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Showing papers on "Upstream activating sequence published in 1994"


Journal ArticleDOI
Krassimir Yankulov1, Justin Blau1, Tracey Purton1, Sadia Roberts1, David Bentley1 
03 Jun 1994-Cell
TL;DR: It is suggested that setting the competence of polymerase II to elongate is an integral part of the initiation step that is controlled by activators cooperating with the general transcription factors.

249 citations


Journal ArticleDOI
TL;DR: The presence of A‐factor homologues in a wide variety of Streptomyces species and distantly related bacteria implies the generality of γ‐butyrolactones as chemical cellular signalling molecules in microorganisms.
Abstract: Summary A-factor, containing a γ-butyrolactone in its structure, is an autoregulatory factor or a‘microbiai hormone’controlling secondary metabolism and cellular differentiation in Streptomyces griseus. A-factor exerts its regulatory role by binding to a specific receptor protein which, in the absence of A-factor, acts as a repressor-type regulator for morphological and physiological differentiation, in the signal relay leading to streptomycin production in S. griseus, the A-factor signal is transferred from the A-factor receptor to the upstream activation sequence of a regulatory gene, strR, in the streptomycin biosynthetic gene cluster via an A-factor-dependent protein that serves as a transcription factor for strR. The StrR protein thus Induced appears to activate the transcription of other streptomycin-production genes. The presence of A-factor homologues in a wide variety of Streptomyces species and distantly related bacteria implies the generality of γ-butyrolactones as chemical cellular signalling molecules in microorganisms.

185 citations


Journal ArticleDOI
TL;DR: The results establish that amino acids 258-265 of alpha constitute an "activation target" essential for CAP-dependent transcription at the lac promoter but not essential forCAP-independent transcription, and amino acid 261 is the most critical amino acid of the activation target.
Abstract: We have isolated and characterized single-amino-acid substitution mutants of RNA polymerase a subunit defective in CAP-dependent transcription at the lac promoter but not defective in CAP-independent transcription. Our results establish that (1) amino acids 258- 265 of a constitute an "activation target" essential for CAP-dependent transcription at the lac promoter but not essential for CAP-independent transcription, (2) amino acid 261 is the most critical amino acid of the activation target, (3) amino acid 261 is distinct from the determinants for a-DNA interaction, and (4) the activation target may fold as a surface amphipathic a-helix. We propose a model for transcriptional activation at the lac promoter that integrates these and other recent results regarding transcriptional activation and RNA polymerase structure and function.

110 citations


Journal ArticleDOI
TL;DR: The results of these studies reveal that sequences that bind weakly to hER in vitro are fully functional as EREs in yeast and are conditionally responsive to estrogen in mammalian cells.
Abstract: A powerful and versatile system for the identification of novel response elements for members of the intracellular receptor family is presented as applied to the human estrogen receptor. In the past, a limited number of estrogen response elements (EREs) have been functionally identified in the promoter regions of estrogen-regulated genes. From these a consensus ERE has been defined that is identical to the ERE of the Xenopus laevis vitellogenin gene, i.e., 5'-GGTCA NNN TGACC-3'. In order to investigate without bias the range of sequences that could function as EREs in vivo, we have developed a genetic selection in yeast expressing the human estrogen receptor (hER) and transformed with a random oligonucleotide library in a vector where expression of a selectable marker requires insertion of an upstream activating sequence. More than 1,000,000 transformants were screened and of 726 clones that contained activating sequences, 65 were found to be hormone-dependent. Sequencing revealed that the majority contai...

