scholarly journals Separase cooperates with Zds1 and Zds2 to activate Cdc14 phosphatase in early anaphase

2008 ◽  
Vol 182 (5) ◽  
pp. 873-883 ◽  
Author(s):  
Ethel Queralt ◽  
Frank Uhlmann
Keyword(s):  
Cell Cycle ◽  
Budding Yeast ◽  
Mitotic Exit ◽  
Interacting Proteins ◽  
Cyclin Dependent Kinase ◽  
Down Regulation ◽  
Anaphase Onset ◽  
Early Anaphase ◽  
Key Enzyme ◽  
Mitotic Cyclin

Completion of mitotic exit and cytokinesis requires the inactivation of mitotic cyclin-dependent kinase (Cdk) activity. A key enzyme that counteracts Cdk during budding yeast mitotic exit is the Cdc14 phosphatase. Cdc14 is inactive for much of the cell cycle, sequestered by its inhibitor Net1 in the nucleolus. At anaphase onset, separase-dependent down-regulation of PP2ACdc55 allows phosphorylation of Net1 and consequent Cdc14 release. How separase causes PP2ACdc55 down-regulation is not known. Here, we show that two Cdc55-interacting proteins, Zds1 and Zds2, contribute to timely Cdc14 activation during mitotic exit. Zds1 and Zds2 are required downstream of separase to facilitate nucleolar Cdc14 release. Ectopic Zds1 expression in turn is sufficient to down-regulate PP2ACdc55 and promote Net1 phosphorylation. These findings identify Zds1 and Zds2 as new components of the mitotic exit machinery, involved in activation of the Cdc14 phosphatase at anaphase onset. Our results suggest that these proteins may act as separase-regulated PP2ACdc55 inhibitors.

scholarly journals DNA Damage Checkpoints Inhibit Mitotic Exit by Two Different Mechanisms

2007 ◽  
Vol 27 (14) ◽  
pp. 5067-5078 ◽  
Author(s):  
Fengshan Liang ◽  
Yanchang Wang
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Cell Cycle Progression ◽  
Budding Yeast ◽  
Mitotic Exit ◽  
Yeast Cells ◽  
Cyclin Dependent Kinase ◽  
Cycle Progression ◽  
Dna Damage Checkpoints ◽  
Early Anaphase

ABSTRACT Cyclin-dependent kinase (CDK) governs cell cycle progression, and its kinase activity fluctuates during the cell cycle. Mitotic exit pathways are responsible for the inactivation of CDK after chromosome segregation by promoting the release of a nucleolus-sequestered phosphatase, Cdc14, which antagonizes CDK. In the budding yeast Saccharomyces cerevisiae, mitotic exit is controlled by the FEAR (for “Cdc-fourteen early anaphase release”) and mitotic exit network (MEN) pathways. In response to DNA damage, two branches of the DNA damage checkpoint, Chk1 and Rad53, are activated in budding yeast to prevent anaphase entry and mitotic exit, allowing cells more time to repair damaged DNA. Here we present evidence indicating that yeast cells negatively regulate mitotic exit through two distinct pathways in response to DNA damage. Rad53 prevents mitotic exit by inhibiting the MEN pathway, whereas the Chk1 pathway prevents FEAR pathway-dependent Cdc14 release in the presence of DNA damage. In contrast to previous data, the Rad53 pathway negatively regulates MEN independently of Cdc5, a Polo-like kinase essential for mitotic exit. Instead, a defective Rad53 pathway alleviates the inhibition of MEN by Bfa1.

scholarly journals Budding yeast relies on G1 cyclin specificity to couple cell cycle progression with morphogenetic development

2021 ◽  
Vol 7 (23) ◽  
pp. eabg0007
Author(s):  
Deniz Pirincci Ercan ◽  
Florine Chrétien ◽  
Probir Chakravarty ◽  
Helen R. Flynn ◽  
Ambrosius P. Snijders ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Budding Yeast ◽  
Quantitative Model ◽  
Cyclin Dependent Kinase ◽  
Cycle Progression ◽  
Model Cell ◽  
Mitotic Cyclin ◽  
Substrate Phosphorylation ◽  
The Cost

