scholarly journals Drosophila Mcm10 Interacts with Members of the Prereplication Complex and Is Required for Proper Chromosome Condensation

2003 ◽  
Vol 14 (6) ◽  
pp. 2206-2215 ◽  
Author(s):  
Tim W. Christensen ◽  
Bik K. Tye
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Drosophila Melanogaster ◽  
Dna Replication ◽  
Dna Content ◽  
Chromatin Condensation ◽  
Chromosome Condensation ◽  
Null Mutant ◽  
Aberrant Chromosome ◽  
Initiation Of Dna Replication

Mcm10 is required for the initiation of DNA replication in Saccharomyces cerevisiae. We have cloned MCM10 from Drosophila melanogaster and show that it complements a ScMCM10 null mutant. Moreover, Mcm10 interacts with key members of the prereplication complex: Mcm2, Dup (Cdt1), and Orc2. Interactions were also detected between Mcm10 and itself, Cdc45, and Hp1. RNAi depletion of Orc2 and Mcm10 in KC cells results in loss of DNA content. Furthermore, depletion of Mcm10, Cdc45, Mcm2, Mcm5, and Orc2, respectively, results in aberrant chromosome condensation. The condensation defects observed resemble previously published reports for Orc2, Orc5, and Mcm4 mutants. Our results strengthen and extend the argument that the processes of chromatin condensation and DNA replication are linked.

scholarly journals Quiescent Saccharomyces cerevisiae forms telomere hyperclusters at the nuclear membrane vicinity through a multifaceted mechanism involving Esc1, the Sir complex, and chromatin condensation

2016 ◽  
Vol 27 (12) ◽  
pp. 1875-1884 ◽  
Author(s):  
Damien Laporte ◽  
Fabien Courtout ◽  
Sylvain Tollis ◽  
Isabelle Sagot
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nuclear Membrane ◽  
Chromatin Condensation ◽  
Histone H1 ◽  
Chromosome Condensation ◽  
Histone H4 ◽  
Microtubule Bundle ◽  
Quiescent Cells ◽  
Nuclear Reorganization ◽  
Sir Complex

Like other eukaryotes, Saccharomyces cerevisiae spatially organizes its chromosomes within the nucleus. In G1 phase, the yeast’s 32 telomeres are clustered into 6–10 foci that dynamically interact with the nuclear membrane. Here we show that, when cells leave the division cycle and enter quiescence, telomeres gather into two to three hyperclusters at the nuclear membrane vicinity. This localization depends on Esc1 but not on the Ku proteins. Telomere hypercluster formation requires the Sir complex but is independent of the nuclear microtubule bundle that specifically assembles in quiescent cells. Importantly, mutants deleted for the linker histone H1 Hho1 or defective in condensin activity or affected for histone H4 Lys-16 deacetylation are impaired, at least in part, for telomere hypercluster formation in quiescence, suggesting that this process involves chromosome condensation. Finally, we establish that telomere hypercluster formation is not necessary for quiescence establishment, maintenance, and exit, raising the question of the physiological raison d’être of this nuclear reorganization.

scholarly journals Expression of p60v-src in Saccharomyces cerevisiae results in elevation of p34CDC28 kinase activity and release of the dependence of DNA replication on mitosis.

1993 ◽  
Vol 13 (8) ◽  
pp. 5112-5121 ◽  
Author(s):  
F Boschelli
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Dna Content ◽  
Protein Tyrosine Kinase ◽  
Flow Cytometric Analysis ◽  
Yeast Saccharomyces Cerevisiae ◽  
Genetic Studies ◽  
Flow Cytometric ◽  
Oncogenic Protein ◽  
Cdc28 Kinase

