scholarly journals A fission yeast general translation factor reveals links between protein synthesis and cell cycle controls

2000 ◽  
Vol 113 (8) ◽  
pp. 1447-1458 ◽  
Author(s):  
B. Grallert ◽  
S.E. Kearsey ◽  
M. Lenhard ◽  
C.R. Carlson ◽  
P. Nurse ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Fission Yeast ◽  
Single Gene ◽  
Mitotic Cell ◽  
Anaphase Promoting Complex ◽  
Wild Type ◽  
Rna Helicases ◽  
Translation Factor ◽  
And Function ◽  
General Translation

In two independent screens we isolated fission yeast mutations with phenotypes suggesting defects in B-cyclin function or expression. These mutations define a single gene which we call ded1. We show that ded1 encodes a general translation factor that is related in sequence and function to RNA helicases required for translation in other species. Levels of the B-cyclins Cig2 and Cdc13 are dramatically reduced upon inactivation of Ded1, and this reduction is independent of degradation by the anaphase promoting complex. When a ded1 mutant is grown under semi-restrictive conditions, the translation of Cig2 (and to a lesser extent Cdc13), is impaired relative to other proteins. We show that B-cyclin translation is specifically inhibited upon nitrogen starvation of wild-type cells, when B-cyclin/Cdc2 inactivation is a prerequisite for G(1) arrest and subsequent mating. Our data suggest that translational inhibition of B-cyclin expression represents a third mechanism, in addition to cyclin degradation and Rum1 inhibition, that contributes to Cdc2 inactivation as cells exit from the mitotic cell cycle and prepare for meiosis.

scholarly journals pkl1 +andklp2 +: Two Kinesins of the Kar3 Subfamily in Fission Yeast Perform Different Functions in Both Mitosis and Meiosis

2001 ◽  
Vol 12 (11) ◽  
pp. 3476-3488 ◽  
Author(s):  
Cynthia L. Troxell ◽  
Mark A. Sweezy ◽  
Robert R. West ◽  
Karen D. Reed ◽  
Bryan D. Carson ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Sexual Reproduction ◽  
Fission Yeast ◽  
Wild Type ◽  
Mitotic Chromosomes ◽  
Double Deletion ◽  
Early Anaphase ◽  
Spindle Disassembly ◽  
And Function ◽  
Mitosis And Meiosis

We have identified Klp2p, a new kinesin-like protein (KLP) of the KAR3 subfamily in fission yeast. The motor domain of this protein is 61% identical and 71% similar to Pkl1p, another fission yeast KAR3 protein, yet the two enzymes are different in behavior and function. Pkl1p is nuclear throughout the cell cycle, whereas Klp2p is cytoplasmic during interphase. During mitosis Klp2p enters the nucleus where it forms about six chromatin-associated dots. In metaphase-arrested cells these migrate back and forth across the nucleus. During early anaphase they segregate with the chromosomes into two sets of about three, fade, and are replaced by other dots that form on the spindle interzone. Neitherklp2 + norpkl1 + is essential, and the double deletion is also wild type for both vegetative and sexual reproduction. Each deletion rescues different alleles ofcut7 ts , a KLP that contributes to spindle formation and elongation. When either or both deletions are combined with a dynein deletion, vegetative growth is normal, but sexual reproduction fails: klp2Δ,dhc1-d1 in karyogamy, pkl1Δ,dhc1-d1 in multiple phases of meiosis, and the triple deletion in both. Deletion of Klp2p elongates a metaphase-arrested spindle, but pkl1Δshortens it. The anaphase spindle of klp2Δ becomes longer than the cell, leading it to curl around the cell's ends. Apparently, Klp2p promotes spindle disassembly and contributes to the behavior of mitotic chromosomes.

