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.

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.

Changes in the Activity of Alcohol Dehydrogenase During the Cell Cycle of the Fission Yeast Schizosaccharomyces Pombe

1991 ◽  
Vol 99 (1) ◽  
pp. 193-199
Author(s):  
J. VICENTE-SOLER ◽  
J. CREANOR ◽  
Y. BISSET ◽  
J. M. MITCHISON
Keyword(s):  
Cell Cycle ◽  
Alcohol Dehydrogenase ◽  
Schizosaccharomyces Pombe ◽  
Rate Change ◽  
Co2 Production ◽  
Activity Increase ◽  
Normal Cycle ◽  
Linear Pattern ◽  
Synchronous Cultures ◽  
Coordinate Control

The activity of alcohol dehydrogenase (ADHase) was followed in synchronous cultures of Schizosaccharomyces pombe. In selection synchronised cultures of wild-type cells, it followed a linear pattern in which there was a constant rate of increase of activity followed by a doubling of this rate at the end of the cycle. The same pattern was also found in selection synchronised cells of wee mutants except that the point of rate change was shifted to 0.27 of the cycle. A similar linear pattern was also found in the shortened cell cycles produced by induction synchrony (block and release of the mutant cdc2.33) but the rate change point was at about 0.75 of the cycle. In the mutant cdcl3.117, there was a marked fall in the rate of activity increase at 35°C but not at 37°C. In all these situations, the ADHase activity closely paralleled in pattern and in timing the rate of production of CO2 established in earlier papers. This suggests a coordinate control of the flux through glycolysis and the activity of the last enzyme in the glycolytic pathway in yeast. However, an interesting difference indicating a loss of the coordinate control occurred in ‘synchronous’ cultures of cdc2.33 in which small cells had been selected but in which the DNAdivision cycle had been blocked by a shift-up to the restrictive temperature. Rate changes both in CO2 production and in ADHase activity continued in these blocked synchronous cultures but the timing was different. With ADHase activity the timing was 15% greater than that in a normal cell cycle whereas with CO2 production it was 15% less. We suggest that these and other periodic events are subject to independent oscillatory controls in these blocked cultures with timings that differ from each other and from the normal cycle but in the normal cycle the oscillators are all entrained by one or more events of the DNA-division cycle.

Change in the rate of CO2 production in synchronous cultures of the fission yeast Schizosaccharomyces pombe: a periodic cell cycle event that persists after the DNA-division cycle has been blocked

1986 ◽  
Vol 86 (1) ◽  
pp. 191-206
Author(s):  
B. Novak ◽  
J.M. Mitchison
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Rate Change ◽  
Co2 Production ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Normal Cycle ◽  
Periodic Cell ◽  
Linear Pattern ◽  
Synchronous Cultures ◽  
Oscillatory Pattern

CO2 production has been followed by manometry in synchronous and asynchronous cultures of Schizosaccharomyces pombe prepared by elutriation from the same initial culture. The rate of production follows a linear pattern in synchronous cultures with a rate change once per cycle at the time of cell division. This pattern is most clearly shown in oscillations of the difference between values of the second differential (acceleration) for the synchronous and asynchronous cultures. The association between the rate change and the time of division is maintained during growth speeded up in rich medium and slowed down in poor medium and at lower temperature. It is also maintained after a shift-up in temperature. Results with wee mutants suggest that the association is with the S period rather than division itself. The rate and acceleration of CO2 production are approximately proportional to cell size (protein content) in asynchronous cultures. When synchronous cultures of the temperature-sensitive mutants cdc2.33 and cdc2.33 wee1.6 are shifted up to the restrictive temperature, the DNA-division cycle is blocked. The oscillatory pattern of CO2 production, however, continues for one to two cycles until the acceleration reaches a constant value, after which the oscillations are undetectable. This point is reached later in the double mutant and there is a phase difference in the oscillations compared to those in the single mutant. With both blocked mutants the ‘free-running’ oscillations are about 15% shorter than the normal cycle time. There are well-known examples of such oscillations in eggs but they are rare in growing systems.

