Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P20226 (TATA-binding protein)
 1,297 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Biochemical experiments indicate that the general transcription factor IIB (TFIIB) can interact directly with acidic activation domains and that activators can stimulate transcription by increasing recruitment of TFIIB to promoters. For promoters at which recruitment of TFIIB to promoters is limiting in vivo, one would predict that transcriptional activity should be particularly sensitive to TFIIB mutations that decrease the association of TFIIB with promoter DNA and/or with activation domains; i.e., such TFIIB mutations should exacerbate a limiting step that occurs in wild-type cells. Here, we describe mutations on the DNA-binding surface of TFIIB that severely affect both TATA-binding protein (TBP)-TFIIB-TATA complex formation and interaction with the VP16 activation domain in vitro. These TFIIB mutations affect the stability of the TBP-TFIIB-TATA complex in vivo because they are synthetically lethal in combination with TBP mutants impaired for TFIIB binding. Interestingly, these TFIIB derivatives support viability, and they efficiently respond to Gal4-VP16 and natural acidic activators in different promoter contexts. These results suggest that in vivo, recruitment of TFIIB is not generally a limiting step for acidic activators. However, one TFIIB derivative shows reduced transcription of GAL4, suggesting that TFIIB may be limiting at a subset of promoters in vivo.
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PMID:Transcriptional activation by TFIIB mutants that are severely impaired in interaction with promoter DNA and acidic activation domains. 937 10

Varicella-zoster virus open reading frame 4-encoded protein (IE4) possesses transactivating properties for varicella-zoster virus genes as well as for those of heterologous viruses such as the human immunodeficiency virus type 1 (HIV-1). Mechanisms of HIV-1 LTR (long terminal repeat) transactivation were investigated in HeLa cells transiently transfected with an IE4 expression plasmid and a CAT reporter gene under the control of the HIV-1 LTR. These results demonstrated that IE4-mediated transactivation of the HIV-1 LTR in HeLa cells required transcription factor kappaB (NF-kappaB). Using the gel retardation assay, it was shown that transfection of the IE4 expression vector in HeLa cells was not associated with induction of NF-kappaB under the p50.p65 heterodimeric form and that no direct binding of IE4 to the kappaB sites could be detected. Both Western blot and immunofluorescence analyses suggested that the ability of IE4 to activate transcription through kappaB motives was not connected with its capacity to override the inhibitory activities of IkappaB-alpha or p105. Finally, in vitro protein-protein interactions involving IE4 and basal transcription factors such as TATA-binding protein and transcription factor IIB were carried out. A direct interaction between IE4 and TATA-binding protein or transcription factor IIB components of the basal complex of transcription was evidenced, as well as binding to the p50 and p65 NF-kappaB subunits. Mutagenesis analysis of IE4 indicated that the COOH-terminal cysteine-rich and arginine-rich regions (residues 82-182) were critical for transactivation, whereas the first 81 amino acids appeared dispensable. Moreover, the arginine-rich region is required for the in vitro binding activity, whereas the COOH-terminal end did not appear essential.
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PMID:Activation of the human immunodeficiency virus long terminal repeat by varicella-zoster virus IE4 protein requires nuclear factor-kappaB and involves both the amino-terminal and the carboxyl-terminal cysteine-rich region. 959 2

