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V M Williamson

Publications and source records attributed to V M Williamson.

31 records · Page 2Linked to original sources
PubMed1987

Homology of Saccharomyces cerevisiae ADH4 to an iron-activated alcohol dehydrogenase from Zymomonas mobilis.

Insertion of the transposable element Ty at the ADH4 locus results in increased levels of a new alcohol dehydrogenase (ADH) activity in Saccharomyces cerevisiae. The DNA sequence of this locus has been determined. It contains a long open reading frame which is not homologous to the other ADH isozymes that have been characterized in S. cerevisiae nor does it show obvious homology to Drosophila ADH. The hypothetical ADH does, however, show strong homology to the sequence of an iron-activated ADH from the bacterium Zymomonas mobilis. Thus ADH4 appears to encode an ADH structural gene which, along with the Zymomonas enzyme, may define a new family of alcohol dehydrogenases.

Aeromonas↗
PubMed1987

Purification and properties of Acid phosphatase-1 from a nematode resistant tomato cultivar.

In tomato the acid phosphatase-1 isozyme (Apase-1) is inherited as a single locus linked to the nematode resistance gene (Mi). The Apase-1(1) electrophoretic variant has been purified from a tomato cell suspension culture using ion exchange and concanavalin A sepharose affinity chromatography. A cellulose acetate electrophoresis method was used to distinguish Apase-1(1) rapidly from other Apase isozymes in tomato. The subunit molecular weight of the purified enzyme was estimated to be 31,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The native size of the enzyme, which is reported to be a dimer, was determined to be approximately 51,000 by high performance liquid chromatography gel filtration. Apase-1(1) has a lower pH optimum and a distinct substrate specificity as compared to Apases extracted from tomato fruit or from other plant species. The amino acid composition of Apase-1(1) is similar to that of a potato Apase.

Journal Article↗
PubMed1986

Resistance to antimycin A in yeast by amplification of ADH4 on a linear, 42 kb palindromic plasmid.

A yeast strain lacking a functional copy of ADH1 has been isolated that is resistant to antimycin A because of the presence of multiple copies of a nuclear gene, ADH4. The amplified copies of ADH4 exist on linear molecules 42 kb in length, which can be separated from chromosomal DNA by orthogonal-field-alternation gel electrophoresis. These amplified molecules are palindromes that reanneal rapidly after denaturation to form linear, snap-back molecules 21 kb in length. The amplified ADH4 sequences are bounded by telomere-homologous sequences. The chromosomal copy of ADH4 is the most distal marker on the left arm of chromosome VII, and the amplified ADH4-containing molecules appear to contain two copies of the region extending from ADH4 to the telomere.

Alcohol Dehydrogenase↗
PubMed1986

Ty insertions at two loci account for most of the spontaneous antimycin A resistance mutations during growth at 15 degrees C of Saccharomyces cerevisiae strains lacking ADH1.

The mutation rate to antimycin A resistance was determined for strains of Sacchromyces cerevisiae lacking a functional copy of the structural gene for alcohol dehydrogenase I (ADH1). One type of mutation that can cause antimycin A resistance in these strains is insertion of the transposable element Ty 5' to ADH2, the structural gene for the glucose-repressed isozyme of alcohol dehydrogenase, resulting in expression of this gene during growth on glucose. Here we show that after growth at 15 or 20 degrees C on glucose, 30% of the antimycin A resistance mutations are Ty insertions at ADH2 and another 65% of the mutations are Ty insertions at ADH4, a new locus identified and cloned as described in this paper. At 30 degrees C only 6% of the mutations are Ty insertions at either of these two loci. In addition, we show that the transposition rate is lower in mating-incompetent (a/alpha) cells than in either haploid or diploid mating-competent cells. Our results suggest that under certain conditions Ty transposition may be a major cause of spontaneous mutations in S. cerevisiae.

Alcohol Dehydrogenase↗
PubMed1983

Nucleotide sequence of the yeast alcohol dehydrogenase II gene.

