scholarly journals Absence of step changes in activity of certain enzymes during the cell cycle of budding and fission yeasts in synchronous cultures

1983 ◽  
Vol 61 (1) ◽  
pp. 339-349
Author(s):  
J. Creanor ◽  
S.G. Elliott ◽  
Y.C. Bisset ◽  
J.M. Mitchison
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Acid Phosphatase ◽  
Kluyveromyces Lactis ◽  
The Other ◽  
Linear Pattern ◽  
Synchronous Cultures ◽  
Asynchronous Control ◽  
Alpha Glucosidase ◽  
Beta Galactosidase

Synchronous cultures prepared by selection from an elutriating rotor were used to measure activity changes during the cell cycle of the following enzymes: acid phosphatase in Schizosaccharomyces pombe and Saccharomyces cerevisiae, alpha-glucosidase in S. cerevisiae and beta-galactosidase in Kluyveromyces lactis. There was no sign of step rises in activity in acid phosphatase but there were indications in S. cerevisiae of the linear pattern with rate doublings once per cycle that had been found previously in S. pombe. There was also no sign of step rises in the other two enzymes, in contrast to earlier results using different techniques. Asynchronous control cultures showed little or no perturbations after the first hour.

Linear Synthesis of Sucrase and Phosphatases during the Cell Cycle of Schizosaccharomyces Pombe

1969 ◽  
Vol 5 (2) ◽  
pp. 373-391
Author(s):  
J. M. MITCHISON ◽  
J. CREANOR ◽  
D. A. WILLAMS
Keyword(s):  
Cell Cycle ◽  
Alkaline Phosphatase ◽  
Acid Phosphatase ◽  
Schizosaccharomyces Pombe ◽  
Critical Point ◽  
Eukaryotic Cell ◽  
Sharp Rise ◽  
Linear Pattern ◽  
Synchronous Cultures ◽  
Tentative Model

The synthesis of sucrase, acid phosphatase and alkaline phosphatase has been followed in synchronous cultures of the fission yeast Schizosaccharomyces pombe prepared by gradient sedimentation. These three enzymes follow a linear pattern of synthesis through the cell cycle, with a doubling in rate at a ‘critical point’ about one-fifth of the way through the cycle. Sucrase can be rapidly derepressed by lowering the glucose concentration in the medium. This has been used to measure the sucrase ‘potential’ or capacity to synthesize sucrase on derepression. The potential exists at all times in the cycle, and follows a stepwise pattern with a sharp rise at the critical point. These results suggest that the functional genome doubles at the critical point. Since, however, the period of DNA synthesis is nearly one-third of a cycle before this point, there must be an appreciable delay between chemical replication and functional replication of the genome. In this respect S. pombe, a eukaryotic cell, differs markedly from bacteria. Other physiological events take place near the critical point, and a tentative model is suggested of what may be happening at the chromosomal level. Experiments with cycloheximide indicate that there is a delay between the synthesis and the appearance of the active enzyme in the case of sucrase and alkaline phosphatase.

Sit4p Protein Phosphatase Is Required for Sensitivity of Saccharomyces cerevisiae to Kluyveromyces lactis Zymocin

Genetics ◽  
2001 ◽  
Vol 159 (4) ◽  
pp. 1479-1489
Author(s):  
Daniel Jablonowski ◽  
Andrew R Butler ◽  
Lars Fichtner ◽  
Donald Gardiner ◽  
Raffael Schaffrath ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Rna Polymerase Ii ◽  
Protein Phosphatase ◽  
Kluyveromyces Lactis ◽  
The Other ◽  
G1 Phase ◽  
Elongator Complex ◽  
Intracellular Target ◽  
Related Proteins

Abstract We have identified two Saccharomyces cerevisiae genes that, in high copy, confer resistance to Kluyveromyces lactis zymocin, an inhibitor that blocks cells in the G1 phase of the cell cycle prior to budding and DNA replication. One gene (GRX3) encodes a glutaredoxin and is likely to act at the level of zymocin entry into sensitive cells, while the other encodes Sap155p, one of a family of four related proteins that function positively and interdependently with the Sit4p protein phosphatase. Increased SAP155 dosage protects cells by influencing the sensitivity of the intracellular target and is unique among the four SAP genes in conferring zymocin resistance in high copy, but is antagonized by high-copy SAP185 or SAP190. Since cells lacking SIT4 or deleted for both SAP185 and SAP190 are also zymocin resistant, our data support a model whereby high-copy SAP155 promotes resistance by competition with the endogenous levels of SAP185 and SAP190 expression. Zymocin sensitivity therefore requires a Sap185p/Sap190p-dependent function of Sit4p protein phosphatase. Mutations affecting the RNA polymerase II Elongator complex also confer K. lactis zymocin resistance. Since sit4Δ and SAP-deficient strains share in common several other phenotypes associated with Elongator mutants, Elongator function may be a Sit4p-dependent process.

scholarly journals Recombination of Ty elements in yeast can be induced by a double-strand break.

