scholarly journals Fission yeast Dss1 associates with the proteasome and is required for efficient ubiquitin-dependent proteolysis

2005 ◽  
Vol 393 (1) ◽  
pp. 303-309 ◽  
Author(s):  
Lyne Jossé ◽  
Margaret E. Harley ◽  
Isabel M. S. Pires ◽  
David A. Hughes
Keyword(s):  
Fission Yeast ◽  
Physiological Role ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Growth Defects ◽  
Tumour Suppressor Protein ◽  
Low Concentrations ◽  
Acid Analogue ◽  
Evolutionarily Conserved ◽  
Related Proteins

Human DSS1 associates with BRCA2, a tumour suppressor protein required for efficient recombinational DNA repair, but the biochemical function of DSS1 is not known. Orthologues of DSS1 are found in organisms such as budding yeast and fission yeast that do not have BRCA2-related proteins, indicating that DSS1 has a physiological role independent of BRCA2. The DSS1 orthologue in Saccharomyces cerevisiae has been shown to associate with the 26 S proteasome and, in the present paper, we report that in the distantly related fission yeast Schizosaccharomyces pombe, Dss1 associates with the 19 S RP (regulatory particle) of the 26 S proteasome. A role for S. pombe Dss1 in proteasome function is supported by three lines of evidence. First, overexpression of two components of the 19 S RP, namely Pad1/Rpn11 and Mts3/Rpn12, rescued the temperature-sensitive growth defect of the dss1 mutant. Secondly, the dss1 mutant showed phenotypes indicative of a defect in proteasome function: growth of the dss1 mutant was inhibited by low concentrations of L-canavanine, an amino acid analogue, and cells of the dss1 mutant accumulated high molecular mass poly-ubiquitylated proteins. Thirdly, synthetic growth defects were found when the dss1 mutation was combined with mutations in other proteasome subunit genes. These findings show that DSS1 has an evolutionarily conserved role as a regulator of proteasome function and suggest that DSS1 may provide a link between BRCA2 and ubiquitin-mediated proteolysis in human cells.

scholarly journals Scw1p Antagonizes the Septation Initiation Network To Regulate Septum Formation and Cell Separation in the Fission Yeast Schizosaccharomyces pombe

2003 ◽  
Vol 2 (3) ◽  
pp. 510-520 ◽  
Author(s):  
Quan-Wen Jin ◽  
Dannel McCollum
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Cell Separation ◽  
Deletion Mutant ◽  
Temperature Sensitive ◽  
Growth Defects ◽  
Septum Formation ◽  
Primary Septum ◽  
Septation Initiation Network ◽  
Ring Constriction

ABSTRACT Cytokinesis in the fission yeast Schizosaccharomyces pombe is regulated by a signaling pathway termed the septation initiation network (SIN). The SIN is essential for initiation of actomyosin ring constriction and septum formation. In a screen to search for mutations that can rescue the sid2-250 SIN mutant, we obtained scw1-18. Both the scw1-18 mutant and the scw1 deletion mutant (scw1Δ mutant), have defects in cell separation. Both the scw1-18 and scw1Δ mutations rescue the growth defects of not just the sid2-250 mutant but also the other temperature-sensitive SIN mutants. Other cytokinesis mutants, such as those defective for actomyosin ring formation, are not rescued by scw1Δ. scw1Δ does not seem to rescue the SIN by restoring SIN signaling defects. However, scw1Δ may function downstream of the SIN to promote septum formation, since scw1Δ can rescue the septum formation defects of the cps1-191β-1,3-glucan synthase mutant, which is required for synthesis of the primary septum.