Cytoskeletal and DNA structure abnormalities result from bypass of requirement for the cdc10 start gene in the fission yeast Schizosaccharomyces pombe

1992 ◽  
Vol 101 (3) ◽  
pp. 517-528 ◽  
Author(s):  
J. Marks ◽  
C. Fankhauser ◽  
A. Reymond ◽  
V. Simanis
Keyword(s):  
Cell Cycle ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Null Allele ◽  
Single Gene ◽  
Dna Structure ◽  
Mitotic Cell ◽  
Normal Function ◽  
Mitotic Cell Cycle ◽  
Structural Abnormalities

The cdc10 gene of the fission yeast S. pombe is required for traverse of the start control in late G1 and commitment to the mitotic cell cycle. To increase our understanding of the events which occur at start, a pseudoreversion analysis was undertaken to identify genes whose products may interact with cdc10 or bypass the requirement for it. A single gene, sct1+ (suppressor of cdc ten), has been identified, mutation of which suppresses all conditional alleles and a null allele of cdc10. Bypass of the requirement for cdc10+ function by sct1-1 mutations leads to pleiotropic defects, including microtubule, microfilament and nuclear structural abnormalities. Our data suggest that sct1 encodes a protein that is dependent upon cdc10+ either for its normal function or expression, or is a component of a checkpoint that monitors execution of p85cdc10 function.

Asymmetry of the spindle pole bodies and spg1p GAP segregation during mitosis in fission yeast

1999 ◽  
Vol 112 (14) ◽  
pp. 2313-2321 ◽  
Author(s):  
L. Cerutti ◽  
V. Simanis
Keyword(s):  
Cell Cycle ◽  
Fission Yeast ◽  
Spindle Pole Body ◽  
Mitotic Cell ◽  
Spindle Pole ◽  
The Other ◽  
Septum Formation ◽  
Spindle Pole Bodies ◽  
Interphase Cells ◽  
Early Mitosis

In the fission yeast Schizosaccharomyces pombe, the onset of septum formation is induced by a signal transduction network involving several protein kinases and a GTPase switch. One of the roles of the spg1p GTPase is to localise the cdc7p protein kinase to the poles of the mitotic spindle, from where the onset of septation is thought to be signalled at the end of mitosis. Immunofluorescence studies have shown that cdc7p is located on both spindle pole bodies early in mitosis, but only on one during the later stages of anaphase. This is mediated by inactivation of spg1p on one pole before the other. The GAP for spg1p is a complex of two proteins, cdc16p and byr4p. Localisation of cdc16p and byr4p by indirect immunofluorescence during the mitotic cell cycle showed that both proteins are present on the spindle pole body in interphase cells. During mitosis, byr4p is seen first on both poles of the spindle, then on only one. This occurs prior to cdc7p becoming asymmetric. In contrast, the signal due to cdc16p decreases to a low level during early mitosis, before being seen strongly on the same pole as byr4p. Double staining indicates that this is the opposite pole to that which retains cdc7p in late anaphase. Examination of the effect of inactivating cdc16p at various stages of the cell cycle suggests that cdc16p, together with cdc2p plays a role in restraining septum formation during interphase. The asymmetric inactivation of spg1p is mediated by recruitment of the cdc16p-byr4p GAP to one of the poles of the spindle before the other, and the asymmetry of the spindle pole bodies may be established early during mitosis. Moreover, the spindle pole bodies appear to be non-equivalent even after division has been completed.

Polypeptide synthesis in cell cycle mutants of fission yeast

1981 ◽  
Vol 51 (1) ◽  
pp. 203-217
Author(s):  
D.P. Dickinson
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Fission Yeast ◽  
Gel Electrophoresis ◽  
Mutant Cell ◽  
Wild Type ◽  
Unsuccessful Attempt ◽  
Dependent Sequence ◽  
Polypeptide Synthesis ◽  
Cellular Components

