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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The binding of TATA-binding protein (TBP) to the TATA element is the first step in the initiation of RNA polymerase II transcription from many promoters in vitro. It has been proposed that upstream activator proteins stimulate transcription by recruiting TBP to the promoter, thus facilitating the assembly of a transcription complex. However, the role of activator proteins acting at this step to stimulate transcription in vivo remains largely speculative. To test whether recruitment of TBP to the promoter is sufficient for transcriptional activation in vivo, we constructed a hybrid protein containing TBP of the yeast Saccharomyces cerevisiae fused to the DNA-binding domain of GAL4. Our results show that TBP recruited by the GAL4 DNA-binding domain to promoters bearing a GAL4-binding site can interact with the TATA element and direct high levels of transcription. This finding indicates that binding of TBP to promoters in S. cerevisiae is a major rate-limiting step accelerated by upstream activator proteins.
Mol Cell Biol 1995 Oct
PMID:Recruiting TATA-binding protein to a promoter: transcriptional activation without an upstream activator. 756 28

The core promoter compositions of mammalian protein-coding genes are highly variable; some contain TATA boxes, some contain initiator (Inr) elements, and others contain both or neither of these basal elements. The underlying reason for this heterogeneity remains a mystery, as recent studies have suggested that TATA-containing and Inr-containing core promoters direct transcription initiation by similar mechanisms and respond similarly to a wide variety of upstream activators. To analyze in greater detail the influence of core promoter structure on transcriptional activation, we compared activation by GAL4-VP16 and Sp1 through synthetic core promoters containing a TATA box, an Inr, or both TATA and Inr. Striking differences were found between the two activators, most notably in the relative strengths of the TATA/Inr and Inr core promoters: the TATA/Inr promoter was much stronger than the Inr promoter when transcription was activated by GAL4-VP16, but the strengths of the two promoters were more comparable when transcription was activated by Sp1. To define the domains of Sp1 responsible for efficient activation through an Inr, several Sp1 deletion mutants were tested as GAL4 fusion proteins. The results reveal that the glutamine-rich activation domains, which previously were found to interact with Drosophila TAF110, preferentially stimulate Inr-containing core promoters. In contrast, efficient activation through TATA appears to require additional domains of Sp1. These results demonstrate that activation domains differ in their abilities to function with specific core promoters, suggesting that the core promoter structure found in a given gene may reflect a preference of the regulators of that gene. Furthermore, the core promoter preference of an activation domain may be related to a specific mechanism of action, which may provide a functional criterion for grouping activation domains into distinct classes.
Mol Cell Biol 1995 Nov
PMID:Core promoter specificities of the Sp1 and VP16 transcriptional activation domains. 756 43

Rox1 is a repressor of the hypoxic genes of Saccharomyces cerevisiae. It binds to a specific hypoxic consensus sequence in the upstream region of these genes and represses transcription in conjunction with the general repression complex Tup1-Ssn6. In this study, we demonstrated that the first 100 amino acids comprising the HMG domain of Rox1 were responsible for DNA binding and that when bound, Rox1 bent DNA at an angle of 90 degrees. A mutational analysis resulted in the isolation of seven missense mutations, all located within the HMG domain, that caused loss of DNA binding. The effect of these mutations on the structure of Rox1 was evaluated on the basis of the homology between Rox1 and the human male sex-determining protein SRY, for which a structural model is available. The failure to isolate missense mutations in the carboxy-terminal three-quarters of the protein prompted a deletion analysis of this region. The results suggested that this region was responsible for the repression function of Rox1 and that the repression information was redundant. This hypothesis was confirmed by using a set of fusions between sequences encoding the GAL4 DNA-binding domain and portions of ROX1. Those fusions containing either the entire carboxy-terminal region or either half of it were capable of repression. Repression by selected fusions was demonstrated to be dependent on Ssn6.
Mol Cell Biol 1995 Nov
PMID:Mutational analysis of Rox1, a DNA-bending repressor of hypoxic genes in Saccharomyces cerevisiae. 756 63

