and further characterized with interference assays. Antibodies reveal that the TATA-binding protein is in the complex. Interference assays localize specific contacts in the TATA element, the start site, and in a region approximately 25 bp downstream of the start site that contribute to either the assembly or the maintenance of the complex. Contact at the TATA element is made in the minor groove, as has been reported for the recombinant TATA-binding protein. Mutation in the TATA element or the start site of hsp70 causes complex formation to be more strongly dependent on contacts in the +25 region than in the normal core promoter. Examination of the hsp26 and histone H4 genes indicates that similar contacts contribute to the TATA complexes that form on these promoters. The results suggest that specific contacts downstream of the TATA element could play a key role in establishing the transcriptional potential of a gene by contributing to the interaction of the TATA-binding protein.

Contribution of sequences downstream of the TATA element to a protein-DNA complex containing the TATA-binding protein

1993 ◽  
Vol 13 (4) ◽  
pp. 2593-2603
Author(s):  
B A Purnell ◽  
D S Gilmour
Keyword(s):  
Binding Protein ◽  
Core Promoter ◽  
Tata Binding Protein ◽  
Start Site ◽  
Histone H4 ◽  
Normal Core ◽  
Tata Element ◽  
Gel Shift Assay ◽  
Dna Complex ◽  
Made In

A TATA complex that forms on the hsp70 promoter has been found to depend on sequence-specific interactions that occur at the transcription start and regions further downstream. The complex was detected with a gel shift assay and further characterized with interference assays. Antibodies reveal that the TATA-binding protein is in the complex. Interference assays localize specific contacts in the TATA element, the start site, and in a region approximately 25 bp downstream of the start site that contribute to either the assembly or the maintenance of the complex. Contact at the TATA element is made in the minor groove, as has been reported for the recombinant TATA-binding protein. Mutation in the TATA element or the start site of hsp70 causes complex formation to be more strongly dependent on contacts in the +25 region than in the normal core promoter. Examination of the hsp26 and histone H4 genes indicates that similar contacts contribute to the TATA complexes that form on these promoters. The results suggest that specific contacts downstream of the TATA element could play a key role in establishing the transcriptional potential of a gene by contributing to the interaction of the TATA-binding protein.

SPN1, a Conserved Gene Identified by Suppression of a Postrecruitment-Defective Yeast TATA-Binding Protein Mutant

Genetics ◽  
2002 ◽  
Vol 162 (4) ◽  
pp. 1605-1616
Author(s):  
Julie A Fischbeck ◽  
Susan M Kraemer ◽  
Laurie A Stargell
Keyword(s):  
Binding Protein ◽  
Single Point ◽  
Tata Binding Protein ◽  
Single Point Mutation ◽  
Temperature Sensitive ◽  
Yeast Gene ◽  
Tata Element ◽  
Conserved Gene ◽  
Temperature Sensitive Phenotype

Abstract Little is known about TATA-binding protein (TBP) functions after recruitment to the TATA element, although several TBP mutants display postrecruitment defects. Here we describe a genetic screen for suppressors of a postrecruitment-defective TBP allele. Suppression was achieved by a single point mutation in a previously uncharacterized Saccharomyces cerevisiae gene, SPN1 (suppresses postrecruitment functions gene number 1). SPN1 is an essential yeast gene that is highly conserved throughout evolution. The suppressing mutation in SPN1 substitutes an asparagine for an invariant lysine at position 192 (spn1K192N). The spn1K192N strain is able to suppress additional alleles of TBP that possess postrecruitment defects, but not a TBP allele that is postrecruitment competent. In addition, Spn1p does not stably associate with TFIID in vivo. Cells containing the spn1K192N allele exhibit a temperature-sensitive phenotype and some defects in activated transcription, whereas constitutive transcription appears relatively robust in the mutant background. Consistent with an important role in postrecruitment functions, transcription from the CYC1 promoter, which has been shown to be regulated by postrecruitment mechanisms, is enhanced in spn1K192N cells. Moreover, we find that SPN1 is a member of the SPT gene family, further supporting a functional requirement for the SPN1 gene product in transcriptional processes.

scholarly journals Mutations on the DNA-binding surface of TATA-binding protein can specifically impair the response to acidic activators in vivo.