98 citations


Journal ArticleDOI
TL;DR: It is proposed that the CSRE functions as a UAS element common to genes of the gluconeogenic pathway, similar to the fructose-1,6-bisphosphatase gene FBP1, which is found in coregulated genes involved in gluconeogenesis.
Abstract: The expression of yeast genes encoding gluconeogenic enzymes depends strictly on the carbon source available in the growth medium. We have characterized the control region of the isocitrate lyase gene ICL1, which is derepressed more than 200-fold after transfer of cells from fermentative to nonfermentative growth conditions. Deletion analysis of the ICL1 promoter led to the identification of an upstream activating sequence element, UASICL1 (5' CATTCATCCG 3'), necessary and sufficient for conferring carbon source-dependent regulation on a heterologous reporter gene. Similar sequence motifs were also found in the upstream regions of coregulated genes involved in gluconeogenesis. This carbon source-responsive element (CSRE) interacts with a protein factor, designated Ang1 (activator of nonfermentative growth), detectable only in extracts derived from derepressed cells. Gene activation mediated by the CSRE requires the positively acting derepression genes CAT1 (= SNF1 and CCR1) and CAT3 (= SNF4). In the respective mutants, Ang1-CSRE interaction was no longer observed under repressing or derepressing conditions. Since binding of Ang1 factor to the CSRE could be competed for by an upstream sequence derived from the fructose-1,6-bisphosphatase gene FBP1, we propose that the CSRE functions as a UAS element common to genes of the gluconeogenic pathway.

91 citations


Patent
Joan C. McPherson1, Robert Kay1
11 Jul 1994
TL;DR: In this paper, novel transcription initiation regions that provide for enhanced transcription of a DNA sequence, particularly a plant sequence, are provided. But the authors do not discuss the role of these initiation regions in gene expression.
Abstract: Novel transcription initiation regions that provide for enhanced transcription of a DNA sequence, particularly a plant sequence, are provided.

85 citations


Journal ArticleDOI
TL;DR: Specific protein binding is demonstrated to the short TEF promoter segment carrying the UASrpg homologous sequences, which is an essential promoter element in genes coding for highly expressed components of the translational apparatus.
Abstract: Ashbya gossypii carries only a single gene (TEF) coding for the abundant translation elongation factor 1α. Cloning and sequencing of this gene and deletion analysis of the promoter region revealed an extremely high degree of similarity with the well studied TEF genes of the yeast Saccharomyces cerevisiae including promoter upstream activation sequence (UAS) elements. The open reading frames in both species are 458 codons long and show 88.6% identity at the DNA level and 93.7% identity at the protein level. A short DNA segment in the promoter, between nucleotides -268 and -213 upstream of the ATG start codon, is essential for high-level expression of the A. gossypii TEF gene. It carries two sequences, GCCCATACAT and ATCCATACAT, with high homology to the UASrpg sequence of S. cerevisiae, which is an essential promoter element in genes coding for highly expressed components of the translational apparatus. UASrpg sequences are binding sites for the S. cerevisiae protein TUF, also called RAP1 or GRF1. In gel retardation with A. gossypii protein extracts we demonstrated specific protein binding to the short TEF promoter segment carrying the UASrpg homologous sequences.

84 citations


Journal ArticleDOI
TL;DR: The E6/E7 early promoter of genital human papillomavirus type 18 contains binding sites for the viral regulator E2, tandemly repeated and closely flanked by two crucial promoter elements; the TATA box downstream and an Sp1 binding site upstream.
Abstract: The E6/E7 early promoter (P105) of genital human papillomavirus type 18 contains binding sites for the viral regulator E2, tandemly repeated and closely flanked by two crucial promoter elements; the TATA box downstream and an Sp1 binding site upstream. We showed that binding of purified E2 and Sp1 proteins in vitro to their neighboring sites is mutually exclusive and that Sp1 is displaced by E2. However, this displacement did not result in repression of P105 transcription. In contrast, binding of E2 to its site overlapping the Sp1 binding site activated transcription of P105 derivatives lacking the E2 site most proximal to the TATA box. Surprisingly, a truncated form of E2, deleted of part of the transactivation domain and known as the E2 transcriptional repressor, as well as the E2 DNA-binding domain alone also supported transcription of these P105 derivatives. In the context of P105, the viral E2 protein can thus activate P105 transcription in place of Sp1, even in the absence of its transactivation domain.

83 citations


Journal ArticleDOI
TL;DR: The results obtained suggest that LevR is a multidomain protein, the amino-terminal part of the protein is required for DNA binding whereas the central domain allows the activation of transcription.

