Concerning the map of chromosome I of Schizosaccharomyces pombe

Summary The correct orientation of a large segment (at least 200 cM) on the left arm of chromosome I of S. pombe is inverted relative to the one given in a recent mapping paper.     

A revised chromosome map of the fission yeast Schizosaccharomyces pombe

Summary The genetic map of the nuclear genome of the fission yeast Schizosaccharomyces pombe has been extended by mitotic and meiotic mapping data. A total of 158 markers are now assigned to the three linkage groups known in this organism, and 118 of them have been located on the corresponding chromosome map. Chromosome II and III each consist of one linkage group. There is some indication that the two large fragments which define chromosome I are meiotically linked, but the linkage observed is significant at the P = 0.05 level only. The length of the map is at least 1,700 map units, corresponding to an average of about 8 kilobases per map unit. The latter figure is comparable to the one obtained for intragenic recombination in the sup3 gene (Hofer et al. 1979). The basic frequency of gene conversion as measured for 21 genes varies according to a distribution of Poisson (with a modal value of 0.6% conversion per meiosis and per gene), in sharp contrast with Saccharomyces cerevisiae (Fogel et al. 1980) and Ascobolus immersus (Nicolas 1979). This may reflect the rarity of gene or region-specific rec alleles in S. pombe and may be related to the homothallism of this organism.     

Switching genes in Schizosaccharomyces pombe

Summary In homothallic (h ⁹⁰) Schizosaccharomyces pombe strains mutants occur which exhibit reduced frequencies of mating-type switching. The colonies of such mutants show a mottled iodine reaction. The underlying mutations map either in a switching signal at matl or in switching (swi) genes which are not linked to the mating-type region. Forty-nine swi mutants were examined. They map in ten different swi genes, swi1 to swi10. Seven swi genes were assigned to chromosomes I and II, respectively. Two classes of swi genes can be distinguished: when plated, class I mutants yield only mottled colonies, whereas class Il mutants yield mottled and iodine-negative colonies (most of the latter are h ¹).     

Two genes control three alkaline phosphatases in Schizosaccharomyces pombe

Summary Mutants of Schizosaccharomyces pombe defective in alkaline phosphatase activity have been selected and analyzed. The mutations map in two different loci, pho2 and pho3, both located on the right arm of chromosome II. Pho2-mutants lack the activity of a soluble alkaline phosphatase specific for nitrophenyl-phosphate. Its activity requires Mg ions and it is abolished with Zn ions. The pho3-mutations block the activities of a soluble and of a membrane-bound nonspecific alkaline phosphatase. Both pho3-controlled enzyme forms exhibit similar substrate specificities, pH-optima and Mg ions are essential for both activities. Zn ions can partially replace the Mg ions. The three enzyme forms migrate differently on nondenaturing polyacrylamide gels. We speculate that pho2 and pho3 represent the structural genes for a substrate-specific and two forms of a nonspecific alkaline phosphatase.     

Putative frameshift suppressors in Schizosaccharomyces pombe

Summary Nine genetically distinct suppressors of ICR-170-induced ade6 and ade7 mutations have been identified in Schizosaccharomyces pombe. The nine suppressors of ICR-170-induced and spontaneous origin have been assigned to the three chromosomes by haploidization and meiotic analysis. They do not suppress missense or nonsense mutations and are therefore likely to be frameshift suppressors. Based on the spectrum of suppression, the nine suppressors fall into two mutually exclusive groups. Group I comprises the two dominant suppressors sufl and suf11. Group II consists of the seven dominant suppressors suf2 through suf8. The suppressors of both groups are inefficient and all lead to a marked reduction of growth rate. Within suppressor groups, combinations of suppressors lead to drastic reductions of growth rates and to an increased efficiency of suppression. Freely segregating modifiers of suppression increasing and decreasing the efficiency of supression have been found for all the suppressors. The two omnipotent suppressors sup1 and sup2 increase the efficiency of suppression of some frameshift suppressors. The suf5 locus is unstable and reverts at very high frequency both meiotically and mitotically.     

