Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.23 (beta-galactosidase)
14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The LPD1 gene of Saccharomyces cerevisiae, encoding lipoamide dehydrogenase (LPDH), is subject to catabolite repression. The promoter of this gene contains a number of motifs for DNA-binding transcriptional activators, including three which show strong sequence homology to the core HAP2/HAP3/HAP4 binding motif. Here we report that transcription of LPD1 requires HAP2, HAP3 and HAP4 for release from glucose repression. In the wild-type strain, specific activity of LPDH was increased 12-fold by growth on lactate, 10-fold on glycerol and four- to five-fold on galactose or raffinose, compared to growth on glucose. In hap2, hap3 and hap4 null mutants, the specific activities of LPDH in cultures grown on galactose and raffinose showed only slight induction above the basal level on glucose medium. Similar results were obtained upon assaying for beta-galactosidase production in wild-type, or hap2, hap3 or hap4 mutant strains carrying a single copy of the LPD1 promoter fused in frame to the lacZ gene of Escherichia coli and integrated at the URA3 locus. Transcript analysis in wild-type and hap2 mutants confirmed that the HAP2 protein regulates LPD1 expression at the level of transcription in the same way as it does for the CYC1 gene. Site-directed mutagenesis of the putative HAP2/HAP3/HAP4 binding site at -204 relative to the ATG start codon showed that this element was required for full derepression of the LPD1 gene on non-fermentable substrates.
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PMID:Positive regulation of the LPD1 gene of Saccharomyces cerevisiae by the HAP2/HAP3/HAP4 activation system. 131 May 23

In a previous paper we have studied the expression of beta-galactosidase from Escherichia coli, driven from the inducible GAL1-10/CYC1 hybrid promoter, in batch cultures of budding Saccharomyces cerevisiae and have described operating conditions for maximal productivity. In this paper we show that the plasmid instability in continuous cultures can be overcome by utilizing appropriate selection markers and a high copy number vector. The maximal level of expression is influenced by the dilution rate. Moreover, enzyme accumulation appears to depend also upon the degree of oxygenation. A possible explanation of these modulations is discussed, taking into account the interactions of the UAS-GAL and TATA-CYC1 elements.
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PMID:Heterologous gene expression in continuous cultures of budding yeast. 136 26

We have previously shown that nucleosome loss, obtained by repressing histone H4 mRNA synthesis, activates otherwise inactive PHO5, GAL1, and CYC1 gene promoters (fused to the bacterial beta-galactosidase [lacZ] reporter gene) to moderate levels of activity (approximately 2 to 15% of fully induced levels). We now report that nucleosome loss activates the expression of two additional promoters that are normally induced by independent mechanisms: CUP1 (induced by heavy-metal toxicity) and HIS3 (induced by amino acid starvation). Surprisingly, the level of CUP1-lacZ and HIS3-lacZ activation by nucleosome loss approximates fully induced levels of transcription. These CUP1 and HIS3 promoter activities are increased similarly from either episomal or genomic constructs. Our results emphasize the universality of the mechanism by which nucleosome loss activates yeast promoters. Moreover, a comparison of absolute levels of activation for different promoters suggests that activation by nucleosome loss results in a relatively constant level of activation, while levels obtained by normal induction vary considerably. These data argue that nucleosome loss may play a uniquely dominant role in the regulation of certain promoters.
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PMID:Nucleosome loss activates CUP1 and HIS3 promoters to fully induced levels in the yeast Saccharomyces cerevisiae. 154 16

To assess thyroid hormone receptor (TR)-mediated activation of transcription in yeast in the presence or absence of thyroid hormone (T3), we developed a co-expression system using a TR-beta 1 expression vector and a reporter plasmid containing a 16 base pair palindromic thyroid hormone response element (TRE) upstream from a proximal CYC1 promoter that was fused to the beta-galactosidase lac Z gene of Escherichia coli. Although TR-beta 1 functions as a repressor in most mammalian systems, using our system we observed a unique thyroid hormone-independent transcriptional response indicating that wild TR-beta 1 acted as a constitutive activator in yeast; the addition of 1 microM T3 induced a moderate but significant (p less than 0.01) 25-40% further increase in transcriptional activity. Using a series of rat TR-beta 1 mutant constructs, we found that deletion of domain D and portions of E completely eliminated transcriptional activity, whereas truncations of domain F and E permitted a partial (20-40%) response compared to wild TR-beta 1 in the presence or absence of T3. These observations indicate that TR-beta 1 functions as an activator in yeast and that domains D,E and F play important interactive roles in its hormone-independent gene activation with the D domain likely being the most essential. Furthermore, our results suggest that the different transcriptional property of TR-beta 1 in yeast compared to mammalian cells i.e. activator vs repressor function, is likely determined by transcriptional factor differences which are dependent upon cellular origin.
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PMID:Rat liver c-erb A beta 1 thyroid hormone receptor is a constitutive activator in yeast (Saccharomyces cerevisiae): essential role of domains D,E and F in hormone-independent transcription. 165 14