76 citations


Journal ArticleDOI
TL;DR: Analysis of the upstream regions of protein-coding genes from Trichomonas vaginalis revealed the presence of a highly conserved DNA sequence motif immediately upstream of the coding region, which is similar, both structurally and functionally, to initiator elements found in promoters of higher eukaryotes.
Abstract: To identify regulatory elements that play a role in transcription initiation in ancient eukaryotes, we have analyzed the upstream regions of protein-coding genes from Trichomonas vaginalis, one of the most ancient eukaryotes studied to date. Characterization of seven protein-coding genes from this protist invariably revealed the presence of a highly conserved DNA sequence motif immediately upstream of the coding region. This 13-nt motif was shown to surround and contain precise sites for transcription initiation. No typical TATA boxes, positioned at 25-30 nt upstream of the transcription start sites of these genes, were found. The start-site regions from all seven T. vaginalis genes impart strong specific initiation of transcription in a mammalian in vitro transcription assay. This consensus promoter element in an ancient eukaryote is similar, both structurally and functionally, to initiator elements found in promoters of higher eukaryotes.

75 citations


Journal ArticleDOI
TL;DR: The characteristics of this novel UAS suggest that it might have a role in initiating CLN2 expression early in G1 to activate the positive feedback loop that drives maximal Cln accumulation, and is both necessary and sufficient for regulated transcription driven by aCLN2 promoter lacking functional SCBs and MCBs.
Abstract: The budding yeast Saccharomyces cerevisiae CLN1, CLN2, and CLN3 genes encode functionally redundant G1 cyclins required for cell cycle initiation. CLN1 and CLN2 mRNAs accumulate periodically throughout the cell cycle, peaking in late G1. We show that cell cycle-dependent fluctuation in CLN2 mRNA is regulated at the level of transcriptional initiation. Mutational analysis of the CLN2 promoter revealed that the major cell cycle-dependent upstream activating sequence (UAS) resides within a 100-bp fragment. This UAS contains three putative SWI4-dependent cell cycle boxes (SCBs) and two putative MluI cell cycle boxes (MCBs). Mutational inactivation of these elements substantially decreased CLN2 promoter activity but failed to eliminate periodic transcription. Similarly, inactivation of SWI4 decreased CLN2 transcription without affecting its periodicity. We have identified a second UAS in the CLN2 upstream region that can promote cell cycle-dependent transcription with kinetics similar to that of the intact CLN2 promoter. Unlike the major CLN2 UAS, this newly identified UAS promotes transcription in cells arrested in G1 by inactivation of cdc28. This novel UAS is both necessary and sufficient for regulated transcription driven by a CLN2 promoter lacking functional SCBs and MCBs. Although this UAS itself contains no SCBs or MCBs, its activity is dependent upon SWI4 function. The characteristics of this novel UAS suggest that it might have a role in initiating CLN2 expression early in G1 to activate the positive feedback loop that drives maximal Cln accumulation.

Journal ArticleDOI
TL;DR: The results strongly suggest that these UASs are targets for transcriptional factors required for assisting specific regulatory proteins.
Abstract: We have initiated a study of the promoter region of the alkaline extracellular protease gene (XPR2) from Yarrowia lipolytica to identify upstream sequences possibly involved in carbon, nitrogen, and peptone control of XPR2 expression. Deletion analysis showed that the TATA box and two major upstream activation sequences (UASs) were essential for promoter activity under conditions of repression or full induction. Within the distal UAS (UAS1), in vivo footprinting studies with dimethyl sulfate (DMS) identified two sequences similar to Saccharomyces cerevisiae GCN4 (-800 to -792)- and TUF/RAP1 (-790 to -778)-binding sites and two sequences which partially overlap a repeated sequence (-778 to -771 and -720 to -713) similar to the CAR1 upstream repression sequence of S. cerevisiae. Oligonucleotides carrying the TUF/RAP1-like-binding site and adjacent downstream nucleotides restored full transcriptional activity of a UAS1-deleted promoter. Within the proximal UAS (UAS2), a directly repeated decameric sequence (-146 to -137 and -136 to -127) was protected against DMS in vivo. Sequences identical to the ABF1-binding site of S. cerevisiae (-121 to -109) or similar to the GCN4-binding site (-113 to -105) were not clearly protected from DMS in vivo. An oligomer (-150 to -106) carrying these three sequences, inserted into a UAS2-deleted promoter, increased the transcriptional activity. The results from footprints under different physiological conditions suggested that protein binding to both UASs was constitutive. Deletion of both UASs greatly reduced XPR2 expression without abolishing its regulation. Our results strongly suggest that these UASs are targets for transcriptional factors required for assisting specific regulatory proteins.