The complete nucleotide sequence of the glucose-repressed alcohol dehydrogenase II gene (ADR2) from yeast has been established together with its 5'- and 3'-flanking regions. The limits of the gene have been determined by reverse transcriptase sequencing of the 5'-ends and by S1 nuclease mapping of the 3'-ends of the mature ADR2 mRNA. Comparison of the alcohol dehydrogenase I gene (ADC1) sequence (Bennetzen, J. L., and Hall, B.D. (1982) J. Biol. Chem. 257, 3018-3025) with that of ADR2 indicated four regions of sequence conservation in the 5'-flanking DNAs. One of these conserved regions contains the sequence TCAAG which may function as a yeast cap sequence. The coding sequence of ADR2 is 89% homologous with that of ADC1 and exhibits a bias in its codon utilization. Evidence is presented that the intergenic region at the 3'-end of the ADR2 gene is less than 550 base pairs.

Alcohol Dehydrogenase↗
PubMed1983

Characterization of transposable element-associated mutations that alter yeast alcohol dehydrogenase II expression.

Seven cis-dominant, constitutively expressed mutations of the normally glucose-repressible isozyme of alcohol dehydrogenase (ADHII) from the yeast Saccharomyces cerevisiae are caused by insertion of transposable elements from the Ty1 family in front of the ADHII structural gene (ADR2) (V. M. Williamson, E. T. Young, and M. Ciriacy, Cell 23:605-614, 1981). We cloned ADR2 with its associated Ty1 element from five S. cerevisiae strains carrying these mutations. Comparison of the Ty1 elements by heteroduplex studies and restriction enzyme analyses indicated that four were very similar; the fifth, although the same size as the others (about 5.6 kilobases), differed by the presence of two large substitutions of approximately 1 and 2 kilobases. The DNA sequences of the terminal direct repeats (deltas) were very homologous but not identical and were similar to previously reported Ty1 element direct repeats. We determined the 5'-flanking sequences of the ADR2 gene isolated from a wild-type strain and from five Ty1-associated mutations. The 5-base pair target sequence at the site of Ty1 insertion was present at both ends of each Ty1 element. The sites of insertion of the elements were all different and occurred from 125 to 210 base pairs in front of the coding region of ADR2. The 5' end of the major transcript as determined by S1 mapping was the same in wild-type cells and in Ty1-associated constitutive mutants and was approximately 54 base pairs upstream from the coding region. ADR2 transcripts were not detected when a solo delta sequence was present in the 5'-flanking region of this gene.

Alcohol Dehydrogenase↗
PubMed1981

Analysis of mutations affecting Ty-mediated gene expression in Saccharomyces cerevisiae.

Yeast translocatable, Ty, elements can cause constitutive synthesis of the glucose-repressible alcohol dehydrogenase (ADHII) when inserted upstream from the 5' end of the structural gene, ADR2. These insertion mutations, ADR3c, are unstable and give rise to secondary ADHII- mutations. The majority of such mutants, adr3, can be attributed to excision of the insertion sequence, leaving behind a single copy of the delta-sequence which occurs as a direct repeat at the ends of the Ty elements. A few adr3 mutants appear to be generated by DNA-rearrangements in the vicinity of the Ty insertion. The occurrence of recessive mutants, tye, which are unlinked to ADR2 indicates that the constitutive expression of ADR2 caused by the Ty insertions requires the function of trans-acting genes. These results support the idea that regulation of Ty-linked ADR2 is actively mediated by the insertion sequence and is probably not due to a mere disruption of the wild-type controlling site.

Alcohol Oxidoreductases↗
PubMed1981

Transposable elements associated with constitutive expression of yeast alcohol dehydrogenase II.