Triploid meiosis and aneuploidy in Schizosaccharomyces pombe: an unstable aneuploid disomic for chromosome III

Summary Triploid meiosis in crosses between haploid and diploid strains of the fission yeast Schizosaccharomyces-pombe was investigated by tetrad analyses. Viability of the segregants was low. Only about 10% of the total tetrads contained four colony-forniing spores and most of these segregated into two haploids and two diploids. The remaining tetrads were rather normal in germination but were defective in colony formation. More than 50% of the total tetrads had no colony-forming spores while about 30% contained 1–3 colony-forming spores. Among the colony-forming segregants from the defective tetrads, a class of aneuploids disomic for the shortest chromosome III was obtained. In such aneuploid cells combined with a cold-sensitive ß-tubulin mutation, an additional short chromosome corresponding to chromosome III was observed by DAPI staining when the cells were incubated at a restrictive temperature. The other classes of aneuploids appeared able to be to germinate but not to enter into vegetative growth. Detailed genetical analyses indicated that recombination frequencies near the centromere were strikingly different between triploid and normal diploid meiosis.     

Switching genes in Schizosaccharomyces pombe: their influence on cell viability and recombination

Summary In Schizosaccharomyces pombe, mutations in 10 swi genes are known which reduce but not abolish mating-type (MT) switching in homothallic (h ⁹⁰) strains. Three classes (Ia, Ib, and II) of swi genes are distinguished. Three swi1 alleles are nonsense mutations. In strains with mutations in two or three swi genes, a cumulative reduction of MT switching only occurs if the genes belong to different classes. The class 1a combinations swi1 swi7 and swi3 swi7 are lethal. Besides its influence on MT switching, swi5 also reduces the frequencies of meiotic intragenic recombination and gene conversion in the ade6 locus.We dedicate this paper to Professor Kaudewitz on the occasion of his retirement     

Genetic mapping of eleven spo genes essential for ascospore formation in the fission yeast Schizosaccharomyces pombe

Summary Sporulation-deficient mutants of the fission yeast Schizosaccharomyces pombe were isolated from a homothallic strain mutagenized with ethyl methanesulfonate. Complementation tests defined two new genetic loci (spo19 and spo20) essential for ascospore formation, in addition to the 18 known spo loci (Bresch et al. 1968). A novel mapping procedure using random spore analysis prior to tetrad analysis allowed us to map 11 spo genes. Four genes (spo3, spo15, spo19 and spo20) were mapped on chromosome I, 6 genes (spo2, spo4, spoS, spo6, spo14 and spo18) on chromosome II and 1 gene (spo13) on chromosome III. Although there was no noticeable clustering of spo genes on the chromosomes, three pairs of linked genes (spo15-spo20, spo3-spo19 and spo2-spo18) were found.     

Induction control of purine catabolism in Schizosaccharomyces pombe

Summary The purine catabolic enzymes uricase, allantoinase, allantoicase and ureidoglycollase are highly induced by purines in wild-type Schizosaccharomyces pombe cells. In contrast, urease activity is constitutive. Attempts have been made to identify the inducer or inducers involved. There appears to be a sequential form of induction, uricase being induced by uric acid, allantoinase by allantoic acid (and possibly allantoin), allantoicase by allantoic acid, ureidoglycollase by ureidoglycollic acid. This sequential control of purine breakdown is compared with that found in other fungi.     

Sterile mutants of Schizosaccharomyces pombe: Analysis by somatic hybridization

Summary Sixteen sterile mutants of Schizosaccharomyces pombe, isolated from homothallic h ⁹⁰strains, were examined. It was found that they are blocked either in copulation and meiosis or in copulation alone.Protoplasts of the sterile strains were fused with protoplasts of h^+N, h^–S or h ⁹⁰strains. In these somatic crosses, eight of the sterile strains yielded hybrids which were able to form azygotic asci. Tetrad analyses revealed that seven of these sterile strains contain a single mutation; the mutations represent at least five sterility genes (ste2, ste3, ste4, ste5, ste6). One sterile strain contains two unlinked mutations which do not represent sterility genes, but genes the interaction of which results in a sterile phenotype.     