Journal ArticleDOI
TL;DR: An experiment in which the upstream CRP-binding site is replaced by a site for the related transcription factor, FNR, shows that heterologous synergistic interactions between FNR and CRP are possible.

Journal ArticleDOI
TL;DR: A substitution at S73 interferes with FNR‐dependent activation at both this promoter and promoters in which the FNR site is located at 41 1/2 bp from the transcript start, suggesting that FNR may contain a second activating region.
Abstract: Summary We have characterized a number of mutations in fnr that interfere with FNR-dependent transcription activation at two promoters where the FNR-binding site is centred around 41½ bp upstream from the transcription start site. The substituted residues in all but one of these FNR mutants are clustered around a presumed surface-exposed beta-turn containing G85 which, we suggest, forms an activating region that contacts RNA polymerase at these promoters. Using the‘oriented heterodimers’method described elsewhere, we show that this activating region on the promoter-proximal subunit of the FNR dimer is sufficient to activate transcription initiation. In contrast, this region is not essential for activation of a third FNR-dependent promoter where the FNR-binding site is centred at 61 1/2 bp upstream from the transcription start site. However, a substitution at S73 interferes with FNR-dependent activation at both this promoter and promoters in which the FNR site is located at 41 1/2 bp from the transcript start, suggesting that FNR may contain a second activating region.

Journal ArticleDOI
TL;DR: It is hypothesized that the W‐terminal two‐thirds of the DctD receiver domain augments and controls an adjacent subdomain for inhibiting the central domain.
Abstract: Summary Rhizobium meliloti DctD is believed to have three functional domains: an N-terminal, two-component receiver domain; and like other σ54-dependent activators, C-terminal and central domains for DNA binding and transcription activation. We have characterized a progressive series of M-terminal deletions of R meliloti DctD. The N-terminal domain was not needed for binding the dctA upstream activation sequence. Only 25% of the C-terminal end of the receiver domain was needed to significantly inhibit the central domain, and proteins lacking up to 60% of the N-terminal end of the receiver domain were‘inducible’in R. meliloti cells. We hypothesize that the W-terminal two-thirds of the DctD receiver domain augments and controls an adjacent subdomain for inhibiting the central domain.

Journal ArticleDOI
TL;DR: Results indicate that upstream regulatory factors are not required for the in vivo binding of TFIID to the CYC1 promoter and that binding of tfiID to DNA is not necessarily a rate-limiting step in the activation of transcription in cells.
Abstract: Functional transcription initiation complexes can be assembled in vitro without the aid of regulatory factors that bind to upstream activating sequences. However, promoters that lack upstream activating sequences are transcribed poorly if at all in vivo, suggesting that regulatory factors are necessary for the assembly of transcription initiation complexes in cells. To test this possibility, we asked whether the general transcription factor TFIID can bind to a promoter in yeast that lacks upstream activating sequences and is transcriptionally inactive. Analysis of an inactive CYC1 core promoter by high-resolution genomic footprinting revealed efficient binding of TFIID to either of two TATA box elements. Addition of a heat shock element rendered this promoter highly responsive to induction of transcription by heat shock but did not alter the TATA box footprints in the core promoter. Inactivation of all but one TATA box by site-directed mutagenesis did not prevent TFIID from binding to the remaining wild-type TATA box independently of regulatory sequences. These results indicate that upstream regulatory factors are not required for the in vivo binding of TFIID to the CYC1 promoter and that binding of TFIID to DNA is not necessarily a rate-limiting step in the activation of transcription in cells. Differences in chromatin structure may account for why regulatory transcription factors are required for the binding of TFIID to some promoters but not to others.