Genetics ◽  
1995 ◽  
Vol 140 (1) ◽  
pp. 67-77 ◽  
Author(s):  
A Parket ◽  
O Inbar ◽  
M Kupiec
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Strand Break ◽  
Double Strand Break ◽  
The Other ◽  
Direct Repeats ◽  
Yeast Saccharomyces Cerevisiae ◽  
Low Frequencies ◽  
Ty Elements ◽  
Main Family

Abstract The Ty retrotransposons are the main family of dispersed repeated sequences in the yeast Saccharomyces cerevisiae. These elements are flanked by a pair of long terminal direct repeats (LTRs). Previous experiments have shown that Ty elements recombine at low frequencies, despite the fact that they are present in 30 copies per genome. This frequency is not highly increased by treatments that cause DNA damage, such as UV irradiation. In this study, we show that it is possible to increase the recombination level of a genetically marked Ty by creating a double-strand break in it. This break is repaired by two competing mechanisms: one of them leaves a single LTR in place of the Ty, and the other is a gene conversion event in which the marked Ty is replaced by an ectopically located one. In a strain in which the marked Ty has only one LTR, the double-strand break is repaired by conversion. We have also measured the efficiency of repair and monitored the progression of the cells through the cell-cycle. We found that in the presence of a double-strand break in the marked Ty, a proportion of the cells is unable to resume growth.

Trans-acting regulatory mutations that alter transcription of Saccharomyces cerevisiae histone genes

1987 ◽  
Vol 7 (12) ◽  
pp. 4204-4210
Author(s):  
M A Osley ◽  
D Lycan
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Fusion Gene ◽  
Chromosome Replication ◽  
Lacz Fusion ◽  
Histone Genes ◽  
Promoter Elements ◽  
Regulatory Mutations ◽  
Cycle Dependent ◽  
Beta Galactosidase

Using a Saccharomyces cerevisiae strain containing an integrated copy of an H2A-lacZ fusion gene, we screened for mutants which overexpressed beta-galactosidase as a way to identify genes which regulate transcription of the histone genes. Five recessive mutants with this phenotype were shown to contain altered regulatory genes because they had lost repression of HTA1 transcription which occurs upon inhibition of chromosome replication (D. E. Lycan, M. A. Osley, and L. Hereford, Mol. Cell. Biol. 7:614-621, 1987). Periodic transcription was affected in the mutants as well, since the HTA1 gene was transcribed during the G1 and G2 phases of the cell cycle, periods in the cell cycle when this gene is normally not expressed. A similar loss of cell cycle-dependent transcription was noted for two of the three remaining histone loci, while the HO and CDC9 genes continued to be expressed periodically. Using isolated promoter elements inserted into a heterologous cycl-lacZ fusion gene, we demonstrated that the mutations fell in genes which acted through a negative site in the TRT1 H2A-H2B promoter.

scholarly journals Structure-function relationships of the yeast cyclin-dependent kinase Pho85.

1995 ◽  
Vol 15 (10) ◽  
pp. 5482-5491 ◽  
Author(s):  
R C Santos ◽  
N C Waters ◽  
C L Creasy ◽  
L W Bergman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Structure Function ◽  
Cell Cycle Progression ◽  
Substrate Recognition ◽  
The Other ◽  
Cyclin Dependent Kinase ◽  
Cycle Progression ◽  
Cyclin Dependent Kinases ◽  
Induced Mutagenesis

The PHO85 gene of Saccharomyces cerevisiae encodes a cyclin-dependent kinase involved in both transcriptional regulation and cell cycle progression. Although a great deal is known concerning the structure, function, and regulation of the highly homologous Cdc28 protein kinase, little is known concerning these relationships in regard to Pho85. In this study, we constructed a series of Pho85-Cdc28 chimeras to map the region(s) of the Pho85 molecule that is critical for function of Pho85 in repression of acid phosphatase (PHO5) expression. Using a combination of site-directed and ethyl methanesulfonate-induced mutagenesis, we have identified numerous residues critical for either activation of the Pho85 kinase, interaction of Pho85 with the cyclin-like molecule Pho80, or substrate recognition. Finally, analysis of mutations analogous to those previously identified in either Cdc28 or cdc2 of Schizosaccharomyces pombe suggested that the inhibition of Pho85-Pho80 activity in mechanistically different from that seen in the other cyclin-dependent kinases.

scholarly journals The Yeast Elongator Histone Acetylase Requires Sit4-dependent Dephosphorylation for Toxin-Target Capacity

2004 ◽  
Vol 15 (3) ◽  
pp. 1459-1469 ◽  
Author(s):  
Daniel Jablonowski ◽  
Lars Fichtner ◽  
Michael J.R. Stark ◽  
Raffael Schaffrath
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Kluyveromyces Lactis ◽  
Toxin Complex ◽  
Cell Cycle Block ◽  
Resistant Cells