The nuclear receptor hepatocyte nuclear factor 4 (HNF-4) is an important regulator of several genes involved in diverse metabolic and developmental pathways. Mutations in the HNF-4A gene are responsible for the maturity-onset diabetes of the young type 1. Recently, we showed that the 24 N-terminal residues of HNF-4 function as an acidic transcriptional activator, termed AF-1 (Hadzopoulou-Cladaras, M., Kistanova, E., Evagelopoulou, C., Zeng, S. , Cladaras C., and Ladias, J. A. A. (1997) J. Biol. Chem. 272, 539-550). To identify the critical residues for this activator, we performed an extensive genetic analysis using site-directed mutagenesis. We showed that the aromatic and bulky hydrophobic residues Tyr6, Tyr14, Phe19, Lys10, and Lys17 are essential for AF-1 function. To a lesser degree, five acidic residues are also important for optimal activity. Positional changes of Tyr6 and Tyr14 reduced AF-1 activity, underscoring the importance of primary structure for this activator. Our analysis also indicated that AF-1 is bipartite, consisting of two modules that synergize to activate transcription. More important, AF-1 shares common structural motifs and molecular targets with the activators of the tumor suppressor protein p53 and NF-kappaB-p65, suggesting similar mechanisms of action. Remarkably, AF-1 interacted specifically with multiple transcriptional targets, including the TATA-binding protein; the TATA-binding protein-associated factors TAFII31 and TAFII80; transcription factor IIB; transcription factor IIH-p62; and the coactivators cAMP-responsive element-binding protein-binding protein, ADA2, and PC4. The interaction of AF-1 with proteins that regulate distinct steps of transcription may provide a mechanism for synergistic activation of gene expression by AF-1.
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PMID:Critical structural elements and multitarget protein interactions of the transcriptional activator AF-1 of hepatocyte nuclear factor 4. 979 14

To modulate transcription, regulatory factors communicate with basal transcription factors and/or RNA polymerases in a variety of ways. Previously, it has been reported that RNA polymerase II subunit 5 (RPB5) is one of the targets of hepatitis B virus X protein (HBx) and that both HBx and RPB5 specifically interact with general transcription factor IIB (TFIIB), implying that RPB5 is one of the communicating subunits of RNA polymerase II involved in transcriptional regulation. In this context, we screened for a host protein(s) that interacts with RPB5. By far-Western blot screening, we cloned a novel gene encoding a 508-amino-acid-residue RPB5-binding protein from a HepG2 cDNA library and designated it RPB5-mediating protein (RMP). Expression of RMP mRNA was detected ubiquitously in various tissues. Bacterially expressed recombinant RMP strongly bound RPB5 but neither HBx nor TATA-binding protein in vitro. Endogenous RMP was immunologically detected interacting with assembled RPB5 in RNA polymerase in mammalian cells. The central part of RMP is responsible for RPB5 binding, and the RMP-binding region covers both the TFIIB- and HBx-binding sites of RPB5. Overexpression of RMP, but not mutant RMP lacking the RPB5-binding region, inhibited HBx transactivation of reporters with different HBx-responsive cis elements in transiently transfected cells. The repression by RMP was counteracted by HBx in a dose-dependent manner. Furthermore, RMP has an inhibitory effect on transcriptional activation by VP16 in the absence of HBx. These results suggest that RMP negatively modulates RNA polymerase II function by binding to RPB5 and that HBx counteracts the negative role of RMP on transcription indirectly by interacting with RPB5.
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PMID:RMP, a novel RNA polymerase II subunit 5-interacting protein, counteracts transactivation by hepatitis B virus X protein. 981 40

Binding of the TATA-binding protein (TBP) to the "TATA" sequences present in the promoters of eukaryotic class II genes is the first step in the sequential assembly of transcription pre-initiation complexes. Myriad structural changes, including severe bending of the DNA, accompany TBP-TATA complex formation. A detailed kinetic study has been conducted to elucidate the mechanistic details of TBP binding and DNA bending. The binding of Saccharomyces cerevisiae TBP to the adenovirus major late promoter (AdMLP) was followed in real-time through a range of temperatures and TBP concentrations using fluorescence resonance energy transfer (FRET) and stopped-flow mixing. The results of association and relaxation kinetics and equilibrium binding experiments were analyzed globally to obtain the complete kinetic and energetic profile of the reaction. This analysis reveals a complex mechanism with two intermediate species, with the DNA in the intermediates apparently bent similarly to the DNA in the final complex. TBP binding and DNA bending occur simultaneously through the multiple steps of the reaction. The first and third steps in this sequential process show nearly identical large increases in both enthalpy and entropy, whereas the middle step is highly exothermic and proceeds with a large decrease in entropy. The first intermediate is significantly populated at equilibrium and resembles the final complex both structurally and energetically. It is postulated that both this intermediate and the final complex bind transcription factor IIB in the second step of pol II pre-initiation complex assembly. A consequence of such a reactive intermediate is that the rate of assembly of transcriptionally competent pre-initiation complexes from bi-directionally bound TBP is greatly increased.
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PMID:Intermediate species possessing bent DNA are present along the pathway to formation of a final TBP-TATA complex. 1037 70