Journal ArticleDOI
TL;DR: Rules for an upstream activation sequence (UAS) are established for the GAL4, RAP1 (RPG box), GCN4, and the HAP2/HAP3/ HAP4 regulatory proteins, as well as for a motif (PAC) frequently found upstream of the genes of the RNA polymerase A and C subunits.
Abstract: The systematic sequencing of the yeast genome reveals the presence of many potential genes of unknown function. One way to approach their function is to define which regulatory system controls their transcription. This can also be accomplished by the detection of an upstream activation sequence (UAS). Such a detection can be done by computer, provided that the definition of a UAS includes sufficient and precise rules. We have established such rules for the UASs of the GAL4, RAP1 (RPG box), GCN4, and the HAP2/HAP3/HAP4 regulatory proteins, as well as for a motif (PAC) frequently found upstream of the genes of the RNA polymerase A and C subunits. These rules were applied to the chromosome III DNA sequence, and gave precise predictions.

Journal ArticleDOI
TL;DR: Transcription of yeast fatty acid synthase genes is subjected to both the pathway-specific control affecting genes of phospholipid biosynthesis and to the activation by general transcription factors allowing a sufficiently high level of constitutive gene expression.
Abstract: The fatty acid synthase genes FAS1 and FAS2 of the yeast Saccharomyces cerevisiae are under transcriptional control of pathway-specific regulators of phospholipid biosynthesis. However, site-directed mutagenesis of the respective cis-acting elements upstream of FAS1 and FAS2 revealed that additional sequences activating both genes must exist. A deletion analysis of the FAS1 promoter lacking the previously characterized inositol/choline-responsive-element motif defined a region (nucleotides -760 to -850) responsible for most of the remaining activation potency. Gel-retardation experiments and in-vitro DNase footprint studies proved the binding of the general regulatory factors Rap1p, Abf1p and Reb1p to this FAS1 upstream region. Mutation of the respective binding sites led to a drop of gene activation to 8% of the wild-type level. Similarly, we also demonstrated the presence of a Reb1p-binding site upstream of FAS2 and its importance for gene activation. Thus, in addition to the previously characterized FAS-binding factor 1 interacting with the inositol/choline-responsive-element motif, a second motif common to the promoter regions of both FAS genes could be identified. Transcription of yeast fatty acid synthase genes is therefore subjected to both the pathway-specific control affecting genes of phospholipid biosynthesis and to the activation by general transcription factors allowing a sufficiently high level of constitutive gene expression.

Journal ArticleDOI
TL;DR: Results suggest that rather short sequences between nucleotides -103 and -72 from the transcription start site are associated with the specific expression of HrMA4 alpha.

Journal ArticleDOI
TL;DR: A physical assay was used to show that Ty3 did not transpose when yeast cells were arrested in G1 during treatment with the mating pheromone alpha-factor, and transposition was detected within one generation of cell growth after Ty3 transcription was initiated.
Abstract: Host cell cycle genes provide important functions to retroviruses and retroviruslike elements. To define some of these functions, the cell cycle dependence of transposition of the yeast retroviruslike element Ty3 was examined. Ty3 is unique among retroviruslike elements because of the specificity of its integration, which occurs upstream of genes transcribed by RNA polymerase III. A physical assay for Ty3 transposition which takes advantage of this position-specific integration was developed. The assay uses PCR to amplify a product of Ty3 integration into a target plasmid that carries a modified tRNA gene. By using the GAL1 upstream activating sequence to regulate expression of Ty3, transposition was detected within one generation of cell growth after Ty3 transcription was initiated. This physical assay was used to show that Ty3 did not transpose when yeast cells were arrested in G1 during treatment with the mating pheromone alpha-factor. The restriction of transposition was not due to changes in transcription of either Ty3 or tRNA genes or to aspects of the mating pheromone response unrelated to cell cycle arrest. The block of the Ty3 life cycle was reversed when cells were released from G1 arrest. Examination of Ty3 intermediates during G1 arrest indicated that Ty3 viruslike particles were present but that reverse transcription of the Ty3 genomic RNA into double-stranded DNA had not occurred. In G1, the Ty3 life cycle is blocked after particle assembly but before the completion of reverse transcription.

Patent
Joan C. McPherson1, Robert Kay1
09 Mar 1994
TL;DR: In this paper, novel transcription initiation regions that provide for enhanced transcription of a DNA sequence, particularly a plant sequence, are provided. But the authors do not discuss the role of these initiation regions in gene expression.
Abstract: Novel transcription initiation regions that provide for enhanced transcription of a DNA sequence, particularly a plant sequence, are provided.

