John H. Duffus

Effects of CO2 on Budding and Fission Yeasts

SUMMARY: The budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe were grown under CO2 pressure. Cell division was inhibited in both yeasts; RNA and protein content per cell declined but DNA continued to be synthesized and, in the case of Sacch. cerevisiae, reached twice the normal level within 24 h. Changes in mean cell volume occurred in both yeasts within 1 h; an increase occurred in the budding yeast while a decrease was observed in the fission yeast. The results suggest that CO2 may exert its influence on cell characteristics through a change in cell volume.

An Investigation into the Potential Use of Chelating Agents and Antibiotics as Synchrony Inducers in the Fission Yeast Schizosaccharomyces pombe

Summary: Following the discoveries that the divalent cation ionophore A23187 and the divalent cation chelating agent EDTA can be used to synchronize yeast cell division, a study has been undertaken of the possible use of other chelating agents and antibiotics which interact with divalent cations in controlling cell division in the fission yeast Schizosaccharomyces pombe. All the agents studied (five chelators and two antibiotics) arrested cell division in growing cultures of this yeast, but only sodium pyrophosphate and citrate induced synchrony of cell division. Novobiocin produced a transient inhibition of cell division, treated cells exhibiting “endogenous recovery” in the continued presence of the antibiotic. The results obtained are discussed in relation to the hypothesis that the concentration of intracellular Mg2+ regulates cell division.

Chapter 6 The Isolation of Yeast Nuclei and Methods to Study Their Properties

Publisher Summary This chapter discusses the isolation of yeast nuclei and the techniques used to study their properties. Two main approaches have been made to the isolation of yeast nuclei. The first method is using yeast protoplasts. Recently, it has been shown possible to isolate nuclei from normal yeast cells using an Eaton press, a modified French press, or a Biotec X-Press. This has the advantage that the cells, and consequently the nuclei, are initially in a normal state; however, yields are variable and the nuclei may suffer considerable damage. In the chapter, isolation of nuclei from Sarcharomyces carlsbergensis NCYC 74 is discussed. Isolation of nuclei from Kluyveromycesfiagilis H 32 and also from Saccharomyes carlsbergensis H 60 is described. Preparation of a dense crescent (nucleolar) fraction from yeast nuclei is discussed as well.

The Use of Primuline to Identify the Septum Polysaccharide of the Fission Yeast Schizosaccharomyces Pombe

Treatment of cells and purified cell walls of the fission yeast Schizosaccharomyces pombe with primuline reveals the septum as a bright fluorescent band. When polysaccharides containing (1----3)-beta-, (1----6)-beta- or (1----3)-alpha-glucosidic linkages are treated with primuline, only those molecules containing chains of (1----3)-beta-glucosyl residues are stained. This implies that (1----3)-beta-glucan is present in the septum of Schiz. pombe as the main constituent.

Synchronization of Cell Division in the Fission Yeast Schizosaccharomyces pombe by Ethylenediaminetetra-acetic Acid

Summary: When cultures of Schizosaccharomyces pombe growing exponentially in a minimal medium were treated with 50 mM-EDTA for 1 h, nucleic acid synthesis was inhibited but protein synthesis was not. On transfer to fresh medium, 50% of the cells divided synchronously. These phenomena may be explained on the basis of reduced availability of Mg2+ following chelation with EDTA.

THE REGULATION OF CELL DIVISION IN YEAST BY MAGNESIUM

ABSTRACT Cells of the fission yeast, Schizosaccharomyces pombe, and the budding yeast, Kluyveromyces fragilis, fail to divide in Mg-free minimal media but can be induced to divide synchronously on restoration of Mg to “exhausted” cultures. Mg-depleted S. pombe, cells appear abnormally long, lack cell plates and possess elongated nuclei. This, together with evidence of a late cell cycle arrest by the ionophore A23187, suggests that specific Mg limitation blocks cells late in the G2 phase of the cell cycle. Moreover, analysis of cell Mg in cultures synchronized by various techniques reveals that there is a fairly steady fall in Mg concentration as cells grow, terminating in a rapid influx of Mg just before division. These results lead to the hypothesis that intracellular Mg concentration is the transducer for size and consequently time, related control of the cell cycle. KEYWORDS Yeast cell cycle;, magnesium and cell division control;, Schizosaccharomyces pombe/Kluyveromyces fragilis synchronous division.

Glutamine Amido Transferase in Yeast: Changes in Activity during the Cell Cycle

Glutamine aniido transferase activity is of considerable interest i n that it represents the simplest method of assaying glutamine synthetase (Shapiro & Stadtman, 1970). This is a key enzyme in the metabolic processes leading to the incorporation of nitrogen into amino acids and nucleotides in both prokaryotic and eukaryotic cells (Prusiner & Stadtman, 1973; Ferguson & Sims, I 97 I , I 9744 b; Siins & Ferguson, I 974). The present paper reports a study of changes i n glutamine amido transferase activity during the cell cycle in the budding yeast

Correlation of glutamine synthetase activity with cell magnesium concentration during cell division in yeast synchronized by induction

Summary: Synchronization of cell division in the fission yeast Schizosaccharomyces pombe and the budding yeast Kluyveromyces fragilis was achieved by induction using the DNA synthesis inhibitor 2′-deoxyadenosine and by a magnesium-exhaustion technique. The activity of glutamine synthetase in these synchronized cultures oscillated. Variations in the intracellular magnesium concentration were also observed, and peaks in magnesium concentration correlated with peaks in enzyme activity. We suggest that the enzyme from yeast is unstable and that its activity is regulated in vivo by changes in the intracellular concentration of magnesium.