The cell cycle of a growing cel is characterized by 3 main periodic events: DNA synthesis mitosis and cell division. These events generally lie in a dependent sequence, in which one event cannot occur unless preceding events have occurred. The existence of dependent sequences of events raises the possibility that at least some of the gene products involved in the events are synthesized in a dependent sequence parallel to the observable events. To test this hypothesis, the patterns of polypeptide synthesis were investigated in 2 types of cell cycle mutant of the fission yeast Schizosaccharomyces pombe: temperature-sensitive cell cycle (ts cdc) mutants. which become blocked in cell cycle progress at the restrictive temperature; and wee I mutants, which are defective in size control over nuclear division, and which divide at a small size. Cells of mutants and wild-type cells were labelled with [35S[sulphate under conditions designed to maximize any differences between the labelling patterns of wild-type and mutant cell polypeptides. The polypeptides were then separated by O'Farrell 2-dimensional gel electrophoresis, and the patterns compared. Although both types of mutation affect cell cycle control, and cause a considerable alteration in the relative proportions of cellular components, an examination of over 700 polypeptides detected on gels revealed no qualitative differences between wild-type and mutant cell polypeptides. These results suggest that a large majority of the more abundant polypeptides in the growing cell are synthesized independently of cell cycle controls directly related to DNA synthesis and division, and that the synthesis of these polypeptides can occur in the absence of normal progress through the cell cycle. Dependent sequences of gene expression do not appear to make a significant contribution to total polypeptide synthesis during the cell cycle, or to the occurrence of periodic cell cycle events such as mitosis. It is suggested that such cell cycle events may result largely through the reorganization of existing cellular components, rather than by the synthesis of new ones. An unsuccessful attempt was made to detect the wee I gene product on gels by surveying a range of mutants for changes in an individual spot. The limitations of gel electrophoresis for this type of survey, and other cell cycle experiments, are discussed.

scholarly journals Two distinct ubiquitin-proteolysis pathways in the fission yeast cell cycle

1999 ◽  
Vol 354 (1389) ◽  
pp. 1551-1557 ◽  
Author(s):  
Takashi Toda ◽  
Itziar Ochotorena ◽  
Kin-ichiro Kominami
Keyword(s):  
Cell Cycle ◽  
Fission Yeast ◽  
Cell Cycle Control ◽  
Eukaryotic Cell ◽  
Cdk Inhibitor ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Repeat Proteins ◽  
Cycle Dependent ◽  
Universal Mechanism

The SCF complex (Skp1-Cullin-1-F-box) and the APC/cyclosome (anaphase-promoting complex) are two ubiquitin ligases that play a crucial role in eukaryotic cell cycle control. In fission yeast F-box/WD-repeat proteins Pop1 and Pop2, components of SCF are required for cell-cycle-dependent degradation of the cyclin-dependent kinase (CDK) inhibitor Rum1 and the S-phase regulator Cdc18. Accumulation of these proteins in pop1 and pop2 mutants leads to re-replication and defects in sexual differentiation. Despite structural and functional similarities, Pop1 and Pop2 are not redundant homologues. Instead, these two proteins form heterodimers as well as homodimers, such that three distinct complexes, namely SCF Pop1/Pop1 , SCF Pop1/Pop2 and SCF Pop2/Pop2 , appear to exist in the cell. The APC/cyclosome is responsible for inactivation of CDK/cyclins through the degradation of B-type cyclins. We have identified two novel components or regulators of this complex, called Apc10 and Ste9, which are evolutionarily highly conserved. Apc10 (and Ste9), together with Rum1, are required for the establishment of and progression through the G1 phase in fission yeast. We propose that dual downregulation of CDK, one via the APC/cyclosome and the other via the CDK inhibitor, is a universal mechanism that is used to arrest the cell cycle at G1.

scholarly journals Inactivation of the selB Gene in Methanococcus maripaludis: Effect on Synthesis of Selenoproteins and Their Sulfur-Containing Homologs

2003 ◽  
Vol 185 (1) ◽  
pp. 107-114 ◽  
Author(s):  
Michael Rother ◽  
Isabella Mathes ◽  
Friedrich Lottspeich ◽  
August Böck
Keyword(s):  
Gene Product ◽  
Formate Dehydrogenase ◽  
Selenium Supplementation ◽  
Wild Type ◽  
Methanococcus Maripaludis ◽  
Translation Factor ◽  
Type Form ◽  
And Function ◽  
Sulfur Containing ◽  
Wild Type Form

