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

Understanding the mechanism of glucose repression in yeast has proved to be a difficult and challenging problem. A multitude of genes in different pathways are repressed by glucose at the level of transcription. The SUC2 gene, which encodes invertase, is an excellent reporter gene for glucose repression, since its expression is controlled exclusively by this pathway. Genetic analysis has identified numerous regulatory mutations which can either prevent derepression of SUC2 or render its expression insensitive to glucose repression. These mutations allow us to sketch the outlines of a pathway for general glucose repression, which has several key elements: hexokinase PII, encoded by HXK2, which seems to play a role in the sensing of glucose levels; the protein kinase encoded by SNF1, whose activity is required for derepression of many glucose-repressible genes; and the MIG1 repressor protein, which binds to the upstream regions of SUC2 and other glucose-repressible genes. Repression by MIG1 requires the activity of the CYC8 and TUP1 proteins. Glucose repression of other sets of genes seems to be controlled by the general glucose repression pathway acting in concert with other mechanisms. In the cases of the GAL genes and possibly CYC1, regulation is mediated by a cascade in which the general pathway represses expression of a positive transcriptional activator.
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PMID:Glucose repression in the yeast Saccharomyces cerevisiae. 131 Jul 93

In this report we study the effects of internal deletions of the yeast transcriptional activator HAP1 (CYP1) on activity at two dissimilar DNA binding sites, upstream activation sequence 1 (UAS1) of CYC1 (iso-1-cytochrome c) and CYC7 (iso-2-cytochrome c). These deletions remove up to 1061 amino acids of the 1483-residue protein and bring the carboxyl-terminal acidic activation domain closer to the amino-terminal DNA-binding domain. Surprisingly, the deletions have opposite effects at the two sites; activity at UAS1 increases with deletion size, while activity at CYC7 decreases. The mutant with the largest deletion, mini-HAP1, has no measurable activity at CYC7 but binds normally to the site in vitro. In contrast, a protein with the DNA-binding domain of HAP1 fused to the acidic activation domain of GAL4 is active at both UAS1 and CYC7. These findings are discussed in the context of two models that suggest how the DNA sequence can alter the activity of the bound HAP1. In a separate experiment, we generate a mutation in the DNA-binding domain of HAP1 that requires the addition of zinc for binding to either UAS1 or CYC7 in vitro. This finding shows that a zinc finger anchors DNA binding to both types of HAP1 sites.
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PMID:Internal deletions in the yeast transcriptional activator HAP1 have opposite effects at two sequence elements. 216 46

Expression of the yeast pyrimidine biosynthetic gene, URA3, is induced three- to fivefold in response to uracil starvation, and this regulation is mediated by the transcriptional activator PPR1 (pyrimidine pathway regulator 1). In this study, we have analyzed the regulatory elements of the URA3 promoter by DNase I footprinting, using partially purified yeast cell extracts, by deletion mutagenesis, and by 5'-end mapping of RNA transcripts. Two DNA-binding activities have been detected, and at least four distinct cis-acting regions have been identified. A region rich in poly(dA-dT) serves as an upstream promoter element necessary for the basal level of URA3 expression. A 16-base-pair sequence with dyad symmetry acts acts as a uracil-controlled upstream activating site (UASURA) and shows a specific binding only with cell extracts from strains overproducing PPR1. This in vitro binding does not require dihydroorotic acid, the physiological inducer of URA3. The TATA region appears to be composed of two functionally distinct (constitutive and regulatory) elements. Two G + A-rich regions surrounding this TATA box bind an unidentified factor called GA-binding factor. The 5' copy, GA1, is involved in PPR1 induction and overlaps the constitutive TATA region. The 3' region, GA2, is necessary for maximal expression. Neither of these GA sequences acts as a UAS in a CYC1-lacZ context. The promoters of the unlinked but coordinately regulated URA1 and URA4 genes contain highly conserved copies of the UASURA sequence, which prompted us to investigate the effects of many point mutations within this UASURA sequence on PPR1-dependent binding. In this way, we have identified the most important residues of this binding site and found that a nonsymmetrical change of these bases is sufficient to prevent the specific binding and to suppress the UASURA activity in vivo. In addition, we showed that UASURA contains a constitutive activating element which can stimulate transcription from a heterologous promoter independently of dihydroorotic acid and PPR1.
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PMID:cis- and trans-acting regulatory elements of the yeast URA3 promoter. 220 10

