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https://www.readbyqxmd.com/read/28922845/pot1-mediated-%C3%AE-integration-strategy-for-high-copy-stable-expression-of-heterologous-proteins-in-saccharomyces-cerevisiae
#1
Xiaofei Song, Quanli Liu, Jiwei Mao, Yuzhen Wu, Yuanzi Li, Kai Gao, Xiuming Zhang, Yanling Bai, Haijin Xu, Mingqiang Qiao
In biotechnological industry, increased expression cassette stability and copy number serve as important means of maintaining consistently high production levels of heterologous proteins in Saccharomyces cerevisiae. In this study, we combined δ sequences for site-specific integration with TPI1 gene from Schizosaccharomyces pombe (POT1) as a selection marker to realize high-copy integration and stable expression of heterologous proteins in S. cerevisiae. With the newly developed POT1 platform, a 32-copy integration of enhanced green fluorescent protein (EGFP) expression cassette was obtained in a single round and was stably maintained after 100 generations of growth in a rich complex medium...
September 1, 2017: FEMS Yeast Research
https://www.readbyqxmd.com/read/28917635/evolutionary-analysis-of-nucleosome-positioning-sequences-based-on-new-symmetric-relative-entropy
#2
Hu Meng, Hong Li, Yan Zheng, Zhenhua Yang, Yun Jia, Suling Bo
New Symmetric Relative Entropy (NSRE) was applied innovatively to analyze the nucleosome sequences in S. cerevisiae, S. pombe and Drosophila. NSRE distributions could well reflect the characteristic differences of nucleosome sequences among three organisms, and the differences indicate a concerted evolution in the sequence usage of nucleosome. Further analysis about the nucleosomes around TSS shows that the constitutive property of +1/-1 nucleosomes in S. cerevisiae is different from that in S. pombe and Drosophila, which indicates that S...
September 13, 2017: Genomics
https://www.readbyqxmd.com/read/28916539/comparative-analysis-of-alternative-polyadenylation-in-s-cerevisiae-and-s-pombe
#3
Xiaochuan Liu, Mainul Hoque, Marc Larochelle, Jean-François Lemay, Nathan Yurko, James L Manley, François Bachand, Bin Tian
Alternative polyadenylation (APA) is a widespread mechanism that generates mRNA isoforms with distinct properties. Here we have systematically mapped and compared cleavage and polyadenylation sites (PASs) in two yeast species, S. cerevisiae and S. pombe Although >80% of the mRNA genes in each species were found to display APA, S. pombe showed greater 3' UTR size differences among APA isoforms than did S. cerevisiae PASs in different locations of gene are surrounded with distinct sequences in both species and are often associated with motifs involved in the Nrd1-Nab3-Sen1 termination pathway...
September 15, 2017: Genome Research
https://www.readbyqxmd.com/read/28893786/novel-cell-killing-mechanisms-of-hydroxyurea-and-the-implication-towards-combination-therapy-for-the-treatment-of-fungal-infections
#4
Amanpreet Singh, Ameeta Agarwal, Yong-Jie Xu
We have previously reported that an erg11 mutation affecting ergosterol synthesis and a hem13 mutation in the heme synthesis pathway significantly sensitize the fission yeast Schizosaccahromyces pombe to hydroxyurea (HU) (1, 2). Here we show that treatment with the inhibitors of Erg11 and the heme biosynthesis phenocopies the two mutations in sensitizing wild type cells to HU. Importantly, HU synergistically interacts with the heme biosynthesis inhibitor sampangine and several Erg11 inhibitors, the antifungal azoles, in causing cell lethality...
September 11, 2017: Antimicrobial Agents and Chemotherapy
https://www.readbyqxmd.com/read/28882432/analysis-of-ambient-ph-stress-response-mediated-by-iron-and-copper-intake-in-schizosaccharomyces-pombe
#5
Yujiro Higuchi, Hikari Mori, Takeo Kubota, Kaoru Takegawa
The molecular mechanism of tolerance to alkaline pH is well studied in model fungi Aspergillus nidulans and Saccharomyces cerevisiae. However, how fission yeast Schizosaccharomyces pombe survives under alkaline stress remains largely unknown, as the genes involved in the alkaline stress response pathways of A. nidulans and S. cerevisiae were not found in the genome of this organism. Since uptake of iron and copper into cells is important for alkaline tolerance in S. cerevisiae, here we examined whether iron and copper uptake processes were involved in conferring tolerance to alkaline stress in S...