The yeast structural gene ADR2, coding for the glucose-repressible alcohol dehydrogenase (ADHII), has been isolated by complementation of function in transformed yeast. The chromosomal DNA from nine yeast strains with cis-dominant constitutive mutations (ADR3c) has been investigated by restriction enzyme analysis, using the cloned ADR2 DNA as a hybridization probe. Seven mutants appear to have insertions of approximately 5.6 kg near the 5' end of the ADR2-coding region. Four of these insertions have the same restriction pattern as the yeast transposable element Ty1. Two differ from Ty1 by the presence of an additional Hind III site, and a seventh insertion differs from Ty1 at a number of restriction sites. All are inserted in the same orientation with respect to the structural gene. A DNA fragment containing the ADR2 gene and adjacent sequences from a constitutive mutant has been cloned and shown by heteroduplex analysis to contain an insertion near the 5' end of the structural gene. The cloned insertion sequence hybridizes to multiple genomic DNA fragments, indicating that it contains a moderately repetitive sequence. Thus it appears that insertion of a transposable element near the 5' terminus of the structural gene can produce constitutive expression of a normally glucose-repressed enzyme. Such insertions seem to be the most common way of generating cis-dominant constitutive mutations of ADHII.

Alcohol Oxidoreductases↗
PubMed1979

Sigma factor is not released during transcription in Bacillus subtilis.

The relationship between sigma (sigma) and delta (delta) factors of Bacillus subtilis RNA polymerase has been analyzed during initiation of RNA synthesis. When core enzyme (E) containing delta factor (E delta) binds to DNA, the delta factor is released with the formation of an E-DNA complex. The addition of sigma to the E-DNA complex results in the formation of a stable E sigma-DNA complex which can synthesize RNA upon addition of nucleoside triphosphates. Sigma factor, significantly, is not released from the core during RNA synthesis. These results suggest that delta and sigma factors can act sequentially during initiation of RNA synthesis with delta acting as a DNA recognition factor and sigma acting as an initiation factor. The results do not preclude the possibility that E sigma can initiate RNA synthesis correctly since E sigma alone can bind to DNA and initiate RNA synthesis.

Bacillus subtilis↗
PubMed1978

Delta factor can displace sigma factor from Bacillus subtilis RNA polymerase holoenzyme and regulate its initiation activity.

A protein with a molecular weight of 21,000 daltons is found associated with a fraction of Bacillus subtilis RNA polymerase core. This protein (delta) does not react with antibody made against sigma factor and has a peptide map which is significantly different from sigma factor. At ratios of 2:1 to 4:1 (delta:holoenzyme) the delta displaces sigma factor completely from the core and associates in a 1:1 ratio with core to form delta-core. Under the same incubation conditions sigma factor at a ratio of 10:1 (sigma factor:delta-core) does not displace delta from the delta-core. The delta-core has much less activity as compared to holoenzyme on various DNA templates. However, sigma factor does stimulate the activity of delta-core enzyme under conditions of RNA synthesis. These observations and the results of others suggest that delta-core enzyme binds initially to specific DNA sites followed by delta release from the core-DNA complex and that the sigma factor binds to the core-DNA complex to initiate RNA synthesis. Thus both delta and sigma factors are required in a sequential fashion for specific transcription to occur in B subtilis.

Bacillus subtilis↗
PubMedDate not supplied

DNA sequences of two yeast promoter-up mutants.

The yeast Saccharomyces cerevisiae has three genetic loci encoding different alcohol dehydrogenase (ADH) isozymes: ADC1, which encodes the classical fermentative isozyme ADHI; ADR2, which encodes the glucose-repressed isozyme ADHII; and ADM, which encodes an ADH isozyme found associated with mitochondria. When yeast are grown on glucose, the ADC1 gene is expressed, and the ADR2 gene repressed. Conversely, growth on a non-fermentable carbon source such as ethanol or glycerol results in derepression of ADR2, and repression of ADC1. The ADC1 and ADR2 genes have been cloned and sequenced, and a number of cis-acting mutations identified that cause constitutive expression of ADR2, and seem to fall into two classes. The most abundant class consists of mutants that cannot be fully derepressed, and do not revert to wild type at a detectable level: these are caused by the insertion of a transposable element into the 5'-flanking region of the gene. The second class of mutants do revert to a glucose-repressed phenotype at a detectable frequency, and when grown on non-fermentable carbon sources derepress ADR2 to levels up to five times those found in wild-type cells. We report here the sequencing of the 5'-flanking regions of two such promoter-up, constitutive ADR2 mutants, in both of which the mutant phenotype is associated with an increase in length of a poly(A) X poly(T) tract 222 base pairs (bp) upstream of the gene.

Alcohol Oxidoreductases↗