The DNA sequence UAST (TCGTTTTGTACGTTTTTCA) was found to mediate transcription of yeast ribosomal protein gene TCM1. UAST was defined as a transcriptional activator on the basis of loss of transcription accompanying deletions of all or part of UAST, orientation-independent restoration of transcription promoted by a synthetic UAST oligomer inserted either into TCM1 or into the yeast CYC1 gene lacking its transcriptional activation region, and diminished transcription following nucleotide alterations in UAST. UAST bound in vitro to a protein denoted TAF (TCM1 activation factor); TAF was concluded to be a transcriptional activator protein because nucleotide alterations in UAST that diminished transcription in vivo also diminished TAF binding in vitro. The sequence of UAST bore no obvious resemblance to UASrpg, the principal cis-acting element common to most yeast ribosomal protein genes. Likewise, TAF was distinguished from the UASrpg-binding protein TUF, since (i) TAF and TUF were chromatographically separable, (ii) binding of either TAF or TUF to its corresponding UAS was unaffected by an excess of UASrpg or UAST DNA, respectively, and (iii) photochemical cross-linking experiments showed that TAF was a protein of 147 kilodaltons (kDa), while TUF was detected as an approximately 120-kDa polypeptide, consistent with its known size. Cross-linking experiments also revealed that both UAST and UASrpg bound a second heretofore unobserved 82-kDa protein; binding of this additional protein appeared to require binding of TAF or TUF. On the basis of the biochemical characterization of TAF and a lack of sequence similarity between UAST and UASrpg, we suggest that transcription of TCM1 is mediated by a cis-acting sequence and at least one trans-acting factor different from the elements which promote transcription of most other ribosomal protein genes. A second trans-acting factor may be shared by TCM1 and other ribosomal protein genes; this factor could mediate coordinate regulation of these genes.
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PMID:Constitutive transcription of yeast ribosomal protein gene TCM1 is promoted by uncommon cis- and trans-acting elements. 305 14

The GATA motif (WGATAR) is found in the promoter regions of numerous Caenorhabditis elegans genes, including two intestine-specific genes, vit-2 and ges-1, in which it has been shown to be required for promoter function. The protein ELT-1, encoded by a single-copy gene homologous to the GATA family of vertebrate transcription factors, is potentially capable of interacting with this element. In order to determine whether ELT-1 is a transcriptional activator that recognizes this sequence, we have expressed it under the control of the GAL1 promoter in yeast. lacZ driven by the CYC1 promoter lacking an upstream activation sequence (UAS) but containing GATA sequences was used as a reporter. beta-Galactosidase was expressed upon induction only when GATA sequences were present, and expression was increased dramatically by additional binding sites. Deletion analysis demonstrated that the C terminus, containing only one of the two zinc fingers, is sufficient for activation. In addition, the DNA-binding domain and two transactivation regions were identified by fusing these isolated domains to previously defined domains of heterologous transcription factors. While most single base alterations in the GATA core sequence eliminated activity, an A to C change in position four, creating a GATC core, was found to increase activity significantly. The deleted ELT-1 protein containing only the C-terminal Zn finger was sufficient for activation in response to GATA, but both fingers were required for activation at GATC. A variety of sites with non-optimal sequences surrounding the GATA core also were found to be excluded better by the protein containing both Zn fingers. Furthermore, a fusion protein containing the entire ELT-1 DNA binding domain fused to the VP16 activation domain was found to have an even greater preference for the GATC core, as well as the optimal flanking bases. We conclude that, although ELT-1 having only its C-terminal finger is capable of activation in response to the WGATAR site, the presence of the upstream finger supplies additional base specificity.
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PMID:Activity of a C. elegans GATA transcription factor, ELT-1, expressed in yeast. 747 42

In both prokaryotes and eukaryotes nonsense mutations in a gene can enhance the decay rate of the mRNA transcribed from the gene, a phenomenon described as nonsense-mediated mRNA decay. In yeast, the products of the UPF1 and UPF3 genes are required for this decay pathway, and in this report we focus on the identification and characterization of additional factors required for rapid decay of nonsense-containing mRNAs. We present evidence that the product of the UPF2 gene is a new factor involved in this decay pathway. Mutation of the UPF2 gene or deletion of it from the chromosome resulted in stabilization of nonsense-containing mRNAs, whereas the decay of wild-type transcripts was not affected. The UPF2 gene was isolated, and its transcript was characterized. Our results demonstrate that the UPF2 gene encodes a putative 126.7-kD protein with an acidic region at its carboxyl terminus (-D-E)n found in many nucleolar and transcriptional activator proteins. The UPF2 transcript is 3600 nucleotides in length and contains an intron near its 5' end. The UPF2 gene is dispensable for vegetative growth, but upf2 delta strains were found to be more sensitive to the translational elongation inhibitor cycloheximide than UPF2+. A genetic analysis of other alleles proposed to be involved in nonsense-mediated mRNA decay revealed that the UPF2 gene is allelic to the previously identified sua1 allele, a suppressor of an out-of-frame ATG insertion shown previously to reduce translational initiation from the normal ATG of the CYC1 gene. In addition, we demonstrate that another suppressor of this cyc1 mutation, sua6, is allelic to upf3, a previously identified lesion involved in nonsense-mediated mRNA decay.
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PMID:Identification and characterization of genes that are required for the accelerated degradation of mRNAs containing a premature translational termination codon. 788 67

