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Query: UNIPROT:P51532 (transcriptional activator)
 6,546 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We demonstrate that the hormone-binding domain (HBD) of the human estrogen receptor (ER) can function as an autonomous regulatory domain in the budding yeast, Saccharomyces cerevisiae. As in mammalian cells, the HBD can subject the activity of a heterologous protein, which is fused to it, to hormonal control. Thus, a chimeric transcriptional activator consisting of (i) the DNA-binding domain of GAL4, (ii) the ER HBD, and (iii) the activation domain of viral protein 16 (VP16) stimulates both episomal and integrated reporter genes exclusively in the presence of steroid hormone. Steroids being gratuitous signals for yeast, this fusion protein is a convenient tool for highly regulated production of proteins of interest. Notably, it can be exploited to activate the commonly used galactose-inducible expression vectors without switching the carbon source.
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PMID:Fusion of GAL4-VP16 to a steroid-binding domain provides a tool for gratuitous induction of galactose-responsive genes in yeast. 837 May 33

Cooperative DNA binding of the bovine papillomavirus type 1 (BPV-1) E2 transcriptional activator (E2-TA) is thought to play a role in the transcriptional synergism of multiple E2-responsive DNA elements (J. Ham, N. Dostatni, J.-M. Gauthier, and M. Yaniv, Trends Biochem. Sci. 16:440-444, 1991). Binding-equilibrium considerations show that such involvement is unlikely, thereby suggesting that the E2-TA cooperative capacity may have evolved to play other, different roles. The role of cooperative interactions in the antagonistic activity of BPV-1-positive and BPV-1-negative E2 regulatory proteins was investigated by an in vitro quantitative gel shift assay. Viral repressor E2-TR, a truncated peptide encompassing the activator DNA-binding domain, possesses a small but measurable cooperative capacity. Furthermore, the minimal E2 DNA-binding domain interacts with the activator in a positive, heterocooperative manner. As a result, the in vitro competition of full-length and truncated E2 peptides appears to be (macroscopically) noncooperative. This heterocooperative effect is probably dominant in latently infected G0-G1 cells, in which repressor E2-TR is 10- to 20-fold more abundant than the activator. The data are discussed considering the possible role of homo- and heterocooperative DNA binding in E2-conditional gene expression.
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PMID:Cooperative DNA binding of the bovine papillomavirus E2 transcriptional activator is antagonized by truncated E2 polypeptides. 839 66

Cyclic AMP-regulated gene expression frequently involves a DNA element known as the cAMP-regulated enhancer (CRE). Many transcription factors bind to this element, including the protein CREB, which is activated as a result of phosphorylation by protein kinase A. This modification stimulates interaction with one or more of the general transcription factors or, alternatively, allows recruitment of a co-activator. Here we report that CREB phosphorylated by protein kinase A binds specifically to a nuclear protein of M(r) 265K which we term CBP (for CREB-binding protein). Fusion of a heterologous DNA-binding domain to the amino terminus of CBP enables the chimaeric protein to function as a protein kinase A-regulated transcriptional activator. We propose that CBP may participate in cAMP-regulated gene expression by interacting with the activated phosphorylated form of CREB.
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PMID:Phosphorylated CREB binds specifically to the nuclear protein CBP. 841 73

B-myc is a recently described myc gene whose product has not been functionally characterized. The predicted product of B-myc is a 168-amino-acid protein with extensive homology to the c-Myc amino-terminal region, previously shown to contain a transcriptional activation domain. We hypothesized that B-Myc might also function in transcriptional regulation, although its role in regulating gene expression is predicted to be unique, because B-Myc lacks the specific DNA-binding motif found in other Myc proteins. To determine whether B-Myc could interact with the transcriptional machinery, we studied the transcriptional activation properties of a chimeric protein containing B-Myc sequences fused to the DNA-binding domain of the yeast transcriptional activator GAL4 (GAL4-B-Myc). We found that GAL4-B-Myc strongly activated expression of a GAL4-regulated reporter gene in mammalian cells. In addition, full-length B-Myc was able to inhibit or squelch reporter gene activation by a GAL4 chimeric protein containing the c-Myc transcriptional activation domain. We also observed that B-Myc dramatically inhibited the neoplastic cotransforming activity of c-Myc and activated Ras in rat embryo cells. Because B-Myc inhibits both neoplastic transformation and transcriptional activation by c-Myc, we suggest that the transforming activity of c-Myc is related to its ability to regulate transcription. Whether B-Myc functions biologically to squelch transcription and/or to regulate transcription through a specific DNA-binding protein remains unestablished.
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PMID:B-myc inhibits neoplastic transformation and transcriptional activation by c-myc. 842 80