The general transcription factor IIB (TFIIB) plays an essential role in transcription of protein-coding genes by eukaryotic RNA polymerase II. We previously identified a yeast TFIIB mutant (R64E) that exhibited increased activity in the formation of stable TATA-binding protein-TFIIB-DNA (DB) complexes in vitro. We report here that the homologous human TFIIB mutant (R53E) also displayed increased activity in DB complex formation in vitro. Biochemical analyses revealed that the increased activity of the R64E mutant in DB complex formation was associated with an altered protease sensitivity of the protein and an enhanced interaction between the N-terminal region and the C-terminal core domain. These results suggest that the intramolecular interaction in yeast TFIIB stabilizes a productive conformation of the protein for the association with promoter-bound TATA-binding protein.
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PMID:An interaction between the N-terminal region and the core domain of yeast TFIIB promotes the formation of TATA-binding protein-TFIIB-DNA complexes. 1043 92

Protein-protein interactions between human heat shock transcription factor 1 (hHSF1) and general transcription factors TFIIA-gamma, TFIIB, TBP, TAF(II)32, and TAF(II)55 and positive coactivator PC4 were characterized in order to identify potential targets of contact in the transcriptional preinitiation complex. These contacts represent one of the final steps in the signal transfer of heat stress to the transcriptional apparatus. TATA-binding protein (TBP) and transcription factor IIB (TFIIB) were identified as major targets for HSF1 transcriptional activation domains AD1 and AD2 based on in vitro interaction assays. TBP showed affinity for AD2 and a fragment containing AD1, while the core domain of TFIIB interacted primarily with the AD1 fragment. Interactions were also detected between full-length HSF1 and the small subunit (gamma) of TFIIA. PC4 interacted weakly with HSF2 and showed even less affinity for HSF1. Coimmunoprecipitation of transiently expressed TBP in HeLa cells demonstrated that HSF1 AD2 and AD1+AD2 are able to bind TBP in vivo. Assays based on transcriptional interference confirmed predictions that both TBP and TFIIB can interact with HSF1 activation domains in HeLa cells. The negative regulatory region (NR) of HSF1 did not interact with any general factors tested in vitro but did bind TFIID in nuclear extracts through contacts that probably involve TATA associated proteins (TAFs). These results suggest a model for transcriptional regulation by HSF1 that involves a shift between formation of dysfunctional TFIID complexes with the NR and transcriptionally competent complexes with the C-terminal activation domains.
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PMID:Potential targets for HSF1 within the preinitiation complex. 1100 81

The hepatocyte nuclear factor-4 (HNF-4) contains two transcription activation domains. One domain, activation function-1 (AF-1), consists of the extreme N-terminal 24 amino acids and functions as a constitutive autonomous activator of transcription. This short transactivator belongs to the class of acidic activators, and it is predicted to adopt an amphipathic alpha-helical structure. Transcriptional analysis of sequential point mutations of the negatively charged residues (Asp and Glu) revealed a stepwise decrease in activity, while mutation of all acidic residues resulted in complete loss of transcriptional activity. Mutations of aromatic and hydrophobic amino acids surrounding the negatively charged residues had a much more profound effect than mutations of acidic amino acids, since even a single mutation of these residues resulted in a dramatic decrease in transactivation, thus demonstrating the importance of hydrophobic residues in AF-1 activity. Like other acidic activators, the AF-1 of HNF-4 binds the transcription factor IIB and the TATA-binding protein directly in vitro. In addition, the cAMP-response-element-binding-protein, a transcriptional adapter involved in the transactivation of a plethora of transcription factors, interacts with the AF-1 of HNF-4 and co-operates in the process of transactivation by HNF-4. The different protein targets of AF-1 suggest that the AF-1 of HNF-4 may be involved in recruiting both general transcription factors and chromatin remodelling proteins during activation of gene expression.
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PMID:The activation function-1 of hepatocyte nuclear factor-4 is an acidic activator that mediates interactions through bulky hydrophobic residues. 1136 95