Expression of the oncogenic protein tyrosine kinase p60v-src in the yeast Saccharomyces cerevisiae has been shown to result in rapid cell death (J. S. Brugge, G. Jarosik, J. Andersen, A. Queral-Lustig, M. Fedor-Chaiken, and J. R. Broach, Mol. Cell. Biol. 7:2180-2187, 1987). Work described here demonstrates that v-Src expression results in accumulation of large-budded cells and a nuclear division block without blocking cytokinesis. Flow-cytometric analysis indicates that the DNA content of these cells is elevated beyond the G2 DNA content, and genetic studies indicate that v-Src expression causes aneuploidy. The activity of Cdc28 kinase, which controls the G1/S and G2/M transitions in S. cerevisiae, increases during galactose induction in a Src+ strain but not in an isogenic Src- strain. These observations indicate that v-Src expression disrupts p34CDC28 kinase regulation, allowing DNA replication to proceed in the absence of a prior mitotic event.

scholarly journals Cell Cycle Control of Cdc7p Kinase Activity through Regulation of Dbf4p Stability

1999 ◽  
Vol 19 (7) ◽  
pp. 4888-4896 ◽  
Author(s):  
Guy Oshiro ◽  
Julia C. Owens ◽  
Yiqun Shellman ◽  
Robert A. Sclafani ◽  
Joachim J. Li
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Dna Replication ◽  
Kinase Activity ◽  
Cell Cycle Control ◽  
S Phase ◽  
Regulatory Subunit ◽  
Anaphase Promoting Complex ◽  
Initiation Of Dna Replication

ABSTRACT In Saccharomyces cerevisiae, the heteromeric kinase complex Cdc7p-Dbf4p plays a pivotal role at replication origins in triggering the initiation of DNA replication during the S phase. We have assayed the kinase activity of endogenous levels of Cdc7p kinase by using a likely physiological target, Mcm2p, as a substrate. Using this assay, we have confirmed that Cdc7p kinase activity fluctuates during the cell cycle; it is low in the G1 phase, rises as cells enter the S phase, and remains high until cells complete mitosis. These changes in kinase activity cannot be accounted for by changes in the levels of the catalytic subunit Cdc7p, as these levels are constant during the cell cycle. However, the fluctuations in kinase activity do correlate with levels of the regulatory subunit Dbf4p. The regulation of Dbf4p levels can be attributed in part to increased degradation of the protein in G1 cells. This G1-phase instability is cdc16 dependent, suggesting a role of the anaphase-promoting complex in the turnover of Dbf4p. Overexpression of Dbf4p in the G1 phase can partially overcome this elevated turnover and lead to an increase in Cdc7p kinase activity. Thus, the regulation of Dbf4p levels through the control of Dbf4p degradation has an important role in the regulation of Cdc7p kinase activity during the cell cycle.

scholarly journals Gid8p (Dcr1p) and Dcr2p Function in a Common Pathway To Promote START Completion in Saccharomyces cerevisiae

2004 ◽  
Vol 3 (6) ◽  
pp. 1627-1638 ◽  
Author(s):  
Ritu Pathak ◽  
Lydia M. Bogomolnaya ◽  
Jinbai Guo ◽  
Michael Polymenis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Dna Replication ◽  
Cell Cycle Progression ◽  
Cycle Progression ◽  
Dependent Cell ◽  
Promote Cell Cycle Progression ◽  
Initiation Of Dna Replication ◽  
Regulator Genes ◽  
Timing Of Initiation

ABSTRACT How cells determine when to initiate DNA replication is poorly understood. Here we report that in Saccharomyces cerevisiae overexpression of the dosage-dependent cell cycle regulator genes DCR2 (YLR361C) and GID8 (DCR1/YMR135C) accelerates initiation of DNA replication. Cells lacking both GID8 and DCR2 delay initiation of DNA replication. Genetic analysis suggests that Gid8p functions upstream of Dcr2p to promote cell cycle progression. DCR2 is predicted to encode a gene product with phosphoesterase activity. Consistent with these predictions, a DCR2 allele carrying a His338 point mutation, which in known protein phosphatases prevents catalysis but allows substrate binding, antagonized the function of the wild-type DCR2 allele. Finally, we report genetic interactions involving GID8, DCR2, and CLN3 (which encodes a G1 cyclin) or SWI4 (which encodes a transcription factor of the G1/S transcription program). Our findings identify two gene products with a probable regulatory role in the timing of initiation of cell division.