September 4, 2017: Journal of Bioscience and Bioengineering
https://www.readbyqxmd.com/read/28871121/effects-of-cycloheximide-on-the-interpretation-of-ribosome-profiling-experiments-in-schizosaccharomyces-pombe
#6
Caia D S Duncan, Juan Mata
Stress conditions lead to global and gene-specific changes in RNA translation. Ribosome profiling experiments have identified genome-wide alterations in the distribution of ribosomes along mRNAs. However, it is contentious whether these changes reflect real responses, or whether they are artefacts caused by the use of inhibitors of translation (notably cycloheximide). To address this issue we performed ribosome profiling with the fission yeast Schizosaccharomyces pombe under conditions of exponential growth (unstressed) and nitrogen starvation (nutritional stress), and both in the presence and absence of cycloheximide...
September 4, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28844968/aging-and-immortality-in-unicellular-species
#7
REVIEW
Michael Florea
It has been historically thought that in conditions that permit growth, most unicellular species do not to age. This was particularly thought to be the case for symmetrically dividing species, as such species lack a clear distinction between the soma and the germline. Despite this, studies of the symmetrically dividing species Escherichia coli and Schizosaccharomyces pombe have recently started to challenge this notion. They indicate that E. coli and S. pombe do age, but only when subjected to environmental stress...
August 24, 2017: Mechanisms of Ageing and Development
https://www.readbyqxmd.com/read/28821619/dsc-e3-ligase-localization-to-the-golgi-requires-the-atpase-cdc48-and-cofactor-ufd1-for-activation-of-sterol-regulatory-element-binding-protein-in-fission-yeast
#8
Risa Burr, Diedre Ribbens, Sumana Raychaudhuri, Emerson V Stewart, Jason Ho, Peter J Espenshade
Sterol regulatory element-binding proteins (SREBPs) in the fission yeast Schizosaccharomyces pombe regulate lipid homeostasis and the hypoxic response under conditions of low sterol or oxygen availability. SREBPs are cleaved in the Golgi through the combined action of the Dsc E3 ligase complex, the rhomboid protease Rbd2, and the essential ATPases Associated with diverse cellular Activities (AAA+) ATPase Cdc48. The soluble SREBP N-terminal transcription factor domain is then released in the cytosol to enter the nucleus and regulate gene expression...
August 18, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28808035/fission-yeast-myosin-myo2-is-down-regulated-in-actin-affinity-by-light-chain-phosphorylation
#9
Luther W Pollard, Carol S Bookwalter, Qing Tang, Elena B Krementsova, Kathleen M Trybus, Susan Lowey
Studies in fission yeast Schizosaccharomyces pombe have provided the basis for the most advanced models of the dynamics of the cytokinetic contractile ring. Myo2, a class-II myosin, is the major source of tension in the contractile ring, but how Myo2 is anchored and regulated to produce force is poorly understood. To enable more detailed biochemical/biophysical studies, Myo2 was expressed in the baculovirus/Sf9 insect cell system with its two native light chains, Rlc1 and Cdc4. Milligram yields of soluble, unphosphorylated Myo2 were obtained that exhibited high actin-activated ATPase activity and in vitro actin filament motility...
August 29, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28807855/spatial-organization-and-molecular-interactions-of-the-schizosaccharomyces-pombe-ccq1-tpz1-poz1-shelterin-complex
#10
Harry Scott, Jin-Kwang Kim, Clinton Yu, Lan Huang, Feng Qiao, Derek J Taylor
The shelterin complex is a macromolecular assembly of proteins that binds to and protects telomeric DNA, which composes the ends of all linear chromosomes. Shelterin proteins prevent chromosome ends from fusing together and from eliciting erroneous induction of DNA damage response pathways. In addition, shelterin proteins play key roles in regulating the recruitment and activation of telomerase, an enzyme that extends telomeric DNA. In fission yeast, Schizosaccharomyces pombe, interactions between the shelterin proteins Ccq1, Tpz1, and Poz1 are important for regulating telomerase-mediated telomere synthesis and thus telomere length homeostasis...
