• 한국동물학회:학술대회논문집
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• 한국동물학회 1999년도 한국생물과학협회 학술발표대회
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• pp.251.2-251
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• 1999

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• Genomics & Informatics
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• 제19권4호
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• pp.39.1-39.8
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• 2021

Tamoxifen (TAM) is an anticancer drug used to treat estrogen receptor (ER)-positive breast cancer. However, its ER-independent cytotoxic and antifungal activities have prompted debates on its mechanism of action. To achieve a better understanding of the ER-independent antifungal action mechanisms of TAM, we systematically identified TAM-sensitive genes through microarray screening of the heterozygous gene deletion library in fission yeast (Schizosaccharomyces pombe). Secondary confirmation was followed by a spotting assay, finally yielding 13 TAM-sensitive genes under the drug-induced haploinsufficient condition. For these 13 TAM-sensitive genes, we conducted a comparative analysis of their Gene Ontology (GO) 'biological process' terms identified from other genome-wide screenings of the budding yeast deletion library and the MCF7 breast cancer cell line. Several TAM-sensitive genes overlapped between the yeast strains and MCF7 in GO terms including 'cell cycle' (cdc2, rik1, pas1, and leo1), 'signaling' (sck2, oga1, and cki3), and 'vesicle-mediated transport' (SPCC126.08c, vps54, sec72, and tvp15), suggesting their roles in the ER-independent cytotoxic effects of TAM. We recently reported that the cki3 gene with the 'signaling' GO term was related to the ER-independent antifungal action mechanisms of TAM in yeast. In this study, we report that haploinsufficiency of the essential vps54 gene, which encodes the GARP complex subunit, significantly aggravated TAM sensitivity and led to an enlarged vesicle structure in comparison with the SP286 control strain. These results strongly suggest that the vesicle-mediated transport process might be another action mechanism of the ER-independent antifungal or cytotoxic effects of TAM.

• Journal of Microbiology and Biotechnology
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• 제15권4호
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• pp.694-700
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• 2005

Defects in the mitotic spindle or in the attachment of chromosomes to the spindle are believed to release an activated form of spindle checkpoint complex that inhibits APC-dependent ubiquitination and subsequently arrests the cell cycle at metaphase. When the spindle assembly is disrupted, the fission yeast mitotic arrest deficient (mad) mutants fail to arrest and rapidly lose viability. To enhance our understanding of the molecular mechanisms for the pathway of checkpoint function, the functional characterizations of Mad 1 p from Schizosaccharomyces pombe involved in this process have been carried out. Yeast two-hybrid and various deletion analyses of S. pombe Mad1 p reveal that the C terminus of Mad1p is critical for the binding of Mad2p and maintenance of Mad 1 p-Mad2p interaction. In addition, it was found. that the Mad1p region (residues 206-356) is essential for Mad1p-other checkpoint components. Mad1p truncating this region is sufficient to bind Mad2p but abolishes the checkpoint function, indicating that the checkpoint function is necessary for interaction of Mad 1 p-other checkpoint components. The possible functions of S. pombe Mad1p at the cell cycle checkpoint are discussed.

• BMB Reports
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• 제32권1호
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• pp.92-97
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• 1999

Cytokinesis and septation should be coordinated to nuclear division in the cell division cycle for precise transmission of the genome into daughter cells. byr4, an essential gene in fission yeast Schizosaccharomyces pombe, regulates the timing of cytokinesis and septation in a dosage-dependent manner. We examined the intracellular localization of the Byr4 protein by expressing byr4 as a fusion of green fluorescence protein (GFP). The Byr4 protein localizes as a single dot on the nuclear periphery of interphase cells, duplicates before mitosis, and the duplicated dots segregate with the nuclei in anaphase. The behavior of Byr4p throughout the cell cycle strongly suggests that Byr4p is localized to the spindle pole body (SPB), a microtubule organizing center (MTOC) in yeast. The presence of the Byr4 protein in the SPB is consistent with its function to coordinate mitosis and cytokinesis. We also mapped the domains of Byr4p for its proper localization to SPB by expressing various byr4 deletion mutants as GFP fusions. Analyses of the diverse byr4 deletion mutants suggest that the indirect repeats and the regions homologous to the open reading frame (ORF) YJR053W of S. cerevisiae in its C-terminus are essential for its localization to the SPB.