Two new yeast genes, named MBR1 and MBR3, were isolated as multicopy suppressors of the growth defect of a strain lacking the HAP2 transcriptional activator. Both genes when overexpressed can also suppress the growth defect of hap3 and hap4 null mutants. However, overexpression of MBR1 cannot substitute for the HAP2/3/4 complex in activation of the CYC1 gene. Nucleotide sequencing of MBR1 and MBR3 revealed that these two genes encode serine-rich, hydrophilic proteins with regions of significant homology. The functional importance of one of these conserved regions was shown by mutagenesis. Disruption of MBR1 leads to a partial growth defect on glycerol medium. Disruption of MBR3 has no major effect but the double disruptant shows a synthetic phenotype suggesting that the MBR1 and MBR3 gene products participate in common function.
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PMID:MBR1 and MBR3, two related yeast genes that can suppress the growth defect of hap2, hap3 and hap4 mutants. 820 48

Transcription of the PHO84 gene encoding a Pi transporter in Saccharomyces cerevisiae is regulated by the Pi concentration in the medium. The promoter region of PHO84 bears five copies of the motif 5'-CACGT(G/T)-3', a candidate for the upstream activation site (UAS) that binds the transcriptional activator protein of the phosphatase regulon, Pho4p. These motifs are found at nucleotides -880 (site A), -587 (B), -436 (C), -414 (D), and -262 (E) relative to the putative ATG codon of PHO84. The Pho4p binds to all five 6-bp motifs with various affinities. Deletion analysis of the PHO84 promoter using a PHO84-lacZ fusion gene and base substitutions in the 6-bp motif revealed that two copies of the 6-bp motif, either C or D, and E, are necessary and sufficient for full regulation of the PHO84 gene. Results of expression studies with a CYC1-lacZ fusion gene with various 36-bp oligonucleotides including the 30-bp sequences around site D or E, or with modified sequences, inserted in the CYC1 promoter region indicated that the 6-bps motif flanked by a thymine nucleotide at its 5' end is much less effective as a UAS site for Pho4p in vivo than other versions. Thus, the consensus sequences for phosphatase regulation are 5'-GCACGTGGG-3' and 5'-GCACGTTTT-3' which differ from the binding sequences for the Cpflp protein required for transcription of the genes in methionine biosynthesis and for centromere function. However, Pho4p binding in vitro was unaffected by modification of the 5' or 3' flanking sites of the 6-bp motif, while modification inside the 6-bp motif affected it severely. The UAS function of the GCACGTTTT motif with respect to the Pi signal depends on its orientation in the promoter sequence.
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PMID:Structure and distribution of specific cis-elements for transcriptional regulation of PHO84 in Saccharomyces cerevisiae. 855 45

The yeast zinc cluster protein HAP1, a member of the GAL4 family, is a transcriptional activator that binds as a homodimer to target DNA sequences. These targets include the upstream activating sequences of the CYC1 and CYC7 genes, which have no obvious sequence similarity. Even though both sites have the same affinity for HAP1, activation differs at these two sites, even when the sequences are placed in an identical promoter context. In addition, mutants of HAP1 that can bind to both sites but are specifically transcriptionally inactive at CYC7 have been previously isolated. In order to identify nucleotides that are responsible for this differential activity, we have performed random and site-directed mutagenesis of these target sites and assayed their binding to HAP1 in vitro and their activity in vivo in reporter plasmids. Our results show that HAP1 binding sites are degenerate forms of the direct repeat CGG N3 TA N CGG N3 TA. Moreover, we show that activity of HAP1 mutants defective for activation of the CYC7gene is restored by specific mutations in the CYC7 binding site. Conversely, other mutations of the target sites prevent activation by HAP1, without interfering with DNA binding. The results suggest that the sequence of the target sites influences the conformation and, hence, the activity of DNA-bound HAP1.
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PMID:Mutations in target DNA elements of yeast HAP1 modulate its transcriptional activity without affecting DNA binding. 862 77

The yeast repressor Rme1p acts from distant binding sites to block transcription of the chromosomal IME1 gene. Rme1p can also repress the heterologous CYC1 promoter when Rme1p binding sites are placed 250-300 bp upstream of CYC1 transcriptional activator binding sites (UAS1 and UAS2). Here, in vivo footprinting studies indicate that Rme1p acts over this distance by preventing the binding of the CYC1 transcriptional activators to UAS1 and UAS2. Inhibition of activator binding by Rme1p has the same genetic requirements as repression: both depend upon sequences flanking the Rme1p binding sites and upon Rgr1p and Sin4p, two subunits of the RNA polymerase II-associated Mediator complex that are required for normal nucleosome density. Thus Rme1p may alter chromatin to prevent binding of transcriptional activators to distant DNA sequences.
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PMID:Transcriptional repression at a distance through exclusion of activator binding in vivo. 902 35


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