August 12, 2017: Journal of Molecular Biology
https://www.readbyqxmd.com/read/28806726/replication-fork-slowing-and-stalling-are-distinct-checkpoint-independent-consequences-of-replicating-damaged-dna
#11
Divya Ramalingam Iyer, Nicholas Rhind
In response to DNA damage during S phase, cells slow DNA replication. This slowing is orchestrated by the intra-S checkpoint and involves inhibition of origin firing and reduction of replication fork speed. Slowing of replication allows for tolerance of DNA damage and suppresses genomic instability. Although the mechanisms of origin inhibition by the intra-S checkpoint are understood, major questions remain about how the checkpoint regulates replication forks: Does the checkpoint regulate the rate of fork progression? Does the checkpoint affect all forks, or only those encountering damage? Does the checkpoint facilitate the replication of polymerase-blocking lesions? To address these questions, we have analyzed the checkpoint in the fission yeast Schizosaccharomyces pombe using a single-molecule DNA combing assay, which allows us to unambiguously separate the contribution of origin and fork regulation towards replication slowing, and allows us to investigate the behavior of individual forks...
August 2017: PLoS Genetics
https://www.readbyqxmd.com/read/28801462/protein-glutaminylation-is-a-yeast-specific-posttranslational-modification-of-elongation-factor-1a
#12
Thomas Jank, Yury Belyi, Christophe Wirth, Sabine Rospert, Zehan Hu, Jörn Dengjel, Tina Tzivelekidis, Gregers Rom Andersen, Carola Hunte, Andreas Schlosser, Klaus Aktories
Ribosomal translation factors are fundamental for protein synthesis and highly conserved in all kingdoms of life. The essential eukaryotic elongation factor 1A (eEF1A), delivers aminoacyl tRNAs to the A-site of the translating 80S ribosome. Several studies have revealed that eEF1A is posttranslationally modified. Using MS analysis, site-directed mutagenesis, and X-ray structural data analysis of Saccharomyces cerevisiae eEF1A, we identified a posttranslational modification in which the alpha amino group of mono-L-glutamine is covalently linked to the side chain of glutamate 45 in eEF1A...
August 11, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28789907/benzothiazole-analogues-synthesis-characterization-mo-calculations-with-pm6-and-dft-in-silico-studies-and-in-vitro-antimalarial-as-dhfr-inhibitors-and-antimicrobial-activities
#13
Sampark S Thakkar, Parth Thakor, Arabinda Ray, Hiren Doshi, Vasudev R Thakkar
Benzothiazole analogues are of interest due to their potential activity against malarial and microbial infections. In search of suitable antimicrobial and antimalarial agents, we report here the synthesis, characterization and biological activities of benzothiazole analogues (J 1-J 10). The molecules were characterized by IR, Mass, (1)H NMR, (13)C NMR and elemental analysis. The in vitro antimicrobial activity was investigated against pathogenic strains; the results were explained with the help of DFT and PM6 molecular orbital calculations...
July 29, 2017: Bioorganic & Medicinal Chemistry
https://www.readbyqxmd.com/read/28775153/screening-and-purification-of-natural-products-from-actinomycetes-that-affect-the-cell-shape-of-fission-yeast
#14
R A Lewis, J-J Li, N E E Allenby, J Errington, J Hayles, P Nurse
This study was designed to identify bioactive compounds that alter the cellular shape of the fission yeast Schizosaccharomyces pombe, by affecting functions involved in the cell cycle or cell morphogenesis. We used a multidrug sensitive fission yeast strain, SAK950 to screen a library of 657 actinomycete bacteria and identified 242 strains that induced eight different major shape phenotypes in S. pombe. These include the typical cell cycle related phenotype, Elongated cells and cell morphology related phenotype Rounded cells...