The fission yeast bromodomain protein Bdf2 is required for growth of cells with circular chromosomes

2021 ◽  
Author(s):  
Misaki Yasuda ◽  
Ahmed G K Habib ◽  
Kanako Sugiura ◽  
Hossain Mohammad Shamim ◽  
Masaru Ueno
Keyword(s):  
Fission Yeast ◽  
High Temperatures ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
M Phase ◽  
Ts Mutant ◽  
Synthetically Lethal ◽  
Circular Chromosomes ◽  
Synthetic Growth

Abstract Circular chromosomes have frequently been observed in tumors of mesenchymal origin. In the fission yeast Schizosaccharomyces pombe, deletion of pot1+ results in rapid telomere loss, and the resulting survivors have circular chromosomes. Fission yeast has two bromodomains and extra-terminal (BET) proteins, Bdf1 and Bdf2; both are required for maintaining acetylated histones. Here, we found that bdf2, but not bdf1, was synthetically lethal with pot1. We also obtained a temperature-sensitive bdf2-ts mutant, which can grow at high temperatures but becomes camptothecin sensitive. This suggests that Bdf2 is defective at high temperatures. The cell cycle of the pot1 bdf2-ts mutant was delayed in the G2 and/or M phase at a semi-permissive temperature. Furthermore, a temperature-sensitive mutant of mst1, which encodes histone acetyltransferase, showed a synthetic growth defect with a pot1 disruptant at a semi-permissive temperature. Our results suggest that Bdf2 and Mst1 are required for the growth of cells with circular chromosomes.

scholarly journals Fission Yeast dim1+ Encodes a Functionally Conserved Polypeptide Essential for Mitosis

1997 ◽  
Vol 137 (6) ◽  
pp. 1337-1354 ◽  
Author(s):  
Lynne D. Berry ◽  
Kathleen L. Gould
Keyword(s):  
Fission Yeast ◽  
Chromosome Segregation ◽  
Mitotic Spindle ◽  
Kinase Activity ◽  
Nuclear Division ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Sensitive Mutant ◽  
Evolutionarily Conserved ◽  
Mutant Cells

In a screen for second site mutations capable of reducing the restrictive temperature of the fission yeast mutant cdc2-D217N, we have isolated a novel temperature-sensitive mutant, dim1-35. When shifted to restrictive temperature, dim1-35 mutant cells arrest before entry into mitosis or proceed through mitosis in the absence of nuclear division, demonstrating an uncoupling of proper DNA segregation from other cell cycle events. Deletion of dim1 from the Schizosaccharomyces pombe genome produces a lethal G2 arrest phenotype. Lethality is rescued by overexpression of the mouse dim1 homolog, mdim1. Likewise, deletion of the Saccharomyces cerevisiae dim1 homolog, CDH1, is lethal. Both mdim1 and dim1+ are capable of rescuing lethality in the cdh1::HIS3 mutant. Although dim1-35 displays no striking genetic interactions with various other G2/M or mitotic mutants, dim1-35 cells incubated at restrictive temperature arrest with low histone H1 kinase activity. Morevoer, dim1-35 displays sensitivity to the microtubule destabilizing drug, thiabendazole (TBZ). We conclude that Dim1p plays a fundamental, evolutionarily conserved role as a protein essential for entry into mitosis as well as for chromosome segregation during mitosis. Based on TBZ sensitivity and failed chromosome segregation in dim1-35, we further speculate that Dim1p may play a role in mitotic spindle formation and/or function.

scholarly journals Identification of Functionally Interacting SNAREs by Using Complementary Substitutions in the Conserved `0' Layer

2005 ◽  
Vol 16 (5) ◽  
pp. 2263-2274 ◽  
Author(s):  
Carmen T. Graf ◽  
Dietmar Riedel ◽  
Hans Dieter Schmitt ◽  
Reinhard Jahn
Keyword(s):  
Snare Complex ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Central Layer ◽  
Attachment Protein ◽  
Growth Defects ◽  
Protein Receptor ◽  
Sensitive Factor ◽  
Factor Attachment Protein Receptor