Journal ArticleDOI
TL;DR: The discovery of the NRE indicates that HBV gene transcription is controlled by combined effects of both positive and negative regulation and provides a unique system with which to study the mechanism of negative regulation of gene expression.
Abstract: Enhancer II of human hepatitis B virus has dual functions in vivo. Located at nucleotides (nt) 1646 to 1741, it can stimulate the surface and X promoters from a downstream position. Moreover, the same sequence can also function as upstream regulatory element that activates the core promoter in a position- and orientation-dependent manner. In this study, we report the identification and characterization of a negative regulatory element (NRE) upstream of enhancer II (nt 1613 to 1636) which can repress both the enhancer and upstream stimulatory function of the enhancer II sequence in differentiated liver cells. This NRE has marginal inhibitory effect by itself but a strong repressive function in the presence of a functional enhancer II. Mutational analysis reveals that sequence from nt 1616 to 1621 is required for repression of enhancer activity by the NRE. Gel shift analysis reveals that this negative regulatory region can be recognized by a specific protein factor(s) present at the 0.4 M NaCl fraction of HepG2 nuclear extracts. The discovery of the NRE indicates that HBV gene transcription is controlled by combined effects of both positive and negative regulation. It also provides a unique system with which to study the mechanism of negative regulation of gene expression.

Journal ArticleDOI
TL;DR: The results suggest that CPF1 functions to modulate chromatin structure around the CDEI motif but that these changes at the MET25 and MET16 promoters do not explain how CPF 1 functions to maintain methionine-independent growth.
Abstract: CPF1 is an abundant basic-helix-loop-helix-ZIP protein that binds to the CDEI motif in Saccharomyces cerevisiae centromeres and in the promoters of numerous genes, including those encoding enzymes of the methionine biosynthetic pathway. Strains lacking CPF1 are methionine auxotrophs, and it has been proposed that CPF1 might positively influence transcription at the MET25 and MET16 genes by modulating promoter chromatin structure. We test this hypothesis and show that the regions surrounding the CDEI motifs in the MET25 and MET16 promoters are maintained in a nucleosome-free state and that this requires the entire CPF1 protein. However, the chromatin structure around the CDEI motifs does not change on derepression of transcription and does not correlate with the methionine phenotype of the cell. An intact CDEI motif but not CPF1 is required for transcriptional activation from a region of the MET25 upstream activation sequence. Our results suggest that CPF1 functions to modulate chromatin structure around the CDEI motif but that these changes at the MET25 and MET16 promoters do not explain how CPF1 functions to maintain methionine-independent growth. The presence of CPF1-dependent chromatin structures at these promoters leads to a weak repression of transcription.

Journal ArticleDOI
TL;DR: It is demonstrated that NarL‐phosphate, produced by the reaction of purified NarL with acetyl phosphate, specifically binds to a fragment derived from the upstream region of the narG promoter, which appears to result from the formation of a folded protein‐DNA structure created by the binding of NarL-phosphates to multiple sites on either side of an IHF‐induced bend in the upstream regions of the promoter.
Abstract: The stimulation of Fnr-dependent transcription from the narG promoter by NarL-phosphate is known to require a cis-acting sequence, the NarL box, located approximately 195 bp upstream from the transcription start site, and the interaction of integration host factor (IHF) with a binding site in the intervening region (positions -110 to -140) between the NarL box and the transcription start site. By gel retardation and DNase I protection studies, we have demonstrated that NarL-phosphate, produced by the reaction of purified NarL with acetyl phosphate, specifically binds to a fragment derived from the upstream region of the narG promoter. The fragment was protected by NarL-phosphate binding to two distinct regions. One was an extended sequence of approximately 40 bp surrounding the NarL box at -195; the second was located downstream from the IHF-binding region and included a sequence extending from positions -80 to -120. Alteration by site-directed mutagenesis of a putative inverted NarL box sequence identified within the downstream protected region in a plasmid containing a narG-lacZ fusion eliminated the NarL-phosphate-mediated stimulation of transcription. NarL-phosphate bound to the two regions independently from IHF binding and it bound to each site independently when the two sites were separated by cleavage of the promoter fragment. Stimulation of transcription from the narG promoter by NarL-phosphate appears to result from the formation of a folded protein-DNA structure created by the binding of NarL-phosphate to multiple sites on either side of an IHF-induced bend in the upstream region of the promoter.