Two models have been put forward for cyclin-dependent kinase (Cdk) control of the cell cycle. In the qualitative model, cell cycle events are ordered by distinct substrate specificities of successive cyclin waves. Alternatively, in the quantitative model, the gradual rise of Cdk activity from G1 phase to mitosis leads to ordered substrate phosphorylation at sequential thresholds. Here, we study the relative contributions of qualitative and quantitative Cdk control in Saccharomyces cerevisiae. All S phase and mitotic cyclins can be replaced by a single mitotic cyclin, albeit at the cost of reduced fitness. A single cyclin can also replace all G1 cyclins to support ordered cell cycle progression, fulfilling key predictions of the quantitative model. However, single-cyclin cells fail to polarize or grow buds and thus cannot survive. Our results suggest that budding yeast has become dependent on G1 cyclin specificity to couple cell cycle progression to essential morphogenetic events.

scholarly journals Genetic and Biochemical Evaluation of the Importance of Cdc6 in Regulating Mitotic Exit

2003 ◽  
Vol 14 (11) ◽  
pp. 4592-4604 ◽  
Author(s):  
Vincent Archambault ◽  
Caihong X. Li ◽  
Alan J. Tackett ◽  
Ralph Wäsch ◽  
Brian T. Chait ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Mitotic Exit ◽  
Terminal Region ◽  
Cyclin Dependent Kinase ◽  
Replication Control ◽  
Mitotic Cyclin ◽  
Biochemical Evaluation ◽  
Inhibitory Domain ◽  
Late Mitosis ◽  
Control Protein

We evaluated the hypothesis that the N-terminal region of the replication control protein Cdc6 acts as an inhibitor of cyclin-dependent kinase (Cdk) activity, promoting mitotic exit. Cdc6 accumulation is restricted to the period from mid-cell cycle until the succeeding G1, due to proteolytic control that requires the Cdc6 N-terminal region. During late mitosis, Cdc6 is present at levels comparable with Sic1 and binds specifically to the mitotic cyclin Clb2. Moderate overexpression of Cdc6 promotes viability of CLB2Δdb strains, which otherwise arrest at mitotic exit, and rescue is dependent on the N-terminal putative Cdk-inhibitory domain. These observations support the potential for Cdc6 to inhibit Clb2-Cdk, thus promoting mitotic exit. Consistent with this idea, we observed a cytokinesis defect in cdh1Δ sic1Δ cdc6Δ2–49 triple mutants. However, we were able to construct viable strains, in three different backgrounds, containing neither SIC1 nor the Cdc6 Cdk-inhibitory domain, in contradiction to previous work. We conclude, therefore, that although both Cdc6 and Sic1 have the potential to facilitate mitotic exit by inhibiting Clb2-Cdk, mitotic exit nevertheless does not require any identified stoichiometric inhibitor of Cdk activity.

scholarly journals Intrinsic and Cyclin-dependent Kinase-dependent Control of Spindle Pole Body Duplication in Budding Yeast

2008 ◽  
Vol 19 (8) ◽  
pp. 3243-3253 ◽  
Author(s):  
Laura A. Simmons Kovacs ◽  
Christine L. Nelson ◽  
Steven B. Haase
Keyword(s):  
Cell Cycle ◽  
Budding Yeast ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Cyclin Dependent Kinase ◽  
Multipolar Spindle ◽  
Cyclin Dependent Kinases ◽  
Pole Body ◽  
Intrinsic Mechanism ◽  
Mitotic Cyclin