Simian virus 40 (SV40) large T antigen is a potent transcriptional activator of both viral and cellular promoters. Within the SV40 late promoter, a specific upstream element necessary for T-antigen transcriptional activation is the binding site for transcription-enhancing factor 1 (TEF-1). The promoter structure necessary for T-antigen-mediated transcriptional activation appears to be simple. For example, a promoter consisting of upstream TEF-1 binding sites (or other factor-binding sites) and a downstream TATA or initiator element is efficiently activated. It has been demonstrated that transcriptional activation by T antigen does not require direct binding to the DNA; thus, the most direct effect that T antigen could have on these simple promoters would be through protein-protein interactions with either upstream-bound transcription factors, the basal transcription complex, or both. To determine whether such interactions occur, full-length T antigen or segments of it was fused to the glutathione-binding site (GST fusions) or to the Gal4 DNA-binding domain (amino acids 1 to 147) (Gal4 fusions). With the GST fusions, it was found that TEF-1 and the TATA-binding protein (TBP) bound different regions of T antigen. A GST fusion containing amino acids 5 to 172 (region T1) efficiently bound TBP. TEF-1 bound neither region T1 nor a region between amino acids 168 and 373 (region T2); however, it bound efficiently to the combined region (T5) containing amino acids 5 to 383.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Transcriptional activation by simian virus 40 large T antigen: interactions with multiple components of the transcription complex. 842 15

The human ets-2 proto-oncogene is one of the homologs of the v-ets gene, found in avian acutely transforming retrovirus E26 (D. Leprince, A. Gegonne, J. Call, C. de Taisne, A. Schneeberger, C. Lagrou, and D. Stehelin, Nature [London] 306:395-397, 1983; M. F. Nunn, P. H. Seeburg, C. Moscovici, and P. H. Duesberg, Nature [London] 306:391-395, 1983), which causes leukemia in chickens. We used the DNA-binding domain of yeast transcriptional activator GAL4 to locate the transactivation region of human ets-2. The transactivation domain of ets-2 was found in the N-terminal part of the protein, which is homologous to ets-1, and can be disrupted by deletion of a stretch of acidic amino acid residues. A transactivation-deficient mutant of ets-2 failed to transform Rat-1 cells and suppressed the transforming activity of coexpressed wild-type ets-2. A mutation in the putative DNA-binding region of ets-2 abolished transforming activity. We show that the motif crucial for ets-2 transactivation capability is necessary for transforming activity in Rat-1 cells. Mutant ets-2 protein that lacks the transactivation domain has a dominant negative effect on transformation by wild-type ets-2. We were unable to detect ets-2-dependent transcriptional regulation of several enhancers containing ets-binding motifs.
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PMID:Localization of the c-ets-2 transactivation domain. 844 38

The yeast transcriptional activator HAP1 contains a DNA-binding domain homologous to the zinc finger of GAL4 and an adjacent regulatory domain that blocks DNA binding in the absence of the inducer heme. We show that short HAP1 fragments containing the zinc finger are unable to bind to DNA but can be rescued by antibody to the HAP1 zinc finger. These fragments are missing a coiled-coil sequence similar to that within the dimerization domain of GAL4 and dimerization domains of myosin heavy chain. We surmise that the antibody promotes DNA binding by bringing together two monomers. Interestingly, the antibody will also promote DNA binding of a larger HAP1 fragment containing the DNA-binding and the heme-regulatory domains. This suggests that the regulatory domain acts by preventing dimerization of HAP1 in the absence of heme. Consistent with this view is an in vivo assay that also reveals that heme promotes HAP1 dimerization in yeast cells.
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PMID:Antibody-promoted dimerization bypasses the regulation of DNA binding by the heme domain of the yeast transcriptional activator HAP1. 846 99