Oxygen uptake during the cell cycle of the fission yeast Schizosaccharomyces pombe

1978 ◽  
Vol 33 (1) ◽  
pp. 399-411
Author(s):  
J. Creanor
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Oxygen Uptake ◽  
Dna Synthesis ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Nuclear Division ◽  
Synchronous Culture ◽  
Synchronous Cultures ◽  
The Many

Oxygen uptake was measured in synchronous cultures of the fission yeast Schizosaccharomyces pombe. The rate of oxygen uptake was found to increase in a step-wise manner at the beginning of the cycle and again in the middle of the cycle. The increases in rate were such that overall, oxygen uptake doubled in rate once per cell cycle. Addition of inhibitors of DNA synthesis or nuclear division to a synchronous culture did not affect the uptake of oxygen. In an induced synchronous culture, in which DNA synthesis, cell division, and nuclear division, but not ‘growth’ were synchronized, oxygen uptake increased continuously in rate and did not show the step-wise rises which were shown in the selection-synchronized culture. These results were compared with previous measurements of oxygen uptake in yeast and an explanation is suggested for the many different patterns which have been reported.

The Effect of 2-Phenyl Ethanol on the DNA Synthesis Cycle of Schizosaccharomyces Pombe

1970 ◽  
Vol 7 (2) ◽  
pp. 523-530
Author(s):  
C. J. BOSTOCK
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Schizosaccharomyces Pombe ◽  
Normal Growth ◽  
S Phase ◽  
G1 Phase ◽  
Synchronous Cultures ◽  
Number Of Cells

The effect of different concentrations of 2-phenyl ethanol (PE) on growth and DNA synthesis of Schizosaccharomyces pombe is described. o.3% PE inhibits the entry of cells into S phase, but allows a doubling in the number of cells in the culture. The effect of o.2% PE on random and synchronous cultures of S. pombe shows that, in the continued presence of the inhibitor, the S phase is moved to a different point in the cell cycle. Cells continue to grow in the presence of o.2% PE with a G1 phase occupying a significant portion of the cell cycle. This differs from normal growth when the G1 phase is absent.

Patterns of protein synthesis during the cell cycle of the fission yeast Schizosaccharomyces pombe

1982 ◽  
Vol 58 (1) ◽  
pp. 263-285
Author(s):  
J. Creanor ◽  
J.M. Mitchison
Keyword(s):  
Cell Cycle ◽  
Protein Synthesis ◽  
Schizosaccharomyces Pombe ◽  
Messenger Rna ◽  
Model Fitting ◽  
Selection Procedure ◽  
Periodic Pattern ◽  
Synchronous Cultures ◽  
Metabolic Pool ◽  
Cdc 2

The rate of protein synthesis through the cell cycle of Schizosaccharomyces pombe has been determined from the incorporation of pulses of [3H]tryptophan in synchronous cultures prepared by selection in an elutriating rotor. This selection procedure caused minimal perturbations as judged by asynchronous control cultures, which had also been put through the rotor. The rate of synthesis showed a periodic pattern rather than a smooth exponential increase. There was a sharp increase in the rate at an ‘acceleration point’ at about 0.9 of the cycle. Model-fitting by a novel procedure suggests that the average single cell has an increasing rate of protein synthesis for the first 60% of the cycle and a constant rate for the remaining 40%. The same pattern was shown in less extensive experiments with [3H]leucine and [3H]phenylalanine. It was also shown in a series of size mutants, which indicates that the pattern is not size-related, in contrast to earlier work on the rates of synthesis of messenger RNA. However, one large mutant (cdc 2.M35r20) had a significantly earlier acceleration point. Care was taken to justify the assumption that the rate of incorporation of tryptophan was a valid measure of the rate of protein synthesis. A tryptophan auxotroph was used to eliminate the problem of endogenous supply and the size of the metabolic pool was measured through the cycle. This pool did not show cell-cycle related fluctuations. An operational model of the pools is presented.

Rates of synthesis of polyadenylated messenger RNA and ribosomal RNA during the cell cycle of Schizosaccharomyces pombe. With an appendix: calculation of the pattern of protein accumulation from observed changes in the rate of messenger RNA synthesis

1976 ◽  
Vol 21 (3) ◽  
pp. 497-521
Author(s):  
R.S. Fraser ◽  
F. Moreno
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Schizosaccharomyces Pombe ◽  
Ribosomal Rna ◽  
Messenger Rna ◽  
Rna Synthesis ◽  
Protein Accumulation ◽  
Synchronous Culture ◽  
Synchronous Cultures ◽  
Gene Copies

