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R A Singer

Publications and source records attributed to R A Singer.

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RNA synthesis and control of cell division in the yeast S. cerevisiae.

Cells of the yeast Saccharomyces cerevisiae rapidly accumulated in the G1 phase of the cell cycle when exposed to the chelating agents o-phenanthroline (OP) or 8-hydroxyquinoline (HQ). Zinc salts fully reversed the growth-inhibitory effect of both OP and HQ. Cells treated with these chelating agents showed limited RNA accumulation and little RNA degradation. Rates of RNA synthesis were drastically reduced by low concentrations of these compounds. Whereas rates of protein synthesis were essentially unaffected. Rates of synthesis of mRNA and tRNA were less affected than were rates of synthesis of high molecular weight RNA. Processing of ribosomal precursor RNA was altered. these results suggest that the primary effect of OP and HQ is on rRNA synthesis. RNA metabolism must therefore have a key role in the regulation of the cell cycle.

Cell Cycle↗

Methionine analogs and cell division regulation in the yeast Saccharomyces cerevisiae.

Methionine analogs such as ethionine, selenomethionine, and trifluoromethionine all arrest growth and division of the yeast Saccharomyces cerevisiae. One analog, ethionine, caused cells of the yeast to arrest specifically within G1; reciprocal shift experiments showed that ethionine and alpha-factor arrested cells at the same step ("start"). The major effect of ethionine on synthesis of macromolecules was to reduce both the rate of appearance of 35S ribosomal precursor RNA and the rate of production of mature rRNA. Synthesis of protein was relatively unaffected by ethionine. Selenomethionine and trifluoromethionine caused cells to arrest randomly in the cell division cycle. Although treatment of cells with either selenomethionine or trifluoromethionine also reduced the rate of total RNA synthesis, each of these analogs had other effects that presumably prohibited completion of the cell cycle. We propose that the rate of rRNA production is an important regulatory event in the cell cycle.

Cell Cycle↗

Growth and cell division during nitrogen starvation of the yeast Saccharomyces cerevisiae.

During nitrogen starvation, cells of the yeast Saccharomyces cerevisiae increased threefold in number, and little ribonucleic acid (RNA) and protein were accumulated. Both RNA and protein were extensivley degraded during starvation, suggesting that intracellular macromolecules could supply most of the growth requirements. The types and proportions of stable RNA synthesized during nitrogen deprivation were characteristic of exponentially growing cells; however, the complement of proteins synthesized was different. We conclude that, once events in the deoxyribonucleic acid division cycle are initiated, cells can complete division with little dependence on continued net cell growth.

Cell Division↗

Leucine biosynthesis in the blue-green bacterium Anacystis nidulans.

Leucine-requiring auxotrophs of the unicellular blue-green bacterium Anacystis nidulans have been isolated. Extracts of these mutants were deficient in alpha-isopropylmalate synthetase (EC 4.1.3.12). In wild-type cells, this enzyme was subject to feedback inhibition by leucine. However, formation of the enzymes of leucine biosynthesis was little affected by exogenous leucine in either wild-type or mutant strains. Cultures of the latter subjected to extreme leucine deprivation showed no change in specific activity of beta-isopropylmalate isomerase (EC 4.2.1.33) and at most a 50% increase in the specific activity of beta-isopropylmalate dehydrogenase (EC 1.1.1.85). These results are compared with others bearing on the evolution of the control of amino acid biosynthesis in blue-green bacteria.

2-Isopropylmalate Synthase↗

Novel ribonucleic acid species accumulated in the dark in the blue-green alga Anacystis nidulans.

In the dark, the obligately photoautotrophic blue-green alga Anacystis nidulans accumulates large relative amounts of two novel stable ribonucleic acid species (RNAs). These species are also made in illuminated cells but are unstable in them. When darkened cells are reilluminated, these RNAs are rapidly degraded; degradation is inhibited by chloramphenicol. Upon denaturation with heat or urea, one novel species (0.33 x 10(6) daltons) dissociates into two fragments that comigrate with the second novel species (0.16 x 10(6) daltons) on polyacrylamide gels. Both RNAs are associated with particles sedimenting between 30S and 50S through sucrose gradients and are removed from these particles at low magnesium concentration. The function(s) of these RNAs remains unknown.

Cell Fractionation↗

Mutational analysis of dark endogenous metabolism in the blue-green bacterium Anacystis nidulans.

We describe a mutant (strain 704) of the obligate photoautotroph Anacystis nidulans which behaves like the wild type under continuous illumination but which in the dark rapidly loses viability, respires little, and incorporates label into ribonucleic acid and protein at rates considerably less than observed with the darkened wild type. Extracts of this mutant strain show no detectable 6-phosphogluconate dehydrogenase (EC 1.1.1.44) activity. Spontaneous revertants of mutant 704 were selected as survivors of prolonged incubation in darkness. Of 10 such strains examined, none had regained 6-phosphogluconate dehydrogenase activity, and all had lost detectable glucose-6-phosphate dehydrogenase (EC 1.1.1.49) activity. Although dark survival of these revertants paralleled that of the wild type, rates of dark endogenous respiration and incorporation of labeled precursors into ribonucleic acid were still very low, comparable to those observed with strain 704. These results are consistent with the following hypotheses concerning dark endogenous metabolism in unicellular blue-green bacteria. (i) Although the oxidative pentose phosphate cycle (hexose monophosphate shunt) may play a major role in endogenous metabolism in A. nidulans, as proposed by others, it is not the only pathway capable of providing energy for maintenance of viability in darkness. (ii) Much of the endogenous metabolic activity (respiration and macromolecular synthesis) observed in darkened cultures of wild-type A. nidulans is not required for survival alone, and must therefore serve other functions.

Cell Survival↗

G1 cyclins regulate proliferation of the budding yeast Saccharomyces cerevisiae.

The eukaryotic cell cycle is regulated at two points, the G1-S and G2-M boundaries. The molecular basis for these regulatory activities has recently been elucidated, in large part by the use of molecular and genetic analyses using unicellular yeast. The molecular characterization of cell-cycle regulation has revealed striking functional conservation among evolutionarily diverse cell types. For many eukaryotic cells, regulation of cell proliferation occurs primarily in the G1 interval. The G1 regulatory step, termed START, requires the activation of a highly conserved p34 protein kinase by association with a functionally redundant family of proteins, the G1 cyclins. Here we review studies using the genetically tractable budding yeast Saccharomyces cerevisiae, which have provided insight into the role of G1 cyclins in the regulation of START.

Cell Cycle↗