Centrosome duplication must be tightly controlled so that duplication occurs only once each cell cycle. Accumulation of multiple centrosomes can result in the assembly of a multipolar spindle and lead to chromosome mis-segregation and genomic instability. In metazoans, a centrosome-intrinsic mechanism prevents reduplication until centriole disengagement. Mitotic cyclin/cyclin-dependent kinases (CDKs) prevent reduplication of the budding yeast centrosome, called a spindle pole body (SPB), in late S-phase and G2/M, but the mechanism remains unclear. How SPB reduplication is prevented early in the cell cycle is also not understood. Here we show that, similar to metazoans, an SPB-intrinsic mechanism prevents reduplication early in the cell cycle. We also show that mitotic cyclins can inhibit SPB duplication when expressed before satellite assembly in early G1, but not later in G1, after the satellite had assembled. Moreover, electron microscopy revealed that SPBs do not assemble a satellite in cells expressing Clb2 in early G1. Finally, we demonstrate that Clb2 must localize to the cytoplasm in order to inhibit SPB duplication, suggesting the possibility for direct CDK inhibition of satellite components. These two mechanisms, intrinsic and extrinsic control by CDK, evoke two-step system that prevents SPB reduplication throughout the cell cycle.

scholarly journals Mitotic Exit in the Absence of Separase Activity

2009 ◽  
Vol 20 (5) ◽  
pp. 1576-1591 ◽  
Author(s):  
Ying Lu ◽  
Frederick Cross
Keyword(s):  
Nuclear Export ◽  
Spindle Pole Body ◽  
Quantitative Measure ◽  
Spindle Pole ◽  
Mitotic Exit ◽  
Cyclin Dependent Kinase ◽  
Sister Chromatids ◽  
Early Anaphase ◽  
Mitotic Cyclin ◽  
A Minor

In budding yeast, three interdigitated pathways regulate mitotic exit (ME): mitotic cyclin–cyclin-dependent kinase (Cdk) inactivation; the Cdc14 early anaphase release (FEAR) network, including a nonproteolytic function of separase (Esp1); and the mitotic exit network (MEN) driven by interaction between the spindle pole body and the bud cortex. Here, we evaluate the contributions of these pathways to ME kinetics. Reducing Cdk activity is critical for ME, and the MEN contributes strongly to ME efficiency. Esp1 contributes to ME kinetics mainly through cohesin cleavage: the Esp1 requirement can be largely bypassed if cells are provided Esp1-independent means of separating sister chromatids. In the absence of Esp1 activity, we observed only a minor ME delay consistent with a FEAR defect. Esp1 overexpression drives ME in Cdc20-depleted cells arrested in metaphase. We have found that this activity of overexpressed Esp1 depended on spindle integrity and the MEN. We defined the first quantitative measure for Cdc14 release based on colocalization with the Net1 nucleolar anchor. This measure indicates efficient Cdc14 release upon MEN activation; release driven by Esp1 in the absence of microtubules was inefficient and incapable of driving ME. We also found a novel role for the MEN: activating Cdc14 nuclear export, even in the absence of Net1.

scholarly journals Oscillations in Cdc14 release and sequestration reveal a circuit underlying mitotic exit

2010 ◽  
Vol 190 (2) ◽  
pp. 209-222 ◽  
Author(s):  
Romilde Manzoni ◽  
Francesca Montani ◽  
Clara Visintin ◽  
Fabrice Caudron ◽  
Andrea Ciliberto ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Feedback Loop ◽  
Mitotic Exit ◽  
Centrosome Duplication ◽  
Negative Feedback Loop ◽  
Cyclin Dependent Kinase ◽  
Bud Formation ◽  
Mitotic Exit Network ◽  
Early Anaphase ◽  
Periodic Cycles

In budding yeast, the phosphatase Cdc14 orchestrates progress through anaphase and mitotic exit, thereby resetting the cell cycle for a new round of cell division. Two consecutive pathways, Cdc fourteen early anaphase release (FEAR) and mitotic exit network (MEN), contribute to the progressive activation of Cdc14 by regulating its release from the nucleolus, where it is kept inactive by Cfi1. In this study, we show that Cdc14 activation requires the polo-like kinase Cdc5 together with either Clb–cyclin-dependent kinase (Cdk) or the MEN kinase Dbf2. Once active, Cdc14 triggers a negative feedback loop that, in the presence of stable levels of mitotic cyclins, generates periodic cycles of Cdc14 release and sequestration. Similar phenotypes have been described for yeast bud formation and centrosome duplication. A common theme emerges where events that must happen only once per cycle, although intrinsically capable of oscillations, are limited to one occurrence by the cyclin–Cdk cell cycle engine.