The ROX3 gene was identified during a hunt for mutants with increased expression of the heme-regulated CYC7 gene, which encodes the minor species of cytochrome c in the yeast Saccharomyces cerevisiae. The rox3 mutants caused a 10-fold increase in CYC7 expression both in the presence and absence of heme, had slightly increased anaerobic expression of the heme-activated CYC1 gene, and caused decreases in the anaerobic expression of the heme-repressed ANB1 gene and the aerobic expression of its heme-induced homolog. The wild-type ROX3 gene was cloned, and the sequence indicated that it encodes a 220-amino-acid protein. This protein is essential; deletion of the coding sequence was lethal. The coding sequence for beta-galactosidase was fused to the 3' end of the ROX3 coding sequence, and the fusion product was found to be localized in the nucleus, strongly suggesting that the wild-type protein carries out a nuclear function. Mutations in the rox3 gene showed an interesting pattern of intragenic complementation. A deletion of the 5' coding region complemented a nonsense mutation at codon 128 but could not prevent the lethality of the null mutation. These results suggest that the amino-terminal domain is required for an essential function, while the carboxy-terminal domain can be supplied in trans to achieve the wild-type expression of CYC7. Finally, RNA blots demonstrated that the ROX3 mRNA was expressed at higher levels anaerobically but was not subject to heme repression. The nuclear localization and the lack of viability of null mutants suggest that the ROX3 protein is a general regulatory factor.
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PMID:The ROX3 gene encodes an essential nuclear protein involved in CYC7 gene expression in Saccharomyces cerevisiae. 165 37

The RAP1 gene of Saccharomyces cerevisiae encodes an abundant DNA-binding protein, also known as GRF1, TBA, or TUF, that binds to many sites in the yeast genome in vitro. These sites define a consensus sequence, [sequence: see text], and deletion analyses of genes that contain this sequence have implicated the involvement of RAP1 in numerous cellular processes, including gene activation and repression. The MAT alpha locus, required for determination of the alpha cell type in yeast cells, contains a RAP1 binding site; this site coincides with the MAT alpha upstream activating sequence (UAS) and is necessary for expression of the two genes encoded by the MAT alpha locus, MAT alpha 1 and MAT alpha 2. We show that the MAT alpha UAS is sufficient to activate transcription from a promoterless gene fusion of the yeast CYC1 upstream region and the lacZ gene. Constructs containing only the MAT alpha UAS generated elevated levels of beta-galactosidase activity which were indistinguishable from those of constructs containing the entire MAT alpha intergenic region. Further, the MAT alpha UAS has an intrinsic polarity of transcriptional activation; transcription of CYC1-lacZ was six- to sevenfold higher when the UAS was oriented in the direction normally associated with MAT alpha 2 transcription. Point mutations in the MAT alpha UAS that reduce MAT alpha expression three- to fivefold resulted in a bi-mating phenotype, while a mutation that reduced MAT alpha expression still further resulted in an a-mating phenotype. We isolated plasmids from a high-copy-number yeast library that suppressed the bi-mating defect of point mutations in the MAT alpha UAS, and the most effective dosage suppressor contained the gene encoding RAP1. A temperature-sensitive rap1 mutant bi-mates at the semipermissive temperature. Double mutants at rap1 and mat alpha mate exclusively as a cells, at all temperatures, and do not express detectable levels of MAT alpha RNA. These data provide evidence that the RAP1 gene product functions at the MAT alpha UAS in vivo.
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PMID:The role of RAP1 in the regulation of the MAT alpha locus. 199 Feb 67