Thiamin regulates agglutination and zygote formation in Schizosaccharomyces pombe

Summary Nutritional conditions regulate mating of the fission yeast S. pombe. To investigate how nutritional signals are monitored by the cell and translated into appropriate mating behaviour, effects of unique and specific growth factors would be desirable. We show that thiamin can inhibit sexual agglutination and zygote formation in S. pombe. A concentration of 50 nM thiamin in the culture medium is required for full growth of a thiamin auxotrophic strain. At this concentration thiamin starts to inhibit mating of wild-type cells of opposite heterothallic mating type and at a 1M concentration zygote formation is inhibited by more than 95%. Growth conditions modulate the inhibitory effect of thiamin. Thiamin acts only for a restricted period of time and seems to inhibit commitment to zygote formation rather than the cell aggregation and fusion process itself. Pyrithiamin, a thiamin antagonist, inhibits growth as well as mating.     

The mitochondrial genome of Schizosaccharomyces pombe

Summary The open reading frame of the first intron of the mitochondrial cox1 gene (cox1I1) was expressed in Escherichia coli. The putative intron-encoded protein stimulated the formation of intra-chromosomal lac ⁺-recombinants about threefold. No stimulation was found when the reading frame was inserted in the opposite direction, or when it was interrupted by a deletion. The intronic open reading frame did not complement recA ^– or recB ^– mutants of E. coli. In S. pombe, elimination of this intron did not abolish homologous recombination in mitochondria. A possible role of the recombinase activity in yeast mitochondria will be discussed.     

Mapping of rRNA genes by integration of hybrid plasmids in Schizosaccharomyces pombe

Summary The major rRNA genes of the fission yeast Schizosaccharomyces pombe were mapped on chromosome III by plasmid integration. The integration vector YIp33 containing S. cerevisiae LEU2 gene was combined with the S. pombe rDNA. Since LEU2 complements S. pombe leu1 deficiency, it could be used as the genetic marker for integration. The 10.4 kb rDNA repeat contained ARS sequence, and therefore 2.4 kb and 0.7 kb subfragments not containing ARS were subcloned into YIp33 and transformed leu1 S. pombe cells to Leu⁺. Genetic analyses of the transformants indicated that the integrated rDNA resides in the long arm of the shortest chromosome III, tightly linked to ade5 (1.4 cM). This result is consistent with our previous finding that the DAPI-stained smallest chromosomes were associated with the nucleolus (Umesono et al. 1983).Abbreviations ARS autonomously replicating sequence - DAPI 4,6-diamidino-2-phenylindole - kb kilo base pairs - rDNA DNA segment containing ribosomal RNA genes - rRNA ribosomal RNA     

Mating configurations in Schizosaccharomyces pombe strains of different geographical origins

In genetic research with Schizosaccharomyces pombe the strains used are almost exclusively descendants of the clones originally isolated by Leupold. In the standard homothallic (h ⁹⁰) strain three closely linked mating-type (MT) genes are present in the MT region: the actual MT locus, mat1, and two silent cassettes, mat2 and mat3, respectively. Various rearrangements are known in the MT region, e.g., heterothallic h ⁺ or h ^- strains arise by duplications or deletions. In the present paper we analysed the mating behavior and the configurations of the MT regions of 19 S. pombe isolates from different parts of the world. In comparison with the Leupold strains several new MT configurations were found.     