scholarly journals A quantitative model of the initiation of DNA replication in Saccharomyces cerevisiae predicts the effects of system perturbations

2012 ◽  
Vol 6 (1) ◽  
pp. 78 ◽  
Author(s):  
Rohan D Gidvani ◽  
Peter Sudmant ◽  
Grace Li ◽  
Lance F DaSilva ◽  
Brendan J McConkey ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Quantitative Model ◽  
Initiation Of Dna Replication

scholarly journals Interphase Cytogenetic Analysis of Micronucleated and Multinucleated Cells Supports the Premature Chromosome Condensation Hypothesis as the Mechanistic Origin of Chromothripsis

Cancers ◽  
2019 ◽  
Vol 11 (8) ◽  
pp. 1123 ◽  
Author(s):  
Pantelias ◽  
Karachristou ◽  
Georgakilas ◽  
Terzoudi
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Cytogenetic Analysis ◽  
Alternative Hypothesis ◽  
Chromatin Condensation ◽  
Human Lymphocytes ◽  
Chinese Hamster ◽  
Chromosome Condensation ◽  
Premature Chromosome Condensation ◽  
Multinucleated Cells

The discovery of chromothripsis in cancer genomes challenges the long-standing concept of carcinogenesis as the result of progressive genetic events. Despite recent advances in describing chromothripsis, its mechanistic origin remains elusive. The prevailing conception is that it arises from a massive accumulation of fragmented DNA inside micronuclei (MN), whose defective nuclear envelope ruptures or leads to aberrant DNA replication, before main nuclei enter mitosis. An alternative hypothesis is that the premature chromosome condensation (PCC) dynamics in asynchronous micronucleated cells underlie chromosome shattering in a single catastrophic event, a hallmark of chromothripsis. Specifically, when main nuclei enter mitosis, premature chromatin condensation provokes the shattering of chromosomes entrapped inside MN, if they are still undergoing DNA replication. To test this hypothesis, the agent RO-3306, a selective ATP-competitive inhibitor of CDK1 that promotes cell cycle arrest at the G2/M boundary, was used in this study to control the degree of cell cycle asynchrony between main nuclei and MN. By delaying the entrance of main nuclei into mitosis, additional time was allowed for the completion of DNA replication and duplication of chromosomes inside MN. We performed interphase cytogenetic analysis using asynchronous micronucleated cells generated by exposure of human lymphocytes to γ-rays, and heterophasic multinucleated Chinese hamster ovary (CHO) cells generated by cell fusion procedures. Our results demonstrate that the PCC dynamics during asynchronous mitosis in micronucleated or multinucleated cells are an important determinant of chromosome shattering and may underlie the mechanistic origin of chromothripsis.

scholarly journals The mcm5-bob1 Bypass of Cdc7p/Dbf4p in DNA Replication Depends on Both Cdk1-Independent and Cdk1-Dependent Steps in Saccharomyces cerevisiae

Genetics ◽  
2002 ◽  
Vol 161 (1) ◽  
pp. 47-57 ◽  
Author(s):  
Robert A Sclafani ◽  
Marianne Tecklenburg ◽  
Angela Pierce
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Protein Kinases ◽  
Molecular Analysis ◽  
Molecular Conformation ◽  
Ectopic Expression ◽  
The Other ◽  
G1 Phase ◽  
Combined Action ◽  
Initiation Of Dna Replication

