A galactosyltransferase from the fission yeast Schizosaccharomyces pombe

A membrane-associated galactosyltransferase has been purified to homogeneity from the fission yeast, Schizosaccharomyces pombe. The enzyme has a molecular weight of 61,000 and is capable of transfering galactose from UDP-galactose (UDP-Gal) to a variety of mannose-based acceptors to form an alpha-1,2 galactosyl mannoside linkage. Immunofluorescence localization of the protein is consistent with the presence of the enzyme in the Golgi apparatus of S. pombe. This, together with the presence of terminal, alpha-linked galactose on the N- linked oligosaccharides of S. pombe secretory proteins, suggests that the galactosyltransferase is an enzyme involved in the processing of glycoproteins transported through the Golgi apparatus in fission yeast.     

Adenylyl cyclase activity of the fission yeast Schizosaccharomyces pombe is not regulated by guanyl nucleotides

The adenylyl cyclase activity of the fission yeast Schizosaccharomyces pombe is localized to the plasma membrane of the cell. The enzyme utilizes Mn2+/ATP as substrate and free Mn2+ ions as an effector. Unlike the baker yeast Saccharomyces cerevisiae, S. pombe adenylyl cyclase does not utilize Mg2+/ATP as substrate and the activity is not stimulated by guanyl nucleotides. The optimal pH for the S. pombe adenylyl cyclase activity is 6.0. The activity dependence on ATP is cooperative with a Hill coefficient of 1.68 +/- 0.14.     

A review of mitosis in the fission yeast Schizosaccharomyces pombe

Mitosis and cell division are the final events of the cell cycle, resulting in the precise segregation of chromosomes into two daughter cells. A highly controlled and accurate segregation of the chromosomes is required to ensure that each daughter cell receives a complete genome and remains viable. The fission yeast, Schizosaccharomyces pombe, is a unicellular eukaryotic organism which is particularly convenient for investigating these problems. It is very amenable to genetic analysis and its predominantly haploid life cycle has allowed the isolation of recessive temperature-sensitive mutants unable to complete the cell cycle. Classical genetic analysis of these mutants has been used to identify over 40 gene functions that are required for cell cycle progress in S. pombe. Many of these genes have now been cloned and sequenced and in some cases the encoded gene product has been identified. This approach, coupling classical and molecular genetics, allows identification of the molecules important in the mitotic processes and provides a means for establishing what functional roles they may play.     

Programmed cell death in fission yeast Schizosaccharomyces pombe

Yeasts have proven to be invaluable, genetically tractable systems to study various fundamental biological processes including programmed cell death. Recent advances in the elucidation of the molecular pathways underlying apoptotic cell death in yeasts have revealed remarkable similarities to mammalian apoptosis at cellular, organelle and macromolecular levels, thus making a strong case for the relevance of yeast models of regulated cell death. Programmed cell death has been reported in fission yeast Schizosaccharomyces pombe, primarily in the contexts of perturbed intracellular lipid metabolism, defective DNA replication, improper mitotic entry, chronological and replicative aging. Here we review the current understanding of the programmed cell death in fission yeast, paying particular attention to lipid-induced cell death. We discuss our recent findings that fission yeast exhibits plasticity of apoptotic and non-apoptotic modes of cell death in response to different lipid stimuli and growth conditions, and that mitochondria, reactive oxygen species and novel cell death mediators including metacaspase Pca1, SpRad9 and Pck1 are involved in the lipotoxic cell death. We also present perspectives on how various aspects of the cell and molecular biology of this organism can be explored to shed light on the governing principles underlying lipid-mediated signaling and cell demise.     

The fission yeast Schizosaccharomyces pombe in continuous culture

The fission yeast Schizosaccharomyces pombe was cultivated in a chemostat at dilution rates of D = 0.03, 0.05, 0.10, and 0.20 h(-1). After steady state had been reached, the amount of dry matter, number of cells, concentration of residual sugar, yield coefficient (Y), and some morphological properties of the cells were estimated. Curves reflecting the dry mass, number of cells, and cell mean volume show a changing coordination between the growth rate and the rate of cell division, with respect to D. In addition, it could be concluded that in dividing cells the cell septum is localized asymmetrically; Two nonidentical cells differing both in length and volume result. The degree of asymmetry is a function of the dilution rate.     

Transport of L-lysine in the fission yeast Schizosaccharomyces pombe

Systems of L-lysine transport in Schizosaccharomyces pombe are not constitutive, as at no phase of growth in a rich medium is lysine taken up. Transport activity appears only after preincubation of harvested cells with glucose or another suitable source of energy. If cycloheximide is added during this preincubation no transport systems are synthesized. After removal of glucose, the activity of the transport system decays with a half-time of 13 min. The transport of L-lysine into S. pombe cells from the stationary phase of growth preincubated for 60 min with 1% D-glucose is mediated by at least two systems, the high-affinity one with a Kt of 26 mumol/l and Jmax of 4.95 nmol/min per mg dry wt., the low-affinity one with a KT of 1.1 mmol/l and Jmax of 11.8 nmol/min per mg dry wt. The transport of lysine mediated by these two systems proceeds uphill. The high-affinity system has a pH optimum at 4.0-4.2, the accumulation ratio is highest at a cell density 2-5 mg dry wt. per ml and decreases with increasing lysine concentrations. Lysine accumulated by this system does not exit from cells. The only potent competitive inhibitors are L-arginine, L-histidine and D-lysine. The other amino acids tested do not behave as competitive inhibitors. Of the various metabolic inhibitors tested, the most potent were proton conductors and antimycin A.     