To investigate the mechanisms by which transcription factors invade nucleosomal DNA and replace histones at control elements, we have examined the response of the histone octamer to transcription factor binding in the presence of histone-binding proteins (i.e., nucleosome assembly factors). We found that yeast nucleosome assembly protein 1 (NAP-1) stimulated transcription factor binding and nucleosome displacement in a manner similar to that of nucleoplasmin. In addition, disruption of the histone octamer was required both for the stimulation of transcription factor binding to nucleosomal DNA and for transcription factor-induced nucleosome displacement mediated by nucleoplasmin or NAP-1. While NAP-1 and nucleoplasmin stimulated the binding of a fusion protein (GAL4-AH) to control nucleosome cores, this stimulation was lost upon covalent histone-histone cross-linking within the histone octamers. In addition, both NAP-1 and nucleoplasmin were able to mediate histone displacement upon the binding of five GAL4-AH dimers to control nucleosome cores; however, this activity was also forfeited when the histone octamers were cross-linked. These data indicate that octamer disruption is required for both stimulation of factor binding and factor-dependent histone displacement by nucleoplasmin and NAP-1. By contrast, transcription factor-induced histone transfer onto nonspecific competitor DNA did not require disruption of the histone octamer. Thus, histone displacement in this instance occurred by transfer of complete histone octamers, a mechanism distinct from that mediated by the histone-binding proteins nucleoplasmin and NAP-1.
Mol Cell Biol 1995 Nov
PMID:Stimulation of transcription factor binding and histone displacement by nucleosome assembly protein 1 and nucleoplasmin requires disruption of the histone octamer. 756 70

TFIIA is a general transcription factor that interacts with the TFIID-promoter complex required for transcription initiation by RNA polymerase II. Two lines of evidence suggest that TFIIA is directly involved in the mechanism by which some activators stimulate transcription. First, binding of TFIIA to a TFIID-promoter complex is a rate-limiting step that is enhanced by transcriptional activators GAL4-AH and Zta. Second, recombinant TFIIA greatly enhances activator-dependent transcription. In this study, we found that the activation domains of Zta and VP16 bind directly to TFIIA. Both Zta and VP16 stimulated rapid assembly of a stable TFIID-TFIIA complex on promoter DNA. Analysis of deletion derivatives of the VP16 activation domain indicated that the ability to bind to TFIIA correlates with the ability to enhance TFIID-TFIIA-promoter ternary complex assembly. Thus, we propose that a class of activators stimulate transcription initiation through direct interactions with both TFIIA and TFIID, which stimulate the assembly of an activated TFIIA-TFIID-promoter complex.
Mol Cell Biol 1995 Nov
PMID:A class of activation domains interacts directly with TFIIA and stimulates TFIIA-TFIID-promoter complex assembly. 756 98

The expression of the structural genes nit-3 and nit-6, which encode the nitrate assimilatory enzymes nitrate reductase and nitrite reductase, respectively, is highly regulated by the global-acting NIT2 regulatory protein. These structural genes are also controlled by nitrogen catabolite repression and by specific induction via nitrate. A pathway-specific regulatory protein, NIT4, appears to mediate nitrate induction of nit-3 and of nit-6. The NIT4 protein, composed of 1090 amino acids, contains a putative GAL4-like Cys-6 zinc cluster DNA-binding motif, which is joined by a short segment to a stretch of amino acids that appear to constitute a coiled-coil dimerization domain. Chemical crosslinking studies demonstrated that a truncated form of NIT4 forms homodimers. Mobility-shift and DNA-footprinting experiments have identified two NIT4-binding sites of significantly different strengths in the promoter region of the nit-3 gene. The stronger binding site contains a symmetrical octameric sequence, TCCGCGGA, whereas the weaker site has a related sequence. Sequences related to this palindromic element can be found upstream of the nit-6 gene.
Mol Microbiol 1995 Mar
PMID:Sequence-specific DNA binding by NIT4, the pathway-specific regulatory protein that mediates nitrate induction in Neurospora. 759 94

Overproduction of Gcn4p in yeast cells resulted in the inhibition of transcription from promoters controlled by the GAL4 or dA:dT elements. We have demonstrated that this effect is mediated through the activation domain of Gcn4p and that the function of the transcriptional activator at the affected promoter is impaired. The inhibitory effect of Gcn4p and that the function of the transcriptional activator at the affected promoter is impaired. The inhibitory effect of Gcn4p on these promoters persisted in yeast strains disrupted for the ADA2 and/or GCN5 genes, whose products are required for only part of the transcriptional activation capacity of Gcn4p and other activators, but was alleviated by overexpression of gamma TFIIB. These results support the hypothesis that general transcription factors become unavailable at certain promoters when an activator is overexpressed and strongly imply the existence of an Ada2p/Gcn5p-independent pathway of communication between acidic activators and the basic transcription machinery. In a genetic screen, we have isolated a mutation which neutralises the squelching effects of Gcn4p. This AFR1-1 (activation function reduced) mutation is dominant, it affects the transcriptional activation properties of a number of activators and results in lethality when combined with a gcn5 disruption. Our results suggest that the AFR1 gene product is involved in the mediation of transcriptional activation.
Mol Gen Genet 1995 Jun 10
PMID:Transcriptional interference caused by GCN4 overexpression reveals multiple interactions mediating transcriptional activation. 760 36