Transcription of archaeal non-stress genes involves the basal factors TBP and TFB, homologs of the eucaryal TATA-binding protein and transcription factor IIB, respectively. No comparable information exists for the archaeal molecular-chaperone, stress genes hsp70(dnaK), hsp40(dnaJ), and grpE. These do not occur in some archaeal species, but are present in others possibly due to lateral transfer from bacteria, which provides a unique opportunity to study regulation of stress-inducible bacterial genes in organisms with eukaryotic-like transcription machinery. Among the Archaea with the genes, those from the mesophilic methanogen Methanosarcina mazeii are the only ones whose basal (constitutive) and stress-induced transcription patterns have been determined. To continue this work, tbp and tfb were cloned from M. mazeii, sequenced, and the encoded recombinant proteins characterized in solution, separately and in complex with each other and with DNA. M. mazeii TBP ranks among the shortest within Archaea and, contrary to other archaeal TBPs, it lacks tryptophan or an acidic tail at the C terminus and has a basic N-terminal third. M. mazeii TFB is similar in length to archaeal and eucaryal homologs and all have a zinc finger and HTH motifs. Phylogenetically, the archaeal and eucaryal proteins form separate clusters and the M. mazeii molecules are closer to the homologs from Archaeoglobus fulgidus than to any other. Antigenically, M. mazeii TBP and TFB are close to archaeal homologs within each factor family, but the two families are unrelated. The purified recombinant factors were functionally active in a cell-free in vitro transcription system, and were interchangeable with the homologs from Methanococcus thermolithotrophicus. The M. mazeii factors have a similar secondary structure by circular dichroism (CD). The CD spectra changed upon binding to the promoters of the stress genes grpE, dnaK, and dnaJ, with the changes being distinctive for each promoter; in contrast, no effect was produced by the promoter of a non-stress-gene. Factor(s)-DNA modeling predicted that modifications of H bonds are caused by TBP binding, and that these modifications are distinctive for each promoter. It also showed which amino acid residues would contact an extended TATA box with a B recognition element, and evolutionary conservation of the TBP-TFB-DNA complex orientation between two archaeal organisms with widely different optimal temperature for growth (37 and 100 degrees C).
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PMID:The basal transcription factors TBP and TFB from the mesophilic archaeon Methanosarcina mazeii: structure and conformational changes upon interaction with stress-gene promoters. 1139 82

Transcription of the proviral DNA of mouse mammary tumor virus (MMTV) is induced by several classes of hormone-activated steroid receptor proteins. Basal promoter activity in the absence of receptor-mediated activation is selectively repressed by a distal negative regulatory element (dNRE) centered approximately 400 bp upstream of the transcription initiation site. An in vitro transcription system based on synthetic T-free cassette templates was developed to assess MMTV promoter activity, and dNRE-mediated repression was partially reconstituted with this system. Repression was observed with templates in which the dNRE was present in several sequence contexts. The activity of transcription preinitiation complexes formed in vitro in the presence of the dNRE could not be distinguished from that of complexes formed in its absence as assessed by the kinetics of transcript accumulation after addition of nucleoside triphosphates to preformed preinitiation complexes. dNRE-mediated repression in vitro appeared to be the result of decreased efficiency of assembly of functional transcription complexes on the MMTV promoter. However, repression could not be explained by inhibition of assembly of TATA-binding protein or transcription factor IIB into transcription complexes, as neither protein decreased the extent of repression when supplied in excess as a purified recombinant protein.
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PMID:In vitro analysis of transcriptional repression of the mouse mammary tumor virus promoter. 1155 42


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