Cell Division in Yeasts: Movement of Organelles Associated with Cell Plate Growth of Schizosaccharomyces pombe

Electron microscopy of dividing fission yeast cells shows establishment of an annular rudiment (AR) of electron-transparent material under the old cell wall as the first sign of elaboration of the cell plate. The AR grows centripetally, finally closing at the mid-point of the cell. During the inward growth of the AR it is thickened by addition of denser material which becomes the scar plug after fission; the electron-transparent material is lost at fission. Lying always between the cytoplasmic membrane and the cell wall is a dark layer of variable thickness. This layer becomes markedly thickened into a fillet at the base of the centripetally growing cell plate. The fission process begins after the cell plate is completely elaborated. One striking feature of fission is the migration of dense material from the fillet at the base of the cell plate outwardly through the matrix of the cell wall to its final resting place as a dark ring, a "fuscannel," adjacent to the fission scar. The inclusion of Golgi bodies in many sections suggests their involvement in cell plate elaboration, presumably through production of the dense bodies which are seen to fuse with the dark layer proximal to the growing cell plate.     

Genetic Regulation of Cell Division Initiation in Bacillus subtilis

The growth and division properties of a temperature-sensitive mutant of Bacillus subtilis defective in the initiation of cell division have been studied. Log-phase cells transferred from 30 to 45 C continue to increase in length but fail to initiate new divisions. Deoxyribonucleic acid synthesis continues at 45 C, and genomes are segregated along the filament length. When filaments are returned to 30 C, division initiation resumes, and the long multinucleate clones are partitioned into normal-size cells. Occasionally, multiple cross walls initiate in close proximity, resulting in tiny cells, some of which are anucleate. Division resumption is sensitive to protein synthesis inhibitors, suggesting there is a new protein required for the initiation of division in filaments.     

Cell cycle regulation in Schizosaccharomyces pombe

Cdc2, a cyclin-dependent kinase, controls cell cycle progression in fission yeast. New details of Cdc2 regulation and function have been uncovered in recent studies. These studies involve cyclins that associate with Cdc2 in G1-phase and the proteins that regulate inhibitory phosphorylation of Cdc2 during S-phase and G2-phase. Recent investigations have also provided a better understanding of proteins that regulate DNA replication and that are directly or indirectly controlled by Cdc2.     

Mutations induced by x-radiation in the yeast Schizosaccharomyces pombe

Experiments in strains of yeast with different genetic backgrounds were done to evaluate the kinetics of inactivation and mutation induction by X-radiations. A system of forward mutation induction in five loci was used and a specific mutation rate of 0.14·10⁻⁸×locus×rad was evaluated for the wild type.From a comparison of observations with wild type and radiation-sensitive strains, it may be assumed that, in this yeast, mutations are mainly the result of a repair-active process.The range of genotypic and phenotypic influence upon the specific locus mutation rate was evaluated with appropriate biological material and experiments.     

Genetic effects of potassium dichromate in Schizosaccharomyces pombe.

In this review we outline the various factors which may contribute to the non-randomness of intragenic mutational spectra and the occurrence of hot spots. These factors include sample size limitation, particularly for sites of low mutability, and possible regions of low recombination potential. In addition, the nature of the gene product places great restraint on the detectability of either frameshift and premature chain-terminating mutations on one hand, or of the majority of missense mutations on the other. The nature of the Genetic Code itself also limits the mutational spectrum in so far as specific base pair substitutions lead only to a limited number of detectable amino acid replacements.Mutational hot spots may be a special example of the influence of neighbouring base pairs in the mutability of any given base pair. This is apparently true for frameshift mutations which tend to occur in runs of repeated base pairs or base pair doublets. Neigbouring base effects could operate not only at the level of initial reactivity with a mutagen, but also subsequently at the levels of DNA repair, recombination or replication. In some cases rare or modified bases may be responsible for neighbour effects. We suggest specific experimental approaches which seem likely to aid in the elucidation of these problems.     

Genetic studies on cycloheximide-resistant strains of Schizosaccharomyces pombe.

Cycloheximide-resistant mutants of Schizosaccharomyces pombe were isolated either as spontaneous mutants or after mutagenic treatment with nitrous acid, UV and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Twenty-three spontaneous mutants and 64 induced mutants were analysed genetically. Crosses revealed that at least four loci, designated cyh1, cyh2, cyh3 and cyh4 are responsible for resistance. Alleles of cyh1 show good growth on either high (100 mug/ml) or low (40 mug/ml) concentrations of cycloheximide whereas alleles at the cyh2, cyh3 and cyh4 loci gorw well on 40 mug/ml but poorly on 100 mug/ml. Some alleles at the cyh2 and cyh3 loci are also temperature sensitive (ts), the ts phenotype being conferred by the same gene as the resistance. In diploids, cyh1 and cyh4 are re-essive to wild type whereas cyh2 and cyh3 are semi-dominant. There was no intragenic complementation between three cyh1 alleles. Cross-resistance to trichodermin and anisomycin was shown by cyh2, cyh3 and cyh4 but not cyh1. Most cyh1 alleles, of spontaneous and UV origin only, were cold sensitive (cs) at 14 degrees and some of these were also cycloheximide dependent at the same temperature. It is suggested that the cyh1 and cyh4 genes are involved in ribosome formation or function and the other loci probably affect the uptake of cycloheximide by the cells.     