A DNA fragment encoding the yeast GAL4 transcriptional activator DNA-binding domain (amino acids 1-93) was cloned into an Escherichia coli expression vector such that the overproduced protein is tagged with six histidine residues and a factor Xa protease cleavage site. The vector also contains unique restriction sites at the 3' end of the gene to allow the construction of fusion proteins. These fusion proteins can easily be purified to homogeneity and their activity tested in vitro.
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PMID:Overproduction and single-step purification of GAL4 fusion proteins from Escherichia coli. 847 50

Transcription of the genes required for utilization of galactose in Saccharomyces cerevisiae is controlled primarily by the transcriptional activator protein GAL4. The upstream activating sequences for galactose (UASG) of most GAL genes have multiple sites to which GAL4 can bind. In this report we compare the binding properties of wild type GAL4 and derivatives of GAL4 bearing the N-terminal DNA-binding domain to multiple DNA-binding sites in vitro. To produce wild type GAL4, we constructed a recombinant baculovirus for expression in insect cells. Recombinant wild type GAL4 was found to bind efficiently to an oligonucleotide containing a near-consensus 17-mer GAL4 DNA-binding site in electrophoretic mobility shift assays. Footprinting experiments revealed that wild type GAL4 binds cooperatively to the four GAL4 DNA-binding sites of the GAL1-10 UASG; however, in contrast an N-terminal fragment of GAL4 containing only the DNA-binding/dimerization domains binds to each of these sites with slightly different affinity. With increasing concentrations of GAL4(1-147), the four sites become filled in the following order: site II, site IV, site I, and site III. In experiments with wild type GAL4, these four sites become fully occupied at approximately the same concentration of protein. In footprints of wild type GAL4 on the USAG, enhancements and protections of DNase I-sensitive cleavages are detectable between sites III and IV, indicative of formation of a loop between these distantly spaced sites. Binding of wild type GAL4 to a strong near-consensus binding site assists binding to an adjacent mutant site in both electrophoretic mobility shift and footprinting assays. GAL4(1-147) and GAL4(1-147) fused to portions of GAL4's activating region II were incapable of cooperative DNA binding in our assays. We conclude from these observations that wild type GAL4 has a cooperative DNA-binding function that is distinct from the DNA binding and dimerization or transcriptional activation functions, and likely plays and important role in precise regulation of GAL gene transcription.
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PMID:Wild type GAL4 binds cooperatively to the GAL1-10 UASG in vitro. 848 50

We used a yeast-based genetic assay, the two-hybrid system, to characterize the domain of the tumor-suppressor p53 involved in oligomerization. This assay relies on the reconstitution of the function of a transcriptional activator, the yeast GAL4 protein, via the interaction of a protein fused to the DNA-binding domain of GAL4 with a protein fused to the transcriptional activation domain of GAL4. We show by a reconstruction experiment that this approach could detect the interaction of p53 deleted for its N-terminal activation domain with SV40 large T antigen. We then searched a library of human proteins present as activation domain hybrids for proteins that can interact with the hybrid of p53 with the DNA-binding domain. This search identified 36 plasmids containing the p53 gene, representing 10 different classes. These results provide an additional in vivo demonstration of p53 oligomerization. The smallest p53 fragment identified from screening the library contained only amino acids 331-393, indicating that this small C-terminal fragment is sufficient to mediate oligomerization. In addition, a mutant p53 protein could bind to the wild-type protein in this assay, providing support for the idea that mutant forms of p53 act in a dominant-negative manner through C-terminal oligomerization with the wild type.
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PMID:Use of the two-hybrid system to identify the domain of p53 involved in oligomerization. 850 89


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