Soluble N-ethylmaleimide–sensitive factor attachment protein receptor (SNARE) complexes form bundles of four parallel α-helices. The central `0' layer of interacting amino acid side chains is highly conserved and contains one arginine and three glutamines, leading to the classification of SNAREs into R, Qa, Qb, and Qc-SNAREs. Replacing one of the glutamines with arginine in the yeast exocytotic SNARE complex is either lethal or causes a conditional growth defect that is compensated by replacing the R-SNARE arginine with glutamine. Using the yeast SNARE complex mediating traffic from the endoplasmic reticulum to the Golgi apparatus, we now show that functionally interacting SNAREs can be mapped by systematically exchanging glutamines and arginines in the `0' layer. The Q→ R replacement in the Qb-SNARE Bos1p has the strongest effect and can be alleviated by an Q→ R replacement in the R-SNARE Sec22p. Four Q residues in the central layer caused growth defects above 30°C that were rescued by Q→ R substitutions in the Qa and Qc SNAREs Sed5p and Bet1p, respectively. The sec22(Q)/sed5(R) mutant is temperature sensitive and is rescued by a compensating R→ Q replacement in the R-SNARE Ykt6p. This rescue is attributed to the involvement of Sed5p and Ykt6p in a different SNARE complex that functions in intra-Golgi trafficking.

scholarly journals Fission yeast polycystin Pkd2p promotes transition to cell growth during cytokinesis

2021 ◽  
Author(s):  
Debatrayee Sinha ◽  
Denisa Ivan ◽  
Ellie Gibbs ◽  
Madhurya Chetluru ◽  
John W Goss ◽  
...  
Keyword(s):  
Cell Growth ◽  
Fission Yeast ◽  
Genetic Disorders ◽  
Hippo Pathway ◽  
Cell Stiffness ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Size Growth ◽  
Autosomal Dominant Polycystic Kidney ◽  
The Hippo Pathway

Polycystins are evolutionally conserved cation channels. Mutations of human polycystins lead to one of the most common genetic disorders, Autosomal Dominant Polycystic Kidney Disorder. Interestingly, the fission yeast homologue Pkd2p is required for cytokinesis the last stage of cell division, but the mechanism remains unclear. Motivated by our discovery of the epistatic genetic interactions between pkd2 and the Hippo pathway Septation Initiation Network (SIN), we investigated their interplay during cytokinesis. We found that pkd2 modulated the localization as well as the activities of SIN. Most notably, pkd2 promotes a transition to cell size growth during cytokinesis, opposed by SIN. The role of Pkd2p in cell growth is not limited to cytokinesis. A newly isolated pkd2 temperature-sensitive mutant largely blocked the tip expansion of cells during interphase. Such growth defect was accompanied by frequent shrinking, reduced cell volume, and decreased cell stiffness. We conclude that Pkd2p promotes transition to the post-mitosis cell growth in coordination with the Hippo pathway

scholarly journals BAF53/Arp4 Homolog Alp5 in Fission Yeast Is Required for Histone H4 Acetylation, Kinetochore-Spindle Attachment, and Gene Silencing at Centromere

2005 ◽  
Vol 16 (1) ◽  
pp. 316-327 ◽  
Author(s):  
Aki Minoda ◽  
Shigeaki Saitoh ◽  
Kohta Takahashi ◽  
Takashi Toda
Keyword(s):  
Fission Yeast ◽  
Histone Deacetylases ◽  
Specific Inhibitor ◽  
Single Amino Acid ◽  
Temperature Sensitive ◽  
Histone H4 ◽  
Sensitive Mutant ◽  
Histone H4 Acetylation ◽  
Related Proteins ◽  
Yeast Homolog

Nuclear actin-related proteins play vital roles in transcriptional regulation; however, their biological roles remain elusive. Here, we characterize Alp5, fission yeast homolog of Arp4/BAF53. The temperature-sensitive mutant alp5-1134 contains a single amino acid substitution in the conserved C-terminal domain (S402N) and displays mitotic phenotypes, including chromosome condensation and missegregation. Alp5 forms a complex with Mst1-HAT (histone acetyltransferase). Consistently, inhibition of histone deacetylases (HDACs), by either addition of a specific inhibitor or a mutation in HDAC-encoding clr6+ gene, rescues alp5-1134. Immunoblotting with specific antibodies against acetylated histones shows that Alp5 is required for histone H4 acetylation at lysines 5, 8, and 12, but not histone H3 lysines 9 or 14, and furthermore Clr6 plays an opposing role. Mitotic arrest is ascribable to activation of the Mad2/Bub1 spindle checkpoint, in which both proteins localize to the mitotic kinetochores in alp5-1134. Intriguingly, alp5-1134 displays transcriptional desilencing at the core centromere without altering the overall chromatin structure, which also is suppressed by a simultaneous mutation in clr6+. This result shows that Alp5 is essential for histone H4 acetylation, and its crucial role lies in the establishment of bipolar attachment of the kinetochore to the spindle and transcriptional silencing at the centromere.

