Koei Okazaki

Characterization of a Fission Yeast SUMO-1 Homologue, Pmt3p, Required for Multiple Nuclear Events, Including the Control of Telomere Length and Chromosome Segregation

ABSTRACT Unlike ubiquitin, the ubiquitin-like protein modifier SUMO-1 and its budding yeast homologue Smt3p have been shown to be more important for posttranslational protein modification than for protein degradation. Here we describe the identification of the SUMO-1 homologue of fission yeast, which we show to be required for a number of nuclear events including the control of telomere length and chromosome segregation. A disruption of thepmt3 + gene, the Schizosaccharomyces pombe homologue of SMT3, was not lethal, but mutant cells carrying the disrupted gene grew more slowly. Thepmt3Δ cells showed various phenotypes such as aberrant mitosis, sensitivity to various reagents, and high-frequency loss of minichromosomes. Interestingly, we found thatpmt3 + is required for telomere length maintenance. Loss of Pmt3p function caused a striking increase in telomere length. When Pmt3p synthesis was restored, the telomeres became gradually shorter. This is the first demonstration of involvement of one of the Smt3p/SUMO-1 family proteins in telomere length maintenance. Fusion of Pmt3p to green fluorescent protein (GFP) showed that Pmt3p was predominantly localized as intense spots in the nucleus. One of the spots was shown to correspond to the spindle pole body (SPB). During prometaphase- and metaphase, the bright GFP signals at the SPB disappeared. These observations suggest that Pmt3p is required for kinetochore and/or SPB functions involved in chromosome segregation. The multiple functions of Pmt3p described here suggest that several nuclear proteins are regulated by Pmt3p conjugation.

Stress signal, mediated by a Hogl-like MAP kinase, controls sexual development in fission yeast

We identified the phhl + gene that encodes a MAP kinase as the effector of Wis1 MAP kinase kinase in fission yeast, which is highly homologous with HOG1 of S. cerevisiae. Heterothalic phh1 dsiruptant is phenotypically indistinguishable from wis1 deletion mutant, both displaying the same extent of partial sterility and enhanced sensitivity to a variety of stress. In phh1 disruptant, nitrogen starvation‐induced expression of ste11 +, a key controller of sexual differentiation, is markedly diminished. Ectopic expression of ste11 + effectively restores fertility, but not stress resistance, to the phh1 disruptant. These data show that stress signal, mediated by a MAP kinase, is required for efficient start of sexual differentiation.

Novel Factor Highly Conserved among Eukaryotes Controls Sexual Development in Fission Yeast

ABSTRACT In the fission yeast Schizosaccharomyces pombe, the onset of sexual development is controlled mainly by two external signals, nutrient starvation and mating pheromone availability. We have isolated a novel gene named rcd1+ as a key factor required for nitrogen starvation-induced sexual development.rcd1 + encodes a 283-amino-acid protein with no particular motifs. However, genes highly homologous torcd1 + (encoding amino acids with >70% identity) are present at least in budding yeasts, plants, nematodes, and humans. Cells with rcd1 + deleted are sterile if sexual development is induced by nitrogen starvation but fertile if it is induced by glucose starvation. This results largely from a defect in nitrogen starvation-invoked induction ofste11 +, a key transcriptional factor gene required for the onset of sexual development. The striking conservation of the gene throughout eukaryotes may suggest the presence of an evolutionarily conserved differentiation controlling system.

Microtubule-organizing center formation at telomeres induces meiotic telomere clustering

Telomere-localized SUN and KASH proteins induce formation of a microtubule-based “telocentrosome” that fosters microtubule motor-dependent telomere clustering.

Lub1 Participates in Ubiquitin Homeostasis and Stress Response via Maintenance of Cellular Ubiquitin Contents in Fission Yeast

ABSTRACT Ubiquitin-dependent proteolysis plays a pivotal role in stress responses. To investigate the mechanisms of these cellular processes, we have been studying Schizosaccharomyces pombe mutants that have altered sensitivities to various stress conditions. Here, we showed that Lub1, a homologue of Ufd3p/Zzz4p/Doa1p in budding yeast, is involved in the regulation of ubiquitin contents. Disruption of the lub1+ gene resulted in monoubiquitin as well as multiubiquitin depletion without change in mRNA level and in hypersensitivity to various stress conditions. Consistently, overexpression of genes encoding ubiquitin suppressed the defects associated with lub1 mutation, indicating that the phenotypes of the lub1 mutants under stress conditions were due to cellular ubiquitin shortage at the posttranscriptional level. In addition, the lub1-deleted cells showed aberrant functions in ubiquitin/proteasome-dependent proteolysis, with accelerated degradation of ubiquitin. Also Cdc48, a stress-induced chaperon-like essential ATPase, was found to interact with Lub1, and this association might contribute to the stabilization of Lub1. Our results indicated that Lub1 is responsible for ubiquitin homeostasis at the protein level through a negative regulation of ubiquitin degradation.

Dikaryotic Cell Division of the Fission Yeast Schizosaccharomyces pombe

Dikaryons, cells with two haploid nuclei contributed by the members of a mating pair, are part of the life cycle of many filamentous fungi, but the molecular mechanisms underlying the division of dikaryons are largely unknown. We found that the fission yeast Schizosaccharomyces pombe has a latent ability to divide as a dikaryon. Cells capable of restarting the mitotic cycle with two nuclei were prepared by transient inactivation of the septation initiation network. Close pairing of the two nuclei before mitosis was dependent on minus-end-directed kinesin Klp2p and was essential for propagation as a dikaryon. The two spindles extended in opposite directions, keeping their old spindle pole bodies at the prospective site of cell division until the mid-anaphase. The spindles then overlapped, exchanging the inner nuclei. Finally, twin mitosis was followed by a single cytokinesis, producing two daughter dikaryons carrying copies of the original pair of nuclei.