Programmed frameshifting in the synthesis of mammalian antizyme is +1 in mammals, predominantly +1 in fission yeast, but -2 in budding yeast.

The coding sequence for mammalian ornithine decarboxylase antizyme is in two different partially overlapping reading frames with no independent ribosome entry to the second ORF. Immediately before the stop codon of the first ORF, a proportion of ribosomes undergo a quadruplet translocation event to shift to the +1 reading frame of the second and main ORF. The proportion that frameshifts is dependent on the polyamine level and, because the product antizyme is a negative regulator of intracellular polyamine levels, the frameshifting acts to complete an autoregulatory circuit by sensing polyamine levels. An mRNA element just 5' of the shift site and a 3' pseudoknot are important for efficient frameshifting. Previous work has shown that a cassette with the mammalian shift site and associated signals directs efficient shifting in the budding yeast Saccharomyces cerevisiae at the same codon to the correct frame, but that the shift is -2 instead of +1. The product contains an extra amino acid corresponding to the shift site. The present work shows efficient frameshifting also occurs in the fission yeast, Schizosaccharomyces pombe. This frameshifting is 80% +1 and 20% -2. The response of S. pombe translation apparatus to the mammalian antizyme recoding signals is more similar to that of the mammalian system than to that of S. cerevisiae. S. pombe provides a good model system for genetic studies on the mechanism of at least this type of programmed mammalian frameshifting.     

Apoptosis and lipoapoptosis in the fission yeast Schizosaccharomyces pombe

Yeasts being simple eukaryotes are established genetic systems that are often employed to solve important biological questions. Recently, it has become evident that certain cell death programs exist in these unicellular organisms. For example, it has been shown recently that strains of the fission yeast Schizosaccharomyces pombe deficient in triacylglycerol synthesis undergo cell death with prominent apoptotic markers. This minireview is intended to discuss key developments that have rendered fission yeast useful both as a tool and as a model for apoptosis and lipoapoptosis research. It is attempted to delineate a putative signaling pathway leading to the execution of lipoapoptosis in the fission yeast. Although in its infancy, apoptosis research in the fission yeast promises exciting breakthroughs in the near future.     

Ammonia assimilation in the fission yeast Schizosaccharomyces pombe 972

Glutamine synthetase (GS) activity of Schizosaccharomyces pombe 972 was high in ammonia-limited cultures, low in phosphate-and sulphate-limited cultures and not detected in glucose-limited cultures. When ammonia was pulsed into an ammonia-limited culture then GS activity decreased at a rate faster than that calculated if enzyme synthesis ceased and enzyme was diluted out by growth. Enzyme activity increased in ammonia-starved, phosphate-limited cultures and in the ammonia pulse system when the added ammonia had been utilised. These increases in enzyme activity were prevented by the presence of 100 g/ml cycloheximide. GS activity was inversely related to the intracellular concentration of glutamate.Abbreviations Gs Glutamine synthetase, EC 6.3.1.2 - GOGAT Glutamine: 2-oxo-glutarate amino transferase, EC 2.6.1.53 - GDH Glutamate dehydrogenase, EC 1.4.1.3     

DNA repair in the fission yeast, Schizosaccharomyces pombe

Mutants of the fission yeast Schizosaccharomyces pombe which are sensitive to UV and/or γ-irradiation have been assigned to 23 complementation groups, which can be assigned to three phenotypic groups. We have cloned genes which correct the deficiency in mutants corresponding to 12 of the complementation groups. Three genes in the excision-repair pathway have a high degree of sequence conservation with excision-repair genes from the evolutionarily distant budding yeast Saccharomyces cerevisiae. In contrast, those genes in the recombination repair pathway which have been characterised so far, show little homology with any previously characterised genes.     

Versatile use of Schizosaccharomyces pombe plasmids in Saccharomyces cerevisiae

The two model yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe appear to have diverged 1000 million years ago. Here, we describe that S.?pombe vectors can be propagated efficiently in S.?cerevisiae as pUR19 derivatives, and the pREP and pJR vector series carrying the S.?cerevisiae LEU2 or the S.?pombe ura4(+) selection marker are maintained in S.?cerevisiae cells. In addition, genes transcribed from the S.?pombe nmt1(+) promoter and derivatives are expressed in budding yeast. Thus, S.?pombe vectors can be used as shuttle vectors in S.?cerevisiae and S.?pombe. Our finding greatly facilitates the testing for functional orthologs of protein families and simplifies the cloning of new S.?pombe plasmids by using the highly efficient in vivo homologous recombination activity of S.?cerevisiae.