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W L Fangman

Publications and source records attributed to W L Fangman.

70 records · Page 4Linked to original sources
PubMed1975

Mitochondrial DNA synthesis in cell cycle mutants of Saccharomyces cerevisiae.

Mitochondrial DNA replication was examined in mutants for seven different Saccharomyces cerevisiae genes which are essential for nuclear DNA replication. In cdc8 and cdc21, mutants defective in continued replication during the S phase of the cell cycle, mitochondrial DNA replication ceases at the nonpermissive temperature. Replication is temperature sensitive even when these mutants are arrested in the G1 phase of the cell cycle with alpha factor, a condition where mitochondrial DNA replication continues for the equivalent of several generations at the permissive temperature. Therefore the cessation of replication results from a defect in mitochondrial replication per se, rather than from an indirect consequence of cells being blocked in a phase of the cell cycle where mitochondrial DNA is not normally synthesized. Since the temperature-sensitive mutations are recessive, the products of genes cdc8 and cdc21 must be required for both nuclear and mitochondrial DNA replication. In contrast to cdc8 and cdc21, mitochondrial DNA replication continues for a long time at the nonpermissive temperature in five other cell division cycle mutants in which nuclear DNA synthesis ceases within one cell cycle: cdc4, cdc7, and cdc28, which are defective in the initiation of nuclear DNA synthesis, and cdc14 and cdc23, which are defective in nuclear division. The products of these genes, therefore, are apparently not required for the initiation of mitochondrial DNA replication.

Cell Division↗
PubMed1975

Temporal order in yeast chromosome replication.

Previous work with bacteria has shown that a gene is maximally sensitive to mutagenesis by N-methyl-N'-nitro-N-nitrosoguanidine (NG) at the time it is being replicated. NG was used to test for temporal order in the replication of the genome of the unicellular eucaryote, Saccaromyces cerevisiae. Yeast cells growing exponentially were more sensitive to mutagenesis by NG than cells in which DNA synthesis had been inhibited. Further, in a synchronized population of cells, individual genetic markers exhibited maximum sensitivity to muta-genesis at distinct limited intervals within the DNA synthesis period. The peaks of sensitivity are interpreted as reflecting the times of replication of different genes. Since markers for five genes on four different chromosomes showed discrete periods of maximum sensitivity, it is likely that temporal ordering of replication exists for most genes in the yeast genome. These results imply that sites for initiation of DNA replication occur at fairly specific regions along yeast chromosomal DNA moleucles, and are activated at predetermined times in the DNA synthesis period.

Cell Division↗
PubMed1973

Size and structure of yeast chromosomal DNA.

Electron microscopic analysis indicates that yeast nuclear DNA can be isolated as linear molecules ranging in size from 50 mum (1.2 x 10(8) daltons) to 355 mum (8.4 x 10(8) daltons). Analysis indicates the data is consistent with the hypothesis that each yeast chromosome contains a single, linear DNA duplex. Mitochondrial DNA molecules have a contour length of 21 +/- 2 mum and are mostly linear.

Carbon Radioisotopes↗
PubMed1973

Single-strand breaks in deoxyribonucleic acid and viability loss during deoxyribonucleic acid synthesis inhibition in Escherichia coli.

The effects of deoxyribonucleic acid (DNA) synthesis inhibition brought about in four different ways-thymidine starvation, nalidixic acid, hydroxyurea, and dnaB mutation-were examined in isogenic strains of Escherichia coli K-12. Three parameters were examined to determine whether there are strict correlations among them: (i) the extent of DNA synthesis inhibition; (ii) cell survival; and (iii) the rate of breakage of DNA molecules. There was no significant correlation between the extent of DNA synthesis inhibition and the rate of viability loss caused by the four DNA synthesis inhibitors, nor was there a strict correlation between the rate of occurrence of single-strand breaks in DNA and loss of viability. During treatment with hydroxyurea (0.1 M), no viability loss was observed and little, if any, single-strand breakage of DNA occurred. Both thymidine starvation and nalidixic-acid (20 mug/ml) treatment resulted in viability loss and breakage of DNA. For these latter two inhibitors, the two events appeared to be associated because greater rates of both viability loss and DNA breakage were observed for nalidixic acid compared with thymidine starvation. However, viability loss need not be associated with extensive breakage of DNA as demonstrated with a temperature-sensitive DNA synthesis mutant; at 39 C, viability loss occurred at a high rate without significant DNA breakage. With the other agents, the amount of DNA breakage accumulated when a cell population has sustained an average of one lethal hit was estimated to be about 30 single-strand breaks per genome. Differences in chromosomal and episomal breakage rates were observed.

Adenine↗
PubMed1972

Sedimentation properties of yeast chromosomal DNA.

Sedimentation analysis of nuclear DNA released from spheroplasts of the yeast Saccharomyces cerevisiae indicates that it has a number average molecular weight of 6.2 x 10(8). The chromosomal DNA molecules range in size from as small as 5 x 10(7) daltons to as large as 1.4 x 10(9) daltons. Based on these values and estimates of the total DNA content of the yeast nucleus, it is proposed that each yeast chromosome contains a single DNA duplex.

Carbon Isotopes↗
PubMed1969

Specificity and efficiency of thymidine incorporation in Escherichia coli lacking thymidine phosphorylase.

A mutant of Escherichia coli lacking the catabolic enzyme thymidine phosphorylase readily incorporates exogenous thymidine into deoxyribonucleic acid (DNA) even when provided at concentrations as low as 0.2 mug/ml. Incorporation by this prototrophic strain occurs specifically into DNA, since, with radioactively labeled thymidine, (i) more than 98% is incorporated into alkali-stable material, (ii) at least 90% is recovered as thymine after brief formic acid hydrolysis, and (iii) at least 90% is incorporated into material with the buoyant density of DNA. During growth in medium containing thymidine, the bacteria obtain approximately half of their DNA thymines from the exogenous thymidine and half from endogenous synthesis. The thymines and cytosines of DNA can be simultaneously and specifically labeled by thymidine-2-(14)C and uridine-5-(3)H, respectively. The mutant, which does not degrade thymidine, retains the ability to degrade the thymidine analogue 5-bromodeoxyuridine.

Adenine↗
PubMedDate not supplied

Creation of ARS activity in yeast through iteration of non-functional sequences.

Replication origins in Saccharomyces cerevisiae have been identified through the cloning of autonomous replication sequence (ARS) elements that allow the extrachromosomal maintenance of plasmid molecules. ARS activity requires a close match to an 11 bp consensus sequence and A + T-rich flanking DNA. ARS elements with a wide range of capacities for promoting plasmid maintenance have been described. We determined the ARS activity of plasmids with inserts consisting of repetitions of a 64 bp 100% A + T sequence that has sequence similarities to known ARS elements. An insert with approximately four repeats did not yield transformants, but inserts with either eight or eleven repeats did. The cooperative production of ARS activity did not require a contiguous arrangement since a plasmid containing two inserts of four repeats each, separated by about 1 kb, was functional. Our results show that a change from non-function to function can be accomplished by the cumulative action of individually inactive sequences. We conclude that the probability of replication initiation is too low with only four repeats to allow plasmid maintenance, but the overall probability is increased by further sequence iteration to provide origin activity. We suggest that chromosomes may contain stretches with dispersed, weak origin elements, each undetected by the conventional ARS assay, that in sum provide origin function.

DNA Replication↗