scholarly journals The Molecular Function of the Yeast Polo-like Kinase Cdc5 in Cdc14 Release during Early Anaphase

2009 ◽  
Vol 20 (16) ◽  
pp. 3671-3679 ◽  
Author(s):  
Fengshan Liang ◽  
Fengzhi Jin ◽  
Hong Liu ◽  
Yanchang Wang
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Phosphatase ◽  
Budding Yeast ◽  
Molecular Function ◽  
Cyclin Dependent Kinase ◽  
Yeast Saccharomyces Cerevisiae ◽  
Protein Levels ◽  
Early Anaphase ◽  
Show Evidence ◽  
Mitotic Cyclin

In the budding yeast Saccharomyces cerevisiae , Cdc14 is sequestered within the nucleolus before anaphase entry through its association with Net1/Cfi1, a nucleolar protein. Protein phosphatase PP2ACdc55 dephosphorylates Net1 and keeps it as a hypophosphorylated form before anaphase. Activation of the Cdc fourteen early anaphase release (FEAR) pathway after anaphase entry induces a brief Cdc14 release from the nucleolus. Some of the components in the FEAR pathway, including Esp1, Slk19, and Spo12, inactivate PP2ACdc55, allowing the phosphorylation of Net1 by mitotic cyclin-dependent kinase (Cdk) (Clb2-Cdk1). However, the function of another FEAR component, the Polo-like kinase Cdc5, remains elusive. Here, we show evidence indicating that Cdc5 promotes Cdc14 release primarily by stimulating the degradation of Swe1, the inhibitory kinase for mitotic Cdk. First, we found that deletion of SWE1 partially suppresses the FEAR defects in cdc5 mutants. In contrast, high levels of Swe1 impair FEAR activation. We also demonstrated that the accumulation of Swe1 in cdc5 mutants is responsible for the decreased Net1 phosphorylation. Therefore, we conclude that the down-regulation of Swe1 protein levels by Cdc5 promotes FEAR activation by relieving the inhibition on Clb2-Cdk1, the kinase for Net1 protein.

scholarly journals Disappearance of the budding yeast Bub2–Bfa1 complex from the mother-bound spindle pole contributes to mitotic exit

2006 ◽  
Vol 172 (3) ◽  
pp. 335-346 ◽  
Author(s):  
Roberta Fraschini ◽  
Claudio D'Ambrosio ◽  
Marianna Venturetti ◽  
Giovanna Lucchini ◽  
Simonetta Piatti
Keyword(s):  
Cell Cycle ◽  
Budding Yeast ◽  
Spindle Pole ◽  
Mitotic Exit ◽  
Gtpase Activating Protein ◽  
Septin Ring ◽  
Anaphase Onset ◽  
Spindle Pole Bodies ◽  
Bud Neck ◽  
Spindle Position

Budding yeast spindle position checkpoint is engaged by misoriented spindles and prevents mitotic exit by inhibiting the G protein Tem1 through the GTPase-activating protein (GAP) Bub2/Bfa1. Bub2 and Bfa1 are found on both duplicated spindle pole bodies until anaphase onset, when they disappear from the mother-bound spindle pole under unperturbed conditions. In contrast, when spindles are misoriented they remain symmetrically localized at both SPBs. Thus, symmetric localization of Bub2/Bfa1 might lead to inhibition of Tem1, which is also present at SPBs. Consistent with this hypothesis, we show that a Bub2 version symmetrically localized on both SPBs throughout the cell cycle prevents mitotic exit in mutant backgrounds that partially impair it. This effect is Bfa1 dependent and can be suppressed by high Tem1 levels. Bub2 removal from the mother-bound SPB requires its GAP activity, which in contrast appears to be dispensable for Tem1 inhibition. Moreover, it correlates with the passage of one spindle pole through the bud neck because it needs septin ring formation and bud neck kinases.

scholarly journals NAP1 acts with Clb1 to perform mitotic functions and to suppress polar bud growth in budding yeast.