scholarly journals Differential Regulation of the Kar3p Kinesin-related Protein by Two Associated Proteins, Cik1p and Vik1p

1999 ◽  
Vol 144 (6) ◽  
pp. 1219-1233 ◽  
Author(s):  
Brendan D. Manning ◽  
Jennifer G. Barrett ◽  
Julie A. Wallace ◽  
Howard Granok ◽  
Michael Snyder
Keyword(s):  
Spindle Pole Body ◽  
Structural Similarity ◽  
Spindle Pole ◽  
Body Region ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Dependent Manner ◽  
Related Protein ◽  
Phenotypic Analysis ◽  
Related Proteins

The mechanisms by which kinesin-related proteins interact with other proteins to carry out specific cellular processes is poorly understood. The kinesin-related protein, Kar3p, has been implicated in many microtubule functions in yeast. Some of these functions require interaction with the Cik1 protein (Page, B.D., L.L. Satterwhite, M.D. Rose, and M. Snyder. 1994. J. Cell Biol. 124:507–519). We have identified a Saccharomyces cerevisiae gene, named VIK1, encoding a protein with sequence and structural similarity to Cik1p. The Vik1 protein is detected in vegetatively growing cells but not in mating pheromone-treated cells. Vik1p physically associates with Kar3p in a complex separate from that of the Kar3p-Cik1p complex. Vik1p localizes to the spindle-pole body region in a Kar3p-dependent manner. Reciprocally, concentration of Kar3p at the spindle poles during vegetative growth requires the presence of Vik1p, but not Cik1p. Phenotypic analysis suggests that Cik1p and Vik1p are involved in different Kar3p functions. Disruption of VIK1 causes increased resistance to the microtubule depolymerizing drug benomyl and partially suppresses growth defects of cik1Δ mutants. The vik1Δ and kar3Δ mutations, but not cik1Δ, partially suppresses the temperature-sensitive growth defect of strains lacking the function of two other yeast kinesin-related proteins, Cin8p and Kip1p. Our results indicate that Kar3p forms functionally distinct complexes with Cik1p and Vik1p to participate in different microtubule-mediated events within the same cell.

scholarly journals Fission yeast profilin is tailored to facilitate actin assembly by the cytokinesis formin Cdc12

2015 ◽  
Vol 26 (2) ◽  
pp. 283-293 ◽  
Author(s):  
Andrew J. Bestul ◽  
Jenna R. Christensen ◽  
Agnieszka P. Grzegorzewska ◽  
Thomas A. Burke ◽  
Jennifer A. Sees ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Genetic Selection ◽  
Actin Binding ◽  
Temperature Sensitive ◽  
Actin Assembly ◽  
Major Activity ◽  
Evolutionarily Conserved ◽  
Essential Properties

The evolutionarily conserved small actin-monomer binding protein profilin is believed to be a housekeeping factor that maintains a general pool of unassembled actin. However, despite similar primary sequences, structural folds, and affinities for G-actin and poly-l-proline, budding yeast profilin ScPFY fails to complement fission yeast profilin SpPRF temperature-sensitive mutant cdc3-124 cells. To identify profilin's essential properties, we built a combinatorial library of ScPFY variants containing either WT or SpPRF residues at multiple positions and carried out a genetic selection to isolate variants that support life in fission yeast. We subsequently engineered ScPFY(9-Mut), a variant containing nine substitutions in the actin-binding region, which complements cdc3-124 cells. ScPFY(9-Mut), but not WT ScPFY, suppresses severe cytokinesis defects in cdc3-124 cells. Furthermore, the major activity rescued by ScPFY(9-Mut) is the ability to enhance cytokinesis formin Cdc12-mediated actin assembly in vitro, which allows cells to assemble functional contractile rings. Therefore an essential role of profilin is to specifically facilitate formin-mediated actin assembly for cytokinesis in fission yeast.