August 3, 2017: Journal of Cell Science
https://www.readbyqxmd.com/read/28765303/genetic-analysis-of-schizosaccharomyces-pombe
#15
Karl Ekwall, Geneviève Thon
In this introduction we discuss some basic genetic tools and techniques that are used with the fission yeast Schizosaccharomyces pombe Genes commonly used for selection or as reporters are discussed, with an emphasis on genes that permit counterselection, intragenic complementation, or colony-color assays. S. pombe is most stable as a haploid organism. We describe its mating-type system, how to perform genetic crosses and methods for selecting and propagating diploids. We discuss the relative merits of tetrad dissection and random spore preparation in strain construction and genetic analyses...
August 1, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28752814/a-matter-of-scale-and-dimensions-chromatin-of-chromosome-landmarks-in-the-fungi
#16
Allyson A Erlendson, Steven Friedman, Michael Freitag
Chromatin and chromosomes of fungi are highly diverse and dynamic, even within species. Much of what we know about histone modification enzymes, RNA interference, DNA methylation, and cell cycle control was first addressed in Saccharomyces cerevisiae, Schizosaccharomyces pombe, Aspergillus nidulans, and Neurospora crassa. Here, we examine the three landmark regions that are required for maintenance of stable chromosomes and their faithful inheritance, namely, origins of DNA replication, telomeres and centromeres...
July 2017: Microbiology Spectrum
https://www.readbyqxmd.com/read/28733418/a-simple-method-to-induce-meiosis-and-sporulation-semisynchronously-in-the-fission-yeast-schizosaccharomyces-pombe
#17
Akira Yamashita, Takeshi Sakuno, Yoshinori Watanabe, Masayuki Yamamoto
Schizosaccharomyces pombe cells initiate a sexual differentiation program, which comprises meiosis and spore formation, on nitrogen starvation. This protocol describes a simple procedure to induce meiosis and sporulation semisynchronously in heterozygous diploid S. pombe cells. The procedure is appropriate for a variety of applications, including fluorescence-activated cell sorting (FACS) and northern and western blotting. Zygotic meiosis can also be induced by the same procedure, although less synchronously...
July 21, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28733417/analysis-of-schizosaccharomyces-pombe-meiosis
#18
Akira Yamashita, Takeshi Sakuno, Yoshinori Watanabe, Masayuki Yamamoto
Meiosis is a specialized cell cycle that generates haploid gametes from diploid cells. The fission yeast Schizosaccharomyces pombe is one of the best model organisms for studying the regulatory mechanisms of meiosis. S. pombe cells, which normally grow in the haploid state, diploidize by conjugation and initiate meiosis when starved for nutrients, especially nitrogen. Following two rounds of chromosome segregation, spore formation takes place. The switch from mitosis to meiosis is controlled by a kinase, Pat1, and an RNA-binding protein, Mei2...
July 21, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28733412/schizosaccharomyces-japonicus-a-distinct-dimorphic-yeast-among-the-fission-yeasts
#19
Keita Aoki, Kanji Furuya, Hironori Niki
Genomic sequencing data and morphological properties demonstrate evolutionary relationships among groups of the fission yeast, Schizosaccharomyces Phylogenetically, S. japonicus is the furthest removed from other species of fission yeast. The basic characteristics of cell proliferation are shared among all fission yeast, including the process of binary fission during vegetative growth, conjugation and karyogamy with horsetail movement, mating-type switching, and sporulation. However, S. japonicus also exhibits characteristics that are unique to filamentous fungi...
July 21, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28733407/fission-yeast-cell-wall-analysis
#20
Pilar Pérez, Juan C Ribas
The Schizosaccharomyces pombe cell wall is a rigid exoskeletal structure mainly composed of interlinked glucose polysaccharides and galactomannoproteins. It is essential for survival of the fission yeast, as it prevents cells from bursting from internal turgor pressure and protects them from mechanical injuries. Additionally, the cell wall determines the cell shape and, therefore, a better knowledge of cell wall structure and composition could provide valuable data in S. pombe morphogenetic studies. Here, we provide information about this structure and the current reliable methods for rapid analysis of the cell wall polymers by specific enzymatic and chemical degradations of purified cell walls...
July 21, 2017: Cold Spring Harbor Protocols
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