1995 ◽  
Vol 130 (3) ◽  
pp. 675-685 ◽  
Author(s):  
D R Kellogg ◽  
A W Murray
Keyword(s):  
Cell Cycle ◽  
Kinase Activity ◽  
Budding Yeast ◽  
Full Range ◽  
Normal Function ◽  
Cyclin Dependent Kinase ◽  
Mitotic Cyclin ◽  
Range Of Functions ◽  
Bud Growth ◽  
Prolonged Delay

NAP1 is a 60-kD protein that interacts specifically with mitotic cyclins in budding yeast and frogs. We have examined the ability of the yeast mitotic cyclin Clb2 to function in cells that lack NAP1. Our results demonstrate that Clb2 is unable to carry out its full range of functions without NAP1, even though Clb2/p34CDC28-associated kinase activity rises to normal levels. In the absence of NAP1, Clb2 is unable to efficiently induce mitotic events, and cells undergo a prolonged delay at the short spindle stage with normal levels of Clb2/p34CDC28 kinase activity. NAP1 is also required for the ability of Clb2 to induce the switch from polar to isotropic bud growth. As a result, polar bud growth continues during mitosis, giving rise to highly elongated cells. Our experiments also suggest that NAP1 is required for the ability of the Clb2/p34CDC28 kinase complex to amplify its own production, and that NAP1 plays a role in regulation of microtubule dynamics during mitosis. Together, these results demonstrate that NAP1 is required for the normal function of the activated Clb2/p34CDC28 kinase complex, and provide a step towards understanding how cyclin-dependent kinase complexes induce specific events during the cell cycle.

scholarly journals Far3 and Five Interacting Proteins Prevent Premature Recovery from Pheromone Arrest in the Budding Yeast Saccharomyces cerevisiae

2003 ◽  
Vol 23 (5) ◽  
pp. 1750-1763 ◽  
Author(s):  
Hilary A. Kemp ◽  
George F. Sprague,
Keyword(s):  
Cell Cycle ◽  
Budding Yeast ◽  
Open Reading Frames ◽  
Velocity Sedimentation ◽  
Interacting Proteins ◽  
Yeast Saccharomyces Cerevisiae ◽  
Cycle Arrest ◽  
Mutant Cells ◽  
Two Hybrid ◽  
Reading Frames

ABSTRACT In budding yeast, diffusible mating pheromones initiate a signaling pathway that culminates in several responses, including cell cycle arrest. Only a handful of genes required for the interface between pheromone response and the cell cycle have been identified, among them FAR1 and FAR3; of these, only FAR1 has been extensively characterized. In an effort to learn about the mechanism by which Far3 acts, we used the two-hybrid method to identify interacting proteins. We identified five previously uncharacterized open reading frames, dubbed FAR7, FAR8, FAR9, FAR10, and FAR11, that cause a far3-like pheromone arrest defect when disrupted. Using two-hybrid and coimmunoprecipitation analysis, we found that all six Far proteins interact with each other. Moreover, velocity sedimentation experiments suggest that Far3 and Far7 to Far11 form a complex. The phenotype of a sextuple far3far7-far11 mutant is no more severe than any single mutant. Thus, FAR3 and FAR7 to FAR11 all participate in the same pathway leading to G1 arrest. These mutants initially arrest in response to pheromone but resume budding after 10 h. Under these conditions, wild-type cells fail to resume budding even after several days whereas far1 mutant cells resume budding within 1 h. We conclude that the FAR3-dependent arrest pathway is functionally distinct from that which employs FAR1.

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