ABSTRACT The genome of Methanococcus maripaludis harbors genes for at least six selenocysteine-containing proteins and also for homologs that contain a cysteine codon in the position of the UGA selenocysteine codon. To investigate the synthesis and function of both the Se and the S forms, a mutant with an inactivated selB gene was constructed and analyzed. The mutant was unable to synthesize any of the selenoproteins, thus proving that the gene product is the archaeal translation factor (aSelB) specialized for selenocysteine insertion. The wild-type form of M. maripaludis repressed the synthesis of the S forms of selenoproteins, i.e., the selenium-independent alternative system, in selenium-enriched medium, but the mutant did not. We concluded that free selenium is not involved in regulation but rather a successional compound such as selenocysteyl-tRNA or some selenoprotein. Apart from the S forms, several enzymes from the general methanogenic route were affected by selenium supplementation of the wild type or by the selB mutation. Although the growth of M. maripaludis on H2/CO2 is only marginally affected by the selB lesion, the gene is indispensable for growth on formate because M. maripaludis possesses only a selenocysteine-containing formate dehydrogenase.

scholarly journals Validated Bayesian Differentiation of Causative and Passenger Mutations

2017 ◽  
Vol 7 (7) ◽  
pp. 2081-2094 ◽  
Author(s):  
Frederick R Cross ◽  
Michal Breker ◽  
Kristi Lieberman
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Mitotic Cell ◽  
Causative Mutation ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Coding Sequence ◽  
New Genes ◽  
Calculated Probability ◽  
Passenger Mutations

Abstract In many contexts, the problem arises of determining which of many candidate mutations is the most likely to be causative for some phenotype. It is desirable to have a way to evaluate this probability that relies as little as possible on previous knowledge, to avoid bias against discovering new genes or functions. We have isolated mutants with blocked cell cycle progression in Chlamydomonas and determined mutant genome sequences. Due to the intensity of UV mutagenesis required for efficient mutant collection, the mutants contain multiple mutations altering coding sequence. To provide a quantitative estimate of probability that each individual mutation in a given mutant is the causative one, we developed a Bayesian approach. The approach employs four independent indicators: sequence conservation of the mutated coding sequence with Arabidopsis; severity of the mutation relative to Chlamydomonas wild-type based on Blosum62 scores; meiotic mapping information for location of the causative mutation relative to known molecular markers; and, for a subset of mutants, the transcriptional profile of the candidate wild-type genes through the mitotic cell cycle. These indicators are statistically independent, and so can be combined quantitatively into a single probability calculation. We validate this calculation: recently isolated mutations that were not in the training set for developing the indicators, with high calculated probability of causality, are confirmed in every case by additional genetic data to indeed be causative. Analysis of “best reciprocal BLAST” (BRB) relationships among Chlamydomonas and other eukaryotes indicate that the temperature sensitive-lethal (Ts-lethal) mutants that our procedure recovers are highly enriched for fundamental cell-essential functions conserved broadly across plants and other eukaryotes, accounting for the high information content of sequence alignment to Arabidopsis.

scholarly journals Mid2p stabilizes septin rings during cytokinesis in fission yeast

2003 ◽  
Vol 160 (7) ◽  
pp. 1083-1092 ◽  
Author(s):  
Ana Berlin ◽  
Anne Paoletti ◽  
Fred Chang
Keyword(s):  
Fission Yeast ◽  
Cell Separation ◽  
Sequence Similarity ◽  
Ring Closure ◽  
Pleckstrin Homology Domain ◽  
Wild Type ◽  
Contractile Ring ◽  
Single Ring ◽  
And Function ◽  
Cell Cell

Septins are filament-forming proteins with a conserved role in cytokinesis. In the fission yeast Schizosaccharomyces pombe, septin rings appear to be involved primarily in cell–cell separation, a late stage in cytokinesis. Here, we identified a protein Mid2p on the basis of its sequence similarity to S. pombe Mid1p, Saccharomyces cerevisiae Bud4p, and Candida albicans Int1p. Like septin mutants, mid2Δ mutants had delays in cell–cell separation. mid2Δ mutants were defective in septin organization but not contractile ring closure or septum formation. In wild-type cells, septins assembled first during mitosis in a single ring and during septation developed into double rings that did not contract. In mid2Δ cells, septins initially assembled in a single ring but during septation appeared in the cleavage furrow, forming a washer or disc structure. FRAP studies showed that septins are stable in wild-type cells but exchange 30-fold more rapidly in mid2Δ cells. Mid2p colocalized with septins and required septins for its localization. A COOH-terminal pleckstrin homology domain of Mid2p was required for its localization and function. No genetic interactions were found between mid2 and the related gene mid1. Thus, these studies identify a new factor responsible for the proper stability and function of septins during cytokinesis.

scholarly journals A mitotic role for a novel fission yeast protein kinase dsk1 with cell cycle stage dependent phosphorylation and localization.