1995 ◽  
Vol 15 (10) ◽  
pp. 5461-5469 ◽  
Author(s):  
M Lee ◽  
K Struhl
Keyword(s):  
Dna Binding ◽  
Protein Interactions ◽  
Convex Surface ◽  
Binding Protein ◽  
Tata Binding Protein ◽  
Protein Protein Interactions ◽  
Activator Proteins ◽  
Tata Element ◽  
Binding Surface

The TATA-binding protein (TBP) contains a concave surface that interacts specifically with TATA promoter elements and a convex surface that mediates protein-protein interactions with general and gene-specific transcription factors. Biochemical experiments suggest that interactions between activator proteins and TBP are important in stimulating transcription by the RNA polymerase II machinery. To gain insight into the role of TBP in mediating transcriptional activation in vivo, we implemented a genetic strategy in Saccharomyces cerevisiae that involved the use of a TBP derivative with altered specificity for TATA elements. By genetically screening a set of TBP mutant libraries that were biased to the convex surface that mediates protein-protein interactions, we identified TBP derivatives that are impaired in the response to three acidic activators (Gcn4, Gal4, and Ace1) but appear normal for constitutive polymerase II transcription. A genetic complementation assay indicates that the activation-defective phenotypes reflect specific functional properties of the TBP derivatives rather than an indirect effect on transcription. Surprisingly, three of the four activation-defective mutants affect residues that directly contact DNA. Moreover, all four mutants are defective for TATA element binding, but they interact normally with an acidic activation domain and TFIIB. In addition, we show that a subset of TBP derivatives with mutations on the DNA-binding surface of TBP are also compromised in their responses to acidic activators in vivo. These observations suggest that interactions at the TBP-TATA element interface can specifically affect the response to acidic activator proteins in vivo.

scholarly journals Biochemical Characterization of the TATA-binding Protein-Gal4 Activation Domain Complex

2000 ◽  
Vol 275 (41) ◽  
pp. 31914-31920 ◽  
Author(s):  
Yueqing Xie ◽  
Carilee Denison ◽  
Sang-Hwa Yang ◽  
David A. Fancy ◽  
Thomas Kodadek
Keyword(s):  
Biochemical Characterization ◽  
Binding Protein ◽  
Tata Binding Protein ◽  
Activation Domain ◽  
Gal4 Activation Domain

scholarly journals The Yeast TAF145 Inhibitory Domain and TFIIA Competitively Bind to TATA-Binding Protein

1998 ◽  
Vol 18 (2) ◽  
pp. 1003-1012 ◽  
Author(s):  
Tetsuro Kokubo ◽  
Mark J. Swanson ◽  
Jun-ichi Nishikawa ◽  
Alan G. Hinnebusch ◽  
Yoshihiro Nakatani
Keyword(s):  
Convex Surface ◽  
Binding Protein ◽  
Tata Binding Protein ◽  
Tata Box ◽  
Temperature Sensitive ◽  
Dna Binding Domain ◽  
Inhibitory Region ◽  
Inhibitory Domain ◽  
Growth Phenotype ◽  
Tata Box Binding Protein

ABSTRACT The Drosophila 230-kDa TFIID subunit (dTAF230) interacts with the DNA binding domain of TATA box-binding protein (TBP) which exists in the same complex. Here, we characterize the inhibitory domain in the yeast TAF145 (yTAF145), which is homologous to dTAF230. Mutation studies show that the N-terminal inhibitory region (residues 10 to 71) can be divided into two subdomains, I (residues 10 to 37) and II (residues 46 to 71). Mutations in either subdomain significantly impair function. Acidic residues in subdomain II are important for the interaction with TBP. In addition, yTAF145 interaction is impaired by mutating the basic residues on the convex surface of TBP, which are crucial for interaction with TFIIA. Consistently, TFIIA and yTAF145 bind competitively to TBP. A deletion of the inhibitory domain of yTAF145 leads to a temperature-sensitive growth phenotype. Importantly, this phenotype is suppressed by overexpression of the TFIIA subunits, indicating that the yTAF145 inhibitory domain is involved in TFIIA function.

scholarly journals A Human TATA Binding Protein-Related Protein with Altered DNA Binding Specificity Inhibits Transcription from Multiple Promoters and Activators

