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S Wölfl

Publications and source records attributed to S Wölfl.

26 records · Page 2Linked to original sources
PubMed1996

Two glucose transporters in Saccharomyces cerevisiae are glucose sensors that generate a signal for induction of gene expression.

Glucose is the preferred carbon source for most eukaryotic cells and has profound effects on many cellular functions. How cells sense glucose and transduce a signal into the cell is a fundamental, unanswered question. Here we describe evidence that two unusual glucose transporters in the yeast Saccharomyces cerevisiae serve as glucose sensors that generate an intracellular glucose signal. The Snf3p high-affinity glucose transporter appears to function as a low glucose sensor, since it is required for induction of expression of several hexose transporter (HXT) genes, encoding glucose transporters, by low levels of glucose. We have identified another apparent glucose transporter, Rgt2p, that is strikingly similar to Snf3p and is required for maximal induction of gene expression in response to high levels of glucose. This suggests that Rgt2p is a high glucose-sensing counterpart to Snf3p. We identified a dominant mutation in RGT2 that causes constitutive expression of several HXT genes, even in the absence of the inducer glucose. This same mutation introduced into SNF3 also causes glucose-independent expression of HXT genes. Thus, the Rgt2p and Snf3p glucose transporters appear to act as glucose receptors that generate an intracellular glucose signal, suggesting that glucose signaling in yeast is a receptor-mediated process.

Amino Acid Sequence↗
PubMed1996

Transcription of the human corticotropin-releasing hormone gene in NPLC cells is correlated with Z-DNA formation.

The intron of the corticotropin-releasing hormone (corticoliberin; CRH) gene contains a sequence of over 100 bp of alternating purine/pyrimidine residues. We have used binding of a Z-DNA-specific antibody in metabolically active, permeabilized nuclei to study the formation of Z-DNA in this sequence at various levels of transcription. In the NPLC human primary liver carcinoma cell line, activation of cAMP-dependent pathways increased the level of transcription, while adding glucocorticoids inhibited transcription of the CRH gene. These cells respond in a manner similar to hypothalamic cells. Z-DNA formation in this sequence was detected at the basal level of transcription, as well as after stimulation with forskolin. Inhibition of transcription by dexamethasone abolished Z-DNA formation. Z-DNA formation in the WC gene (c-myc) was affected differently in the same experiment. Thus, changes in Z-DNA formation in the CRH gene are gene specific and are linked to the transcription of the gene.

Base Sequence↗
PubMed1996

Analysis of a 26,756 bp segment from the left arm of yeast chromosome IV.

The nucleotide sequence of a 26.7 kb DNA segment from the left arm of Saccharomyces cerevisiae chromosome IV is presented. An analysis of this segment revealed 11 open reading frames (ORFs) longer than 300 bp and one split gene. These ORFs include the genes encoding the large subunit of RNA polymerase II, the biotin apo-protein ligase, an ADP-ribosylation factor (ARF 2), the 'L35'-ribosomal protein, a rho GDP dissociation factor, and the sequence encoding the protein phosphatase 2A. Further sequence analysis revealed a short ORF encoding the ribosomal protein YL41B, an intron in a 5' untranslated region and an extended homology with another cosmid (X83276) located on the same chromosome. The potential biological relevance of these findings is discussed.

ADP-Ribosylation Factors↗
PubMed1995

Identification of transcriptionally induced Z-DNA segments in the human c-myc gene.

Using monoclonal antibodies against Z-DNA three AluI restriction fragments of the human c-myc gene were previously found to form Z-DNA in agarose-embedded, metabolically active permeabilized nuclei. The formation of Z-DNA in these fragments was dependent on negative supercoiling generated by transcription of the gene. Here we show which sequence elements of these three AluI restriction fragments adopt the Z conformation upon negative supercoiling. The three fragments (Z1, Z2 and Z3) were inserted in a suitable plasmid vector. Z-DNA forming elements were detected by comparing DEPC reactivity in relaxed circular and supercoiled plasmid DNA. Z1 and Z3 each contained one major Z-DNA forming region 20-25 nucleotides long, whereas Z2 contained two discrete regions 90 nucleotides apart one about 35 nucleotides the other about 20 nucleotides long.

Base Sequence↗
PubMed1992

Transcription of human c-myc in permeabilized nuclei is associated with formation of Z-DNA in three discrete regions of the gene.

When human U937 cells are placed in agarose microbeads and treated with a detergent, the cytoplasmic membrane is lysed and the nuclear membrane is permeabilized. However, the nuclei remain intact and maintain both replication and transcription. Biotin labeled monoclonal antibodies against Z-DNA have been diffused into this system and used to measure the amount of Z-DNA present in the nuclei. It has previously been shown that the amount of Z-DNA present decreases due to relaxation by topoisomerase I and increases as the level of transcription increases. Here we measure the formation of Z-DNA in the c-myc gene by crosslinking the antibodies to DNA using laser radiation at 266 nm for 10 ns. The crosslinked DNA is isolated by restriction digestion, separation of antibody labeled fractions through the biotin residue, and subsequent proteolysis to remove the crosslinked antibody. Three AluI restriction fragments of the c-myc gene are shown to form Z-DNA when the cell is transcribing c-myc. The Z-DNA forming segments are near the promoter regions of the gene. However, when U937 cells start to differentiate and transcription of the c-myc gene is down-regulated, the Z-DNA content goes to undetectable levels within 30-60 min.

Base Sequence↗
PubMed1991

Lack of correlation between DNA methylation and transcriptional inactivation: the chicken lysozyme gene.

We have analyzed the methylation state of all nine CpG sites in the transcription start region (-420 to +250 base pairs) of the chicken lysozyme gene by genomic sequencing. One of these sites, at -81, lies within the promoter, seven are clustered within the first exon, and the last is in the first intron. Five cell types and tissues have been investigated to study the relationship between methylation level and gene expression. For each cell type used, the majority of CpG sites showed a similar level of methylation. Of two gene-nonexpressing tissues, erythrocytes are hypomethylated, whereas liver is methylated at most of its CpG sites. For gene-expressing tissues, oviduct is completely unmethylated, whereas HD-11 culture cells are methylated. Thus no correlation is observed between degree of CpG methylation and level of expression of the lysozyme gene. The observed methylation patterns are discussed in terms of possible features of the local chromatin structure.

Animals↗
PubMed1990

Direct determination of chloramphenicol acetyltransferase (CAT) activity in homogenates of transfected mammalian cells by high performance liquid chromatography.

The conversion of chloramphenicol to its monoacetylated form by homogenates of pRSV CAT-transfected mammalian cells was assayed by HPLC. The method differed from conventional procedures, in that the extraction of chloramphenicol and its acetylated forms into an organic solvent was replaced by treatment of the cell homogenates with acetonitrile. This allows a rapid, direct analysis by HPLC with few experimental steps.

Acetonitriles↗