Abstract The roles in DNA replication of two distinct protein kinases, Cdc7p/Dbf4p and Cdk1p/Clb (B-type cyclin), were studied. This was accomplished through a genetic and molecular analysis of the mechanism by which the mcm5-bob1 mutation bypasses the function of the Cdc7p/Dbf4p kinase. Genetic experiments revealed that loss of either Clb5p or Clb2p cyclins suppresses the mcm5-bob1 mutation and prevents bypass. These two cyclins have distinct roles in bypass and presumably in DNA replication as overexpression of one could not complement the loss of the other. Furthermore, the ectopic expression of CLB2 in G1 phase cannot substitute for CLB5 function in bypass of Cdc7p/Dbf4p by mcm5-bob1. Molecular experiments revealed that the mcm5-bob1 mutation allows for constitutive loading of Cdc45p at early origins in arrested G1 phase cells when both kinases are inactive. A model is proposed in which the Mcm5-bob1 protein assumes a unique molecular conformation without prior action by either kinase. This conformation allows for stable binding of Cdc45p to the origin. However, DNA replication still cannot occur without the combined action of Cdk1p/Clb5p and Cdk1p/Clb2p. Thus Cdc7p and Cdk1p kinases catalyze the initiation of DNA replication at several distinct steps, of which only a subset is bypassed by the mcm5-bob1 mutation.

Expression of p60v-src in Saccharomyces cerevisiae results in elevation of p34CDC28 kinase activity and release of the dependence of DNA replication on mitosis

1993 ◽  
Vol 13 (8) ◽  
pp. 5112-5121
Author(s):  
F Boschelli
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Dna Content ◽  
Protein Tyrosine Kinase ◽  
Flow Cytometric Analysis ◽  
Yeast Saccharomyces Cerevisiae ◽  
Genetic Studies ◽  
Flow Cytometric ◽  
Oncogenic Protein ◽  
Cdc28 Kinase

Expression of the oncogenic protein tyrosine kinase p60v-src in the yeast Saccharomyces cerevisiae has been shown to result in rapid cell death (J. S. Brugge, G. Jarosik, J. Andersen, A. Queral-Lustig, M. Fedor-Chaiken, and J. R. Broach, Mol. Cell. Biol. 7:2180-2187, 1987). Work described here demonstrates that v-Src expression results in accumulation of large-budded cells and a nuclear division block without blocking cytokinesis. Flow-cytometric analysis indicates that the DNA content of these cells is elevated beyond the G2 DNA content, and genetic studies indicate that v-Src expression causes aneuploidy. The activity of Cdc28 kinase, which controls the G1/S and G2/M transitions in S. cerevisiae, increases during galactose induction in a Src+ strain but not in an isogenic Src- strain. These observations indicate that v-Src expression disrupts p34CDC28 kinase regulation, allowing DNA replication to proceed in the absence of a prior mitotic event.

scholarly journals Cell Cycle Requirements in Assembling Silent Chromatin in Saccharomyces cerevisiae

2006 ◽  
Vol 26 (3) ◽  
pp. 852-862 ◽  
Author(s):  
Ann L. Kirchmaier ◽  
Jasper Rine
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Dna Replication ◽  
Replication Fork ◽  
S Phase ◽  
Silent Chromatin ◽  
Type Locus ◽  
Sir Proteins ◽  
Initiation Of Dna Replication ◽  
Pass Through

ABSTRACT The establishment of silencing at the silent mating-type locus, HMR, in Saccharomyces cerevisiae requires that yeast pass through S phase of the cell cycle, yet requires neither the initiation of DNA replication at the locus destined to become silenced nor the passage of a replication fork through that locus. We tested whether this S-phase requirement reflects a window within the cell cycle permissive for recruitment of Sir proteins to HMR. The S-phase-restricted event necessary for silencing occurred after recruitment of Sir proteins to HMR. Moreover, cells arrested in early S phase formed silent chromatin at HMR, provided HMR was on a nonreplicating template. Replicating templates required a later step for silencing. These results provide temporal resolution of discrete steps in the formation of silent chromatin and suggest that more than one cell cycle-regulated event may be necessary for the establishment of silencing.

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