The rates of polyadenylated messenger RNA and ribosomal RNA synthesis were measured in synchronously dividing cultures of fission yeast (Schizosaccharomyces pombe). Control asynchronous cultures, which had been exposed to the conditions used for preparing synchronous cultures, were investigated to check for effects of the synchronization procedure itself on RNA synthesis. After each period of DNA synthesis in synchronous culture, the rates of messenger and ribosomal RNA synthesis doubled, suggesting that gene number controls the rate of messenger and ribosomal RNA synthesis. This was confirmed by experiments with asynchronous, exponential-phase cultures in which DNA synthesis was inhibited by hydroxyurea. Both synchronous culture and hydroxyurea experiments suggested that there is a delay of 15 min (0-1 of the cell generation time) between replication of the DNA and transcription of both gene copies. A pattern of protein accumulation was calculated from changes in the rate of polyadenylated messenger RNA synthesis during synchronous culture. The simulated pattern indicates that protein is accumulated linearly, with a doubling in the rate of accumulation once per cell cycle. The simulated pattern of protein accumulation is very similar to measurements previously reported by other workers of changes in activities of 3 enzymes in synchronous cultures. It is suggested that the doubling of the rate of messenger RNA synthesis, as a consequence of the replication of the DNA once per cycle, provides the basis of a mechanism for control of the doubling of other cellular constituents during the cell cycle.

Growth and Changes in Pool and Macromolecular Components of Schizosaccharomyces Pombe During the Cell Cycle

1971 ◽  
Vol 9 (3) ◽  
pp. 701-717
Author(s):  
NOWELL STEBBING
Keyword(s):  
Cell Cycle ◽  
Amino Acids ◽  
Amino Acid ◽  
Schizosaccharomyces Pombe ◽  
Minimal Medium ◽  
Linear Growth ◽  
Dry Weight ◽  
Total Cell ◽  
Complex Medium ◽  
Synchronous Cultures

Amino acids, nucleotide and carbohydrate material were found to account for 46% of the total dry weight of pool material in Schizosaccharomyces pombe growing in minimal medium. The composition of the amino acid pool was also determined by autoanalysis and was found to be unaltered during growth in 2 M sorbitol, indicating that pool amino acids are not important in the osmoregulation of the cell. Kinetic analysis of the amino acid pool using 14C-labelled amino acids showed that amino acids accumulated from the medium enter an ‘expandable’ pool distinct from the ‘internal’ pool which is maintained during growth on minimal medium. Total RNA, protein, pool amino acid and pool ‘nucleotide’ material were estimated in synchronous cultures grown in minimal medium. All these components appeared to accumulate in an exponential manner during the cell cycle. Direct estimation of total cellular dry weight and the total pool in synchronous cultures showed that total cell dry weight increased exponentially and the pool did not fluctuate during growth in minimal medium. This contrasts with previous work on single cells of S. pombe grown in complex medium which showed that the dry weight of the pool fluctuates during the cell cycle and total cell dry weight increased linearly. Linear growth of S. pombe in malt extract broth can be accounted for by the presence of the second (‘expandable’) pool of amino acids formed during growth in complex medium. The phenomenon of linear growth during the cell cycle is shown to occur generally only in cells growing in complex medium. The phenomenon is considered in relation to mechanisms for controlling the size of the pool during growth in complex media.

Protein synthesis and its relation to the DNA-division cycle in the fission yeast Schizosaccharomyces pombe

1984 ◽  
Vol 69 (1) ◽  
pp. 199-210
Author(s):  
J. Creanor ◽  
J.M. Mitchison
Keyword(s):  
Cell Cycle ◽  
Protein Synthesis ◽  
Schizosaccharomyces Pombe ◽  
Protein Content ◽  
Fission Yeast ◽  
Nuclear Division ◽  
Restrictive Temperature ◽  
Tentative Conclusion ◽  
Synchronous Cultures

The rate of protein synthesis has been measured with pulse labels of [3H]tryptophan in synchronous and asynchronous cultures of cdc mutants of Schizosaccharomyces pombe shifted up to the restrictive temperature. The cell cycle related fluctuations in rate that occur in normal synchronous cultures vanish when nuclear division is blocked in synchronous cultures of cdc2 and cdc10. But they persist in cdc11 where nuclear division continues and cleavage is stopped. We conclude that nuclear division affects the rate of synthesis and that this effect is inhibitory and probably persists for the last 40% of the cycle. When nuclear division has been blocked, the rate of synthesis continues to increase until a plateau is reached where the rate remains constant. Three size mutants of cdc2 reach the plateau at the same average protein content per cell although their initial protein contents vary over a threefold range. Comparison of these results with those from cdc10 leads to the tentative conclusion that the plateau starts when the cells reach a critical protein/DNA ratio.

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