1999 ◽  
Vol 19 (11) ◽  
pp. 7610-7620 ◽  
Author(s):  
Paul A. Moore ◽  
Josef Ozer ◽  
Moreh Salunek ◽  
Gwenael Jan ◽  
Dennis Zerby ◽  
...  
Keyword(s):  
Dna Binding ◽  
Transcriptional Repression ◽  
Transcription Initiation ◽  
Binding Protein ◽  
Tata Binding Protein ◽  
Related Protein ◽  
Multiple Promoters ◽  
Binding Surface

ABSTRACT The TATA binding protein (TBP) plays a central role in eukaryotic and archael transcription initiation. We describe the isolation of a novel 23-kDa human protein that displays 41% identity to TBP and is expressed in most human tissue. Recombinant TBP-related protein (TRP) displayed barely detectable binding to consensus TATA box sequences but bound with slightly higher affinities to nonconsensus TATA sequences. TRP did not substitute for TBP in transcription reactions in vitro. However, addition of TRP potently inhibited basal and activated transcription from multiple promoters in vitro and in vivo. General transcription factors TFIIA and TFIIB bound glutathioneS-transferase–TRP in solution but failed to stimulate TRP binding to DNA. Preincubation of TRP with TFIIA inhibited TBP-TFIIA-DNA complex formation and addition of TFIIA overcame TRP-mediated transcription repression. TRP transcriptional repression activity was specifically reduced by mutations in TRP that disrupt the TFIIA binding surface but not by mutations that disrupt the TFIIB or DNA binding surface of TRP. These results suggest that TFIIA is a primary target of TRP transcription inhibition and that TRP may modulate transcription by a novel mechanism involving the partial mimicry of TBP functions.

scholarly journals INTERACTION OF RECOMBINANT TATA-BINDING PROTEIN WITH MAMMALS GENES PROMOTER TATA

2007 ◽  
Vol 5 (2) ◽  
pp. 44-49 ◽  
Author(s):  
Ludmila K Savinkova ◽  
Irina A Drachkova ◽  
Michail P Ponomarenko ◽  
Marina V Lysova ◽  
Tatiana V Arshinova ◽  
...  
Keyword(s):  
Experimental Data ◽  
Promoter Region ◽  
Binding Protein ◽  
Tata Binding Protein ◽  
Tata Element ◽  
Quantitative Characteristics

Quantitative characteristics of interaction recombinant TATA-binding protein (TBP) with oligonucleotides identical to natural TATA-containing promoter region genes of mammals are received. In particular, new experimental data about the importance guanine in 8-th position of the TATA-element for affinity to TBP are received. The experimental data, testifying that raised maintenance G and С nucleotides in flanks of TATA-element does the contribution to affinity to TBP are received.

scholarly journals A TATA-Binding Protein Mutant Defective for TFIID Complex Formation In Vivo

1999 ◽  
Vol 19 (6) ◽  
pp. 3951-3957 ◽  
Author(s):  
Ryan T. Ranallo ◽  
Kevin Struhl ◽  
Laurie A. Stargell
Keyword(s):  
Binding Protein ◽  
Tata Binding Protein ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Pol Ii ◽  
Pol I ◽  
Pol Iii ◽  
Polymerase I ◽  
Tfiid Complex

ABSTRACT Using an intragenic complementation screen, we have identified a temperature-sensitive TATA-binding protein (TBP) mutant (K151L,K156Y) that is defective for interaction with certain yeast TBP-associated factors (TAFs) at the restrictive temperature. The K151L,K156Y mutant appears to be functional for RNA polymerase I (Pol I) and Pol III transcription, and it is capable of supporting Gal4-activated and Gcn4-activated transcription by Pol II. However, transcription from certain TATA-containing and TATA-less Pol II promoters is reduced at the restrictive temperature. Immunoprecipitation analysis of extracts prepared after culturing cells at the restrictive temperature for 1 h indicates that the K151L,K156Y derivative is severely compromised in its ability to interact with TAF130, TAF90, TAF68/61, and TAF25 while remaining functional for interaction with TAF60 and TAF30. Thus, a TBP mutant that is compromised in its ability to form TFIID can support the response to Gcn4 but is defective for transcription from specific promoters in vivo.

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