Kluyveromyces lactis zymocin, a heterotrimeric toxin complex, imposes a G1 cell cycle block on Saccharomyces cerevisiae that requires the toxin-target (TOT) function of holo-Elongator, a six-subunit histone acetylase. Here, we demonstrate that Elongator is a phospho-complex. Phosphorylation of its largest subunit Tot1 (Elp1) is supported by Kti11, an Elongator-interactor essential for zymocin action. Tot1 dephosphorylation depends on the Sit4 phosphatase and its associators Sap185 and Sap190. Zymocin-resistant cells lacking or overproducing Elongator-associator Tot4 (Kti12), respectively, abolish or intensify Tot1 phosphorylation. Excess Sit4·Sap190 antagonizes the latter scenario to reinstate zymocin sensitivity in multicopy TOT4 cells, suggesting physical competition between Sit4 and Tot4. Consistently, Sit4 and Tot4 mutually oppose Tot1 de-/phosphorylation, which is dispensable for integrity of holo-Elongator but crucial for the TOT-dependent G1 block by zymocin. Moreover, Sit4, Tot4, and Tot1 cofractionate, Sit4 is nucleocytoplasmically localized, and sit4Δ-nuclei retain Tot4. Together with the findings that sit4Δ and totΔ cells phenocopy protection against zymocin and the ceramide-induced G1 block, Sit4 is functionally linked to Elongator in cell cycle events targetable by antizymotics.

Galactokinase encoded by GAL1 is a bifunctional protein required for induction of the GAL genes in Kluyveromyces lactis and is able to suppress the gal3 phenotype in Saccharomyces cerevisiae

1991 ◽  
Vol 11 (11) ◽  
pp. 5454-5461
Author(s):  
J Meyer ◽  
A Walker-Jonah ◽  
C P Hollenberg
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Kluyveromyces Lactis ◽  
Regulatory Function ◽  
Bifunctional Protein ◽  
Leloir Pathway ◽  
Gal Genes ◽  
Beta Galactosidase ◽  
Gal1 Gene ◽  
Galactokinase Activity ◽  
Negative Mutant

We have analyzed a GAL1 mutant (gal1-r strain) of the yeast Kluyveromyces lactis which lacks the induction of beta-galactosidase and the enzymes of the Leloir pathway in the presence of galactose. The data show that the K. lactis GAL1 gene product has, in addition to galactokinase activity, a function required for induction of the lactose system. This regulatory function is not dependent on galactokinase activity, as it is still present in a galactokinase-negative mutant (gal1-209). Complementation studies in Saccharomyces cervisiae show that K. lactis GAL1 and gal1-209, but not gal1-r, complement the gal3 mutation. We conclude that the regulatory function of GAL1 in K. lactis soon after induction is similar to the function of GAL3 in S. cerevisiae.

scholarly journals The CDC25 protein of Saccharomyces cerevisiae promotes exchange of guanine nucleotides bound to ras.

1991 ◽  
Vol 11 (5) ◽  
pp. 2641-2646 ◽  
Author(s):  
S Jones ◽  
M L Vignais ◽  
J R Broach
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cyclic Amp ◽  
Genetic Analysis ◽  
Fusion Protein ◽  
Guanine Nucleotides ◽  
Biochemical Activity ◽  
Cdc25 Protein ◽  
Beta Galactosidase

The product of the CDC25 gene of Saccharomyces cerevisiae, in its capacity as an activator of the RAS/cyclic AMP pathway, is required for initiation of the cell cycle. In this report, we provide an identification of Cdc25p, the product of the CDC25 gene, and evidence that it promotes exchange of guanine nucleotides bound to Ras in vitro. Extracts of strains containing high levels of Cdc25p catalyze both removal of GDP from and the concurrent binding of GTP to Ras. This same activity is also obtained with an immunopurified Cdc25p-beta-galactosidase fusion protein, suggesting that Cdc25p participates directly in the exchange reaction. This biochemical activity is consistent with previous genetic analysis of CDC25 function.

scholarly journals Galactokinase encoded by GAL1 is a bifunctional protein required for induction of the GAL genes in Kluyveromyces lactis and is able to suppress the gal3 phenotype in Saccharomyces cerevisiae.

1991 ◽  
Vol 11 (11) ◽  
pp. 5454-5461 ◽  
Author(s):  
J Meyer ◽  
A Walker-Jonah ◽  
C P Hollenberg
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Kluyveromyces Lactis ◽  
Regulatory Function ◽  
Bifunctional Protein ◽  
Leloir Pathway ◽  
Gal Genes ◽  
Beta Galactosidase ◽  
Gal1 Gene ◽  
Galactokinase Activity ◽  
Negative Mutant

We have analyzed a GAL1 mutant (gal1-r strain) of the yeast Kluyveromyces lactis which lacks the induction of beta-galactosidase and the enzymes of the Leloir pathway in the presence of galactose. The data show that the K. lactis GAL1 gene product has, in addition to galactokinase activity, a function required for induction of the lactose system. This regulatory function is not dependent on galactokinase activity, as it is still present in a galactokinase-negative mutant (gal1-209). Complementation studies in Saccharomyces cervisiae show that K. lactis GAL1 and gal1-209, but not gal1-r, complement the gal3 mutation. We conclude that the regulatory function of GAL1 in K. lactis soon after induction is similar to the function of GAL3 in S. cerevisiae.

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