The primary structure of the leul + gene of Schizosaccharomyces pombe

Summary A DNA fragment which carries the leul gene encoding beta-isopropylmalate dehydrogenase in Schizosaccharomyces pombe has been isolated by complementation of an E. coli leuB mutation. This 1.5 kb DNA fragment complements not only the S. pombe leul mutation, but also the S. cerevisiae leu2 mutation. The nucleotide sequence of the essential part of the leul gene and its flanking regions was determined. This sequence contains an open reading frame of 371 codons, from which a protein having a Mr = 39,732 can be predicted. The deduced amino acid sequence and its codon usage were compared with those of the S. cerevisiae LEU2 protein. The cloned DNA will be a useful marker when transforming S pombe.     

pH sensitivity of Schizosaccharomyces pombe: effect on the cellular phenotype associated with lacZ gene expression

We report on a series of experiments inSchizosaccharomyces pombe to detect the blue-colour colony phenotype associated with expression of theEscherichia coli lacZ gene. Increasing the pH in solid minimal medium to optimize blue colony colour revealed a pH-sensitive phenotype in auxotrophic strains requiring uracil and leucine as external supplements. This phenotype was observed among commonS. pombe stock strains, 5-fluoroorotic acid (5-FOA)-selected strains, and random genetic segregants. Growth of prototrophicS. pombe strains 972 and 975 or the adenine auxotrophic strain NCYC 1860 were unaffected by an increase in external pH. Analysis of genetic segregants from three independent crosses indicated that a single auxotrophic marker (ura4 ^- orleu1-32) was sufficient for yeast cell-growth inhibition when the medium pH was increased above 6.6. In contrast, growth of aSaccharomyces cerevisiae strain isogenic to AH22, requiring uracil, leucine and histidine, was unaffected by changes in the pH of the medium. These observations suggest that uptake of uracil and leucine intoS. pombe cells is compromised by alterations in external pH. Our results have implications for detection of thelacZ gene-encoded bluecolour colony phenotype inS. pombe, which is optimized by growth in the presence of 5-bromo-4-chloro-3-indolyl--D-galactoside (Xgal) at pH 7.0. We discuss the conditions under which this blue-colour phenotype can be routinely observed inS. pombe.     

Gene database for the fission yeast Schizosaccharomyces pombe

Summary As an aid to the fission yeast genome project, we describe a database for Schizosaccharomyces pombe consisting of both genetic and physical information. As presented, it is therefore both an updated gene list of all the nuclear genes of the fission yeast, and provides an estimate of the physical distance between two mapped genes. Additionally, a field indicates whether the sequence of the gene is available. Currently, sequence information is available for 135 of the 501 known genes.     

Some of the swi genes of Schizosaccharomyces pombe also have a function in the repair of radiation damage

Summary In Schizosaccharomyces pombe the frequency of mating-type (MT) switching is reduced by mutations in the swi genes. The ten hitherto known swi genes can be subdivided into three classes: Ia, Ib and II. Strains having swi5 (class Ib), swi9 (class II) and swi10 (class II) mutations do not only show reduced MT switching, but also exhibit an increased sensitivity to UV- and -rays. For that reason, 19 previously described rad genes were tested for their effect on MT switching. We found that swi9, rad10, rad16 and rad20 are allelic with each other indicating that the former allocation of these rad mutations to three different genes must have been erroneous. Among the remaining 16 rad genes examined, rad22 seems to be a new class II swi gene. The double mutants swi5 swi9 and swi5 swi10, but not swi9 swi10, are much more sensitive to radiation than the respective single mutants. Thus a cumulative increase in sensitivity occurs only if the mutants belong to different classes; previously the same correlation was found with regard to cumulative effects in MT switching.     

Expression and analysis of the green fluorescent protein gene in the fission yeast Schizosaccharomyces pombe

This report demonstrates that the Aequorea victoria green fluorescence protein (gfp) gene product will fluoresce in the fission yeast Schizosaccharomyces pombe when expressed from an episomal expression vector. Fluorescence was readily detectable at both the colony and single cell level. Application of fluorescence-activated cell sorting (FACS) techniques showed that gfp-expressing cells could be detected when they were as rare as 1% of a total yeast population. Quantitative analysis of gfp-expressing cells constituting as little as 5% of a total population was possible. These observations establish the suitability of the gfp gene for use in S. pombe and, in combination with FACS, offers an experimental strategy for quantitative analysis of gene expression in yeast populations.