Dual Functions for the Schizosaccharomyces pombe Inositol Kinase Ipk1 in Nuclear mRNA Export and Polarized Cell Growth

The inositol 1,3,4,5,6-pentakisphosphate (IP₅) 2-kinase (Ipk1) catalyzes the production of inositol hexakisphosphate (IP₆) in eukaryotic cells. Previous studies have shown that IP₆ is required for efficient nuclear mRNA export in the budding yeast Saccharomyces cerevisiae. Here, we report the first functional analysis of ipk1⁺ in Schizosaccharomyces pombe. S. pombe Ipk1 (SpIpk1) is unique among Ipk1 orthologues in that it harbors a novel amino (N)-terminal domain with coiled-coil structural motifs similar to those of BAR (Bin-amphiphysin-Rvs) domain proteins. Mutants with ipk1⁺ deleted (ipk1Δ) had mRNA export defects as well as pleiotropic defects in polarized growth, cell morphology, endocytosis, and cell separation. The SpIpk1 catalytic carboxy-terminal domain was required to rescue these defects, and the mRNA export block was genetically linked to SpDbp5 function and, likely, IP₆ production. However, the overexpression of the N-terminal domain alone also inhibited these functions in wild-type cells. This revealed a distinct noncatalytic function for the N-terminal domain. To test for connections with other inositol polyphosphates, we also analyzed whether the loss of asp1⁺ function, encoding an IP₆ kinase downstream of Ipk1, had an effect on ipk1Δ cells. The asp1Δ mutant alone did not block mRNA export, and its cell morphology, polarized growth, and endocytosis defects were less severe than those of ipk1Δ cells. Moreover, ipk1Δ asp1Δ double mutants had altered inositol polyphosphate levels distinct from those of the ipk1Δ mutant. This suggested novel roles for asp1⁺ upstream of ipk1⁺. We propose that IP₆ production is a key signaling linchpin for regulating multiple essential cellular processes.     

Two-step fermentation of acidic grape musts: Interactions between Schizosaccharomyces pombe and Saccharomyces spp.

The interactions between Schizosaccharomyces pombe and Saccharomyces spp. (S. cerevisiae, S. cerevisiae sake, S. bayanus, S. uvarum) were investigated by growing the yeasts in sterile, partially fermented glucose asparagine medium in flasks, and also in the Ecologen containing either synthetic medium or grape must be separating the adjacent chambers with membranes which allow free movement of medium but not of cells. The growth of Sch. pombe was inhibited by Saccharomyces spp. to a varied extent, but the reverse was not observed. Saccharomyces uvarum, and S. cerevisiae more strongly inhibited Sch. pombe than the other species tested. All three strains of Sch. pombe (ICV-M, BG, ATCC-16979) were inhibited by S. cerevisiae although ICV-M and ATCC strains were more sensitive than BG. The higher growth rate of S. cerevisiae resulted in the exhaustion of nutrients, and its metabolic products could possibly be responsible for the growth inhibition of Sch. pombe. In the light of the present experimental results, the suitability of a two-step fermentation process for making better quality wines from acidic grape musts is discussed.     

Nuclear and Cell Division in Filamentous Bacteria

GROWTH in unicellular bacteria usually takes place as a sequence of events culminating in cell division^1–6. In some cases, however, bacteria grows in chains (referred to here as filaments). Two reasons have been put forward for this: either the individual segments of a filament are complete cells in which the timing of separation from adjacent segments is an imprecise event, not coordinated with overall growth, or the production of filaments may be a controlled process which is a prerequisite to cell division. There is some support for the first possibility⁷, but more recently it has been observed that the chain length of filamentous bacteria depends on the growth medium^8,9. Here we report further evidence that in the bacterium Bacillus subtilis the production of filaments of up to sixteen nuclei is part of a programme of events necessary for the maintenance of particular rates of cell division and that the structure and physiology of this bacterium can be explained along the same lines as those of the models describing growth in the unicellular intestinal bacteria Escherichia coli and Salmonella typhimurium.     

Cell Surface Galactosylation Is Essential for Nonsexual Flocculation in Schizosaccharomyces pombe

We have isolated fission yeast mutants that constitutively flocculate upon growth in liquid media. One of these mutants, the gsf1 mutant, was found to cause dominant, nonsexual, and calcium-dependent aggregation of cells into flocs. Its flocculation was inhibited by the addition of galactose but was not affected by the addition of mannose or glucose, unlike Saccharomyces cerevisiae FLO mutants. The gsf1 mutant coflocculated with Schizosaccharomyces pombe wild-type cells, while no coflocculation was found with galactose-deficient (gms1Δ) cells. Moreover, flocculation of the gsf1 mutant was also inhibited by addition of cell wall galactomannan from wild-type cells but not from gms1Δ cells. These results suggested that galactose residues in the cell wall glycoproteins may be receptors of gsf1-mediated flocculation, and therefore cell surface galactosylation is required for nonsexual flocculation in S. pombe.     

Genetic and biochemical analysis of the adenylyl cyclase of Schizosaccharomyces pombe.

The adenylyl cyclase gene, cyr1, of Schizosaccharomyces pombe has been cloned. We have begun an analysis of the function and regulation of adenylyl cyclase by disrupting this gene and by over-expressing all or parts of this gene in various strains. cyr1- strains are viable and contain no measurable cyclic AMP. They conjugate and sporulate under conditions that normally inhibit wild-type strains. Strains containing the cyr1 coding sequences transcribed from the strong adh1 promoter contain greatly elevated adenylyl cyclase activity, as measured in vitro, but only modestly elevated cAMP levels. Such strains conjugate and sporulate less frequently than wild-type cells upon nutrient limitation. Strains which carry the wild-type cyr1 gene but that also express high levels of the amino terminal domain of adenylyl cyclase behave much like cyr1-strains, suggesting that the amino terminal domain can bind a positive regulator. A protein that copurifies with the adenylyl cyclase of S. pombe cross-reacts to antiserum raised against the S. cerevisiae adenylyl cyclase-associated regulatory protein, CAP.