• Molecules and Cells
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• 제20권1호
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• pp.74-82
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• 2005

Glutaredoxins (Grxs) are thioloxidoreductases which are required for maintaining thiol/disulfide equilibrium in living cells. The Grx3 gene, which encodes one of the three monothiol Grxs in the fission yeast Schizosaccharomyces pombe, was characterized, and its transcriptional regulation studied. Genomic DNA encoding Grx3 was isolated by PCR, and a plasmid pTT3 carrying this DNA was produced. The DNA sequence has 1,267 bp, which would encode a monothiol Grx of 166 amino acids with a molecular mass of 18.3 kDa. The putative protein has 27% homology with Grx5, and contains many hydrophobic amino acid residues in its N-terminal region. S. pombe cells harboring pTT3 had increased Grx activity and enhanced survival on minimal medium plates containing aluminum (5 mM), BSO (0.05 mM), menadione (0.01 mM) or cadmium (0.2 mM). The 568 bp upstream region of Grx3 was fused into the promoterless b-galactosidase gene of the shuttle vector YEp367R to generate fusion plasmid pMJS10. Potassium chloride (KCl) and metals including aluminum and cadmium enhanced the synthesis of ${\beta}$-galactosidase from the fusion gene. The synthesis of ${\beta}$-galactosidase was also enhanced, in a Pap1-dependent manner, by fermentable carbon sources such as glucose (at low concentrations) and sucrose, but not by non-fermentable carbon sources such as ethanol and acetate. Grx3 mRNA increased in response to treatment with BSO. These observations indicate that S. pombe Grx3 is involved in the response to stress, and is regulated by stress.

• BMB Reports
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• 제36권3호
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• pp.326-331
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• 2003

The fission yeast cells that contained the cloned glutathione synthetase (GS) gene showed 1.4-fold higher glutathione (GSB) content and 1.9-fold higher GS activity than the cells without the cloned GS gene. Interestingly, $\gamma$-glutamylcysteine synthetase activity increased 2.1-fold in the S. pombe cells that contained the cloned GS gene. The S. pombe cells that harbored the multi copy-number plasmid pRGS49 (containing the cloned GS gene) showed a higher level of survival on solid media with cadmium chloride (1 mM) or mercuric chloride ($10\;{\mu}M$) than the cells that harbored the YEp357R vector. The 506 bp upstream sequence from the translational initiation point and N-terminal8 amino acid-coding region were fused into the promoteriess $\beta$-galactosidase gene of the shuttle vector YEp367R to generate the fusion plasmid pUGS39. Synthesis of $\beta$-galactosidase from the fusion plasmid pUGS39 was significantly enhanced by cadmium chloride and NO-generating S-nitroso-N-acetylpenicillamine (SNAP) and sodium nitroprusside (SN). It was also induced by L-buthionine-(S,R)-sulfoximine, a specific inhibitor of $\gamma$-glutamylcysteine synthetase (GCS). We also found that the expression of the S. pombe GS gene is regulated by the Atf1-Spc1-Wis1 signal pathway.

• 미생물학회지
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• 제51권4호
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• pp.435-439
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• 2015

THO/TREX 복합체는 전사 신장, mRNA 가공 및 방출, 그리고 유전체 안정성에 중요한 역할을 담당한다. 분열효모 Schizosaccharomyces pombe에서 THO/TREX 복합체의 한 구성요소인 THOC5의 이종상동체를 암호화하고 있는 SPBC 577.04 유전자를 찾아, 그것의 기능을 분석하였다. S. pombe thoc5 (spthoc5) 유전자는 생장과 mRNA의 방출에 필수적이지는 않지만, 결실돌연변이는 야생형에 비해 생장 결함을 보였고 $poly(A)^+$ RNA도 핵 안에 약간 축적되는 현상을 보였다. 또한 정상적인 기능을 가진 spThoc5-GFP 단백질은 주로 핵안에 존재하였다. Co-immunoprecipitation 분석에서 진화적으로 잘 보존된THO/TREX 복합체의 주요 구성인자인 Hpr1(THOC1)는 또 다른 구성인자인 Tho2 (THOC2) 뿐만 아니라 spThoc5와도 상호작용을 하였다. 이와 같은 결과들은 S. pombe의 Thoc5 상동체도 THO/TREX 복합체의 구성인자로 mRNA 방출에 관여하고 있음을 시사한다.