Journal ArticleDOI
TL;DR: A refined functional map of the upstream regulatory element reveals a complex combinatorial code that directs expression of the human alpha subunit gene to placenta.

Journal ArticleDOI
TL;DR: The isolation of seven mutations in the ADR1-5c allele, defining five different amino acid changes, that suppress the enhanced ADH2 expression caused by the ADr1- 5c allele indicate that the DNA-binding region ofADR1 is involved in both transactivation and DNA binding.

Journal ArticleDOI
01 Oct 1994-Yeast
TL;DR: Novel mutants, tye7, are identified, which are affected in Ty1‐mediated expression of ADH2 through a Ty1 sequence distal to the 5′ long terminal repeat sequence.
Abstract: In Saccharomyces cerevisiae, expression of a gene adjacent to the retrotransposon Ty1 is often mediated by Ty-internal sequences. We have identified novel mutants, tye7, which are affected in Ty1-mediated expression of ADH2 through a Ty1 sequence distal to the 5' long terminal repeat sequence. The TYE7 gene has been isolated and characterized. It encodes a 33 kDa protein whose N-terminal third is extremely rich in serine residues (28%). Within its C-terminal sequence, a remarkable similarity to Myc and Max proteins can be found. Thus, TYE7 is a potential member of the basic region/helix-loop-helix/leucine-zipper protein family. TYE7 function is not essential for growth. It may primarily function as a transcriptional activator in Ty1-mediated gene expression, as has been confirmed by the activation of reporter gene expression by a LexA-TYE7 hybrid protein. ADH2 activation by defined Ty1 derivatives revealed that TYE7 acts positively through the more distal Ty1 enhancer element (region D), and negatively in a region between A (the 5' proximal enhancer element) and D.

Journal ArticleDOI
TL;DR: The promoter elements in the Dictyostelium actin 15 and actin 6 genes required for full growth phase expression were identified by assaying promoter/luciferase reporter constructs and it is found that these promoters contain common cis-acting elements, an actin upstream activating sequence (UAS) and sequences proximal to the transcription start site that overlap with a poly(dT) region.
Abstract: The promoter elements in the Dictyostelium actin 15 and actin 6 genes required for full growth phase expression were identified by assaying promoter/luciferase reporter constructs. We find that these promoters contain common cis-acting elements, an actin upstream activating sequence (UAS) and sequences proximal to the transcription start site that overlap with a poly(dT) region. The actin 15 promoter has two additional cis-acting elements not present in the actin 6 promoter that may account for the higher level of expression from the actin 15 promoter. All of the identified promoter elements are unusual for Dictyostelium in that they are all A/T-rich. Two cis-acting elements, the actin UAS and the poly(dT) domain were studied in greater detail. The actin UAS was tested on a heterologous promoter from the prespore-specific gene SP60 and shown to have the ability to confer growth phase expression. The actin UAS also exhibited the ability to function in a distance- and orientation-independent manner and activate expression synergistically when present in two copies. The poly(dT) domain of the actin 15 promoter was studied in greater detail by using a genetic selection scheme to define parameters that effect the strength of this element. This element is comprised of 45 consecutive dT residues immediately upstream of the putative TATA box. We show that the length of the homopolymer dT region correlates with the expression level of the promoter. The poly(dT) element is also shown to function to promote wild-type levels of expression with small deviations in the sequence, indicating that the element is not required to be homopolymeric to function.

Journal ArticleDOI
TL;DR: Three cis-acting elements in the POX1 upstream sequence are characterized, two upstream repression sequences and an upstream activating sequence that was able to regulate the transcription of a heterologous gene construct iso-1-cytochrome-c (CYC1)/lacZ.

Journal ArticleDOI
TL;DR: RNA polymerase is able to bend a DNA fragment containing PctII, suggesting that an increase in the RNA polymerase-DNA contacts is an important step in transcription initiation, and the upstream activating region could be substituted by targets of unrelated DNA-bending proteins, supporting the role of curved DNA as a transcriptional modulator.