Saccharomyces cerevisiae chromosomes end with the sequence C2-3A(CA)1-4, commonly abbreviated as C1-3A. These sequences can function as upstream activators of transcription (UAS's) when placed in front of a CYC1-lacZ fusion gene. When C1-3A sequences are placed between the GAL1,10 UAS and the CYC1-lacZ fusion, the C1-3A UAS still functions and the amount of beta-galactosidase produced in cells grown on glucose is as much or more than that for cells grown on either glycerol medium, or cells grown on glucose medium containing a plasmid with just the C1-3A UAS. These data indicate that the UAS is immune from glucose repression from the upstream GAL1,10 UAS. Because C1-3A sequences are bound in vitro by the transcription factor RAP1, the UAS activity of yeast telomere sequences was compared with that of a similar UAS from the tightly regulated ribosomal protein gene RP39A, which also contains a RAP1 binding site. While transcription from the ribosomal protein gene UAS was responsive to cell density, the amount of transcription from the C1-3A UAS was nearly the same at all cell densities tested. These data show that the transcriptional activation by C1-3A sequences is not regulated by cell density.
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PMID:Properties of the transcriptional enhancer in Saccharomyces cerevisiae telomeres. 211 Jun 55

Ten fragments of higher eucaryotic DNA were tested for upstream activation sequence activity in Saccharomyces cerevisiae by inserting them upstream of a CYC1::lacZ promoter lacking an upstream activation sequence. Fragments containing the 21-base-pair repeat region, the enhancer of simian virus 40 or both strongly stimulated beta-galactosidase synthesis, and three fragments from the polyomavirus enhancer region stimulated moderate levels. Three of the four controls of random DNA sequences failed to stimulate significant levels, and the fourth stimulated moderate levels. The stimulation in all cases was independent of the orientation of the inserted fragment. Two series of clones were examined in which between one and six tandemly arranged copies of a fragment were inserted into the XhoI site of the vector. Very interestingly, we detected an apparent exponential relationship between the number of copies of a fragment and the amount of beta-galactosidase produced. Southern analysis showed that increases in enzyme activity were not a result of increased plasmid copy number. Rather, quantitative S1 nuclease analysis demonstrated that the increases were correlated with steady-state levels of lacZ-specific mRNA. We suggest that there may be an evolutionary relationship between some transcriptional activation sequences in yeast cells and the higher eucaryotic regulatory elements that we tested.
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PMID:Enhancer and promoter elements from simian virus 40 and polyomavirus can substitute for an upstream activation sequence in Saccharomyces cerevisiae. 215 86

To determine whether the 70-kilodalton heat shock proteins of Saccharomyces cerevisiae play a role in regulating their own synthesis, we studied the effect of overexpressing the SSA1 protein on the activity of the SSA1 5'-regulatory region. The constitutive level of Ssa1p was increased by fusing the SSA1 structural gene to the GAL1 promoter. A reporter vector consisting of an SSA1-lacZ translational fusion was used to assess SSA1 promoter activity. In a strain producing approximately 10-fold the normal heat shock level of Ssa1p, induction of beta-galactosidase activity by heat shock was almost entirely blocked. Expression of a transcriptional fusion vector in which the CYC1 upstream activating sequence of a CYC1-lacZ chimera was replaced by a sequence containing a heat shock upstream activating sequence (heat shock element 2) from the 5'-regulatory region of SSA1 was inhibited by excess Ssa1p. The repression of an SSA1 upstream activating sequence by the SSA1 protein indicates that SSA1 self-regulation is at least partially mediated at the transcriptional level. The expression of another transcriptional fusion vector, containing heat shock element 2 and a lesser amount of flanking sequence, is not inhibited when Ssa1p is overexpressed. This suggests the existence of an element, proximal to or overlapping heat shock element 2, that confers sensitivity to the SSA1 protein.
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PMID:Self-regulation of 70-kilodalton heat shock proteins in Saccharomyces cerevisiae. 218 Dec 81

Maltose fermentation in Saccharomyces species requires the presence of at least one of five unlinked MAL loci: MAL1, MAL2, MAL3, MAL4, and MAL6. Each of these loci consists of a complex of genes involved in maltose metabolism; the complex includes maltase, a maltose permease, and an activator of these genes. At the MAL6 locus, the activator is encoded by the MAL63 gene. While the MAL6 locus has been the subject of numerous studies, the binding sites of the MAL63 activator have not been determined. In this study, we used Escherichia coli extracts containing the MAL63 protein to define the binding sites of the MAL63 protein in the divergently transcribed MAL61-62 promotor. When a DNA fragment containing these sites was placed upstream of a CYC1-lacZ gene, maltose induced beta-galactosidase. These sites therefore constitute an upstream activating sequence for the MAL genes.
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PMID:Identification of the upstream activating sequence of MAL and the binding sites for the MAL63 activator of Saccharomyces cerevisiae. 219 62


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