1993 ◽  
Vol 4 (3) ◽  
pp. 247-260 ◽  
Author(s):  
M Takeuchi ◽  
M Yanagida
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Fission Yeast ◽  
Cell Cycle Progression ◽  
Kinase Activity ◽  
Specific Protein ◽  
Wild Type ◽  
Mobility Shift ◽  
Multicopy Suppressor ◽  
Phosphorylation Pattern

The fission yeast dsk1+ gene, a multicopy suppressor for cold-sensitive dis1 mutants, encodes a novel 61-kd protein kinase. It is a phosphoprotein, and phosphoserine is the major phosphorylated amino acid. Hyperphosphorylation of dsk1 causes a mobility shift, resulting in two dsk1-specific protein bands. The phosphorylation pattern is strikingly altered when cell cycle progression is delayed or arrested. The slowly migrating phosphorylated form is prominent in mitotically arrested cells, and the fast migrating form is enriched in interphase-arrested cells. dsk1 is a protein kinase. It auto-phosphorylates as well as phosphorylates myelin basic protein (MBP). Phosphotyrosine as well as phosphoserine/threonine were found in autophosphorylation, but no tyrosine phosphorylation occurs when MBP was used as the substrate. The dsk1 immunoprecipitates from mitotically arrested cells have a several-fold higher kinase activity than that from wild type. The haploid gene disruptant is viable, indicating that the dsk1+ gene is non-essential for viability. High dosage of dsk1+, however, strongly delays the G2/M progression. Immunofluorescence microscopy using anti-dsk1 antibody shows that localization pattern of dsk1 protein strikingly alters depending on cell cycle stages. In G2-arrested cells, dsk1 locates in the cytoplasm, whereas in mitotically arrested cells, nuclear stain is intense. In wild-type cells, nuclear stain is seen only in mitotic cells. Hence dsk1 protein may play an important role in mitotic control by altering cellular location, degree of phosphorylation and kinase activity. We discuss possible roles of dsk1 kinase as an add-on regulator in mitosis.

scholarly journals APC/C Cdh1-mediated degradation of Cdh1 during meiosis in S. cerevisiae

2018 ◽  
Author(s):  
Denis Ostapenko ◽  
Mark J. Solomon
Keyword(s):  
Cell Cycle ◽  
Binding Sites ◽  
Mitotic Cell ◽  
Sporulation Medium ◽  
Cell Cycle Regulators ◽  
Anaphase Promoting Complex ◽  
Cell Cycles ◽  
Activator Proteins ◽  
Subsequent Degradation ◽  
Numerous Cell

ABSTRACTThe Anaphase-Promoting Complex/Cyclosome (APC/C) is a ubiquitin ligase that promotes the ubiquitination and subsequent degradation of numerous cell cycle regulators during mitosis and in G1. Proteins are recruited to the APC/C by activator proteins such as Cdh1. During the cell cycle, Cdh1 is subject to precise regulation so that substrates are not degraded prematurely. We have explored the regulation of Cdh1 during the developmental transition into meiosis and sporulation in the budding yeast S. cerevisiae. Transition to sporulation medium triggers the degradation of Cdh1. Degradation requires that cells be of the a/a mating type and be starved for glucose, but they do not actually need to enter into the meiotic program. Degradation requires an intact SNF1 protein kinase complex (orthologous to the mammalian AMPK nutritional sensor), which is activated by the absence of glucose. Cdh1 degradation is mediated by the APC/C itself in a ‘trans’ mechanism in which one molecule of Cdh1 recruits a second molecule of Cdh1 to the APC/C for ubiquitination. However, Cdh1-Cdh1 recognition does not depend on the degradation motifs or binding sites involved in the recognition of typical APC/C substrates. We hypothesize that Cdh1 degradation is necessary for the preservation of cell cycle regulators and chromosome cohesion proteins between the reductional and equational meiotic divisions, which occur without the intervening Gap or S phases found in mitotic cell cycles.

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