• BMB Reports
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• 제38권5호
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• pp.609-618
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• 2005

$\gamma$-Glutamyl transpeptidase (GGT, EC 2.3.2.2.) catalyzes the transfer of the $\gamma$-glutamyl moiety from $\gamma$-glutamyl containing ompounds, notably glutathione (GSH), to acceptor amino acids and peptides. A second gene (GGTII) encoding GGT was previously isolated and characterized from the fission yeast Schizosaccharomyces pombe. In the present work, the GGTII-lacZ fusion gene was constructed and used to study the transcriptional regulation of the S. pombe GGTII gene. The synthesis of $\beta$-galactosidase from the GGTII-lacZ fusion gene was significantly enhanced by NO-generating SNP and hydrogen peroxide in the wild type yeast cells. The GGTII mRNA level was increased in the wild-type S. pombe cells treated with SNP. However, the induction by SNP was abolished in the Pap1-negative S. pombe cells, implying that the induction by SNP of GGTII is mediated by Pap1. Fermentable carbon sources, such as glucose (at low concentrations), lactose and sucrose, as a sole carbon source, enhanced the synthesis of $\beta$-galactosidase from the GGTII-lacZ fusion gene in wild type KP1 cells but not in Pap1-negative cells. Glycerol, a non-fermentable carbon source, was also able to induce the synthesis of $\beta$-galactosidase from the fusion gene, but other non-fermentable carbon sources such as acetate and ethanol were not. Transcriptional induction of the GGTII gene by fermentable carbon sources was also confirmed by increased GGTII mRNA levels in the yeast cells grown with them. Nitrogen starvation was also able to induce the synthesis of $\beta$-galactosidase from the GGTII-lacZ fusion gene in a Pap1-dependent manner. On the basis of the results, it is concluded that the S. pombe GGTII gene is regulated by oxidative and metabolic stress.

• KSBB Journal
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• 제6권2호
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• pp.189-194
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• 1991

The effect of wtlitc light on unaeraten growth of Baker's yeast and the accompanying ethanol production has been studied in a batch process at 27$^{\circ}C$. Over the 80-hour period of the Champagne yeast process without pH control, the cull growth was inhibited by the fluorescent light. Another observed difference between the runs is that the drop and subsequent rise in redox potential occurred much sooner in the fermentation with light than in the fermentation without light. This preliminary study indicated that ethanol production could be enhanced by light as the cell concentration is repressed. The possible pathway, shift of the sugar substrate toward ethanol and away from cells was manifested by another difference as well. As observed under the microscope, many of the yeast cells grown under light budded without dividing by the normal fission process as they did in the dark. Furthermore, the undivided and branched (light grown) cell did not agglutinate at the end of the fermentation process as did the distinct spherical (dark grown) cells.

• Molecules and Cells
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• 제24권3호
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• pp.316-322
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• 2007

Glutaredoxins (Grxs), also known as thioltransferases (TTases), are thiol oxidoreductases that regulate cellular redox state in a variety of organisms. In the budding yeast Saccharomyces cerevisiae, Grx1 and 2 are cytosolic dithiol Grxs, while Grx3, 4 and 5 are monothiol Grxs. A gene encoding a new monothiol Grx, Grx6, was cloned from the genomic DNA of S. cerevisiae by PCR. Its DNA sequence contains 1,080 bp, and encodes a putative protein of 203 amino acid residues containing Cys-Phe-Tyr-Ser at the active site. Grx6 is similar to other monothiol Grxs in the same organism and to Grx3 in the fission yeast Schizosaccharomyces pombe. and its predicted three-dimensional structure resembles that of S. pombe Grx3. S. pombe cells harboring plasmid pFGRX6 containing the Grx6 gene had about 1.3-fold elevated Grx activity in the exponential phase, and grew better than the control cells under some stressful conditions. Synthesis of ${\beta}$-galactosidase from a Grx6-lacZ fusion gene in S. pombe was enhanced by potassium chloride, aluminum chloride and heat ($37^{\circ}C$) treatment. S. pombe cells harboring plasmid pFGRX6 had elevated ROS levels whereas S. pombe cells harboring extra copies of Grx3 had reduced ROS levels.