Tat strongly activates transcription of the HIV-1 provirus by stimulating both initiation and elongation. This transactivator binds to the TAR RNA element, but can also associate with cellular transcription factors, interacting with upstream promoter sequences. To achieve a better understanding of the role of Tat in the assembly of the transcriptional initiation complex in the living cell, we have examined how the activity of this protein is modified when the general transcription factor involved in the first step of this process, TBP, is overexpressed. The activity of Tat, either wild-type or fused to the DNA binding domain of GAL4 (GBTat), was tested using reporter constructs containing GAL4 binding sites upstream of a minimal promoter corresponding to the HIV-1 TATA box, with or without the TAR element. We found that overexpression of TBP led to a dramatic increase in the activity of the GBTat protein. In order to activate GBTat, TBP must be able to interact with the TATA box. Analysis of several Tat mutants indicated that both the cysteine-rich and the core domains of this transactivator are necessary and sufficient to activate transcription when TBP is overexpressed. In vitro experiments showed that Tat binds specifically to TBP. There was a correlation between the ability of different Tat mutants to bind TBP and their capacity to activate transcription in vivo. With the natural HIV-1 promoter, overexpression of TBP first stimulated and then suppressed the Tat-induced activity. This inhibition was abrogated by an increase in the intracellular levels of Tat. These experimental data indicate that Tat stimulates initiation of transcription by interacting with TBP in vivo.
J Mol Biol 1995 Jul 07
PMID:Evidence for functional interaction between the HIV-1 Tat transactivator and the TATA box binding protein in vivo. 760 68

A human protein that is 92% identical and 97% homologous at the amino acid level to RanBP1 from mouse was identified by the two-hybrid method, using two types of target cDNAs fused to sequences encoding the GAL4 DNA-binding domain. The target cDNAs encoded the human Ran/TC4 and human RCC1 proteins, respectively. An in vitro binding experiment showed that RanBP1 binds to RCC1 with the aid of Ran. Partially purified, GST-fused RanBP1 inhibited RCC1-stimulated guanine nucleotide release from Ran in vitro. Consistent with this in vitro finding, overproduction of human RanBP1 was detrimental to growth of tsBN2, a temperature-sensitive BHK21 hamster cell line defective in the RCC1 gene, and inhibited the growth of the Saccharomyces cerevisiae rcc1 mutants prp20, mtr1 and srm1. The specific effect of RanBP1 on rcc1- cells was confirmed by the finding that overproduction of RanBP1 induces significant levels of expression of a FUS1-lacZ gene and an increase in mating efficiencies in a ste3, pheromone receptor-deficient yeast mutant. This phenotype is similar to the srm1, a mutant isolated as a suppressor that restores mating to receptorless mutants. These findings indicate that RanBP1 negatively regulates RCC1.
Mol Gen Genet 1995 Jun 25
PMID:RanBP1, a Ras-like nuclear G protein binding to Ran/TC4, inhibits RCC1 via Ran/TC4. 761 57

We have characterized a stress-responsive transcriptional activation domain of mouse heat shock factor 1 (HSF1) by using chimeric GAL4-HSF1 fusion proteins. Fusion of the GAL4 DNA-binding domain to residues 124 to 503 of HSF1 results in a chimeric factor that binds DNA yet lacks any transcriptional activity. Transactivation is acquired upon exposure to heat shock or by deletion of a negative regulatory domain including part of the DNA-binding-domain-proximal leucine zippers. Analysis of a collection of GAL4-HSF1 deletion mutants revealed the minimal region for the constitutive transcriptional activator to map within the extreme carboxyl-terminal 108 amino acids, corresponding to a region rich in acidic and hydrophobic residues. Loss of residues 395 to 425 or 451 to 503, which are located at either end of this activation domain, severely diminished activity, indicating that the entire domain is required for transactivation. The minimal activation domain of HSF1 also confers enhanced transcriptional response to heat shock or cadmium treatment. These results demonstrate that the transcriptional activation domain of HSF1 is negatively regulated and that the signal for stress induction is mediated by interactions between the amino-terminal negative regulator and the carboxyl-terminal transcriptional activation domain.
Mol Cell Biol 1995 Aug
PMID:The carboxyl-terminal transactivation domain of heat shock factor 1 is negatively regulated and stress responsive. 762 25


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