scholarly journals Saccharomyces cerevisiae SMT4 Encodes an Evolutionarily Conserved Protease With a Role in Chromosome Condensation Regulation

Genetics ◽  
2001 ◽  
Vol 158 (1) ◽  
pp. 95-107 ◽  
Author(s):  
Alexander V Strunnikov ◽  
L Aravind ◽  
Eugene V Koonin
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Active Site ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Site Specific ◽  
Active Site Cysteine ◽  
Condensin Complex ◽  
Evolutionarily Conserved ◽  
Synthetically Lethal

Abstract In a search for regulatory genes affecting the targeting of the condensin complex to chromatin in Saccharomyces cerevisiae, we identified a member of the adenovirus protease family, SMT4. SMT4 overexpression suppresses the temperature-sensitive conditional lethal phenotype of smc2-6, but not smc2-8 or smc4-1. A disruption allele of SMT4 has a prominent chromosome phenotype: impaired targeting of Smc4p-GFP to rDNA chromatin. Site-specific mutagenesis of the predicted protease active site cysteine and histidine residues of Smt4p abolishes the SMT4 function in vivo. The previously uncharacterized SIZ1 (SAP and Miz) gene, which encodes a protein containing a predicted DNA-binding SAP module and a Miz finger, is identified as a bypass suppressor of the growth defect associated with the SMT4 disruption. The SIZ1 gene disruption is synthetically lethal with the SIZ2 deletion. We propose that SMT4, SIZ1, and SIZ2 are involved in a novel pathway of chromosome maintenance.

scholarly journals Systematic Humanization of the Yeast Cytoskeleton Discerns Functionally Replaceable from Divergent Human Genes

Genetics ◽  
2020 ◽  
Vol 215 (4) ◽  
pp. 1153-1169 ◽  
Author(s):  
Riddhiman K. Garge ◽  
Jon M. Laurent ◽  
Aashiq H. Kachroo ◽  
Edward M. Marcotte
Keyword(s):  
Gene Families ◽  
Growth Defect ◽  
Gene Duplications ◽  
Yeast Gene ◽  
Macromolecular Transport ◽  
Cellular Processes ◽  
Growth Defects ◽  
Yeast Strains ◽  
Human Genes ◽  
Interaction Partners

Many gene families have been expanded by gene duplications along the human lineage, relative to ancestral opisthokonts, but the extent to which the duplicated genes function similarly is understudied. Here, we focused on structural cytoskeletal genes involved in critical cellular processes, including chromosome segregation, macromolecular transport, and cell shape maintenance. To determine functional redundancy and divergence of duplicated human genes, we systematically humanized the yeast actin, myosin, tubulin, and septin genes, testing ∼81% of human cytoskeletal genes across seven gene families for their ability to complement a growth defect induced by inactivation or deletion of the corresponding yeast ortholog. In five of seven families—all but α-tubulin and light myosin, we found at least one human gene capable of complementing loss of the yeast gene. Despite rescuing growth defects, we observed differential abilities of human genes to rescue cell morphology, meiosis, and mating defects. By comparing phenotypes of humanized strains with deletion phenotypes of their interaction partners, we identify instances of human genes in the actin and septin families capable of carrying out essential functions, but failing to fully complement the cytoskeletal roles of their yeast orthologs, thus leading to abnormal cell morphologies. Overall, we show that duplicated human cytoskeletal genes appear to have diverged such that only a few human genes within each family are capable of replacing the essential roles of their yeast orthologs. The resulting yeast strains with humanized cytoskeletal components now provide surrogate platforms to characterize human genes in simplified eukaryotic contexts.

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