scholarly journals Punctuated cyclin synthesis drives early embryonic cell cycle oscillations

2012 ◽  
Vol 23 (2) ◽  
pp. 284-296 ◽  
Author(s):  
Qing Kang ◽  
Joseph R. Pomerening
Keyword(s):  
Feedback Loop ◽  
Proteasome Inhibition ◽  
Cyclin B1 ◽  
Embryonic Cell ◽  
Cyclin B ◽  
Synthesis Rate ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Effective Removal ◽  
Embryonic Cell Cycle

Cyclin B activates cyclin-dependent kinase 1 (CDK1) at mitosis, but conflicting views have emerged on the dynamics of its synthesis during embryonic cycles, ranging from continuous translation to rapid synthesis during mitosis. Here we show that a CDK1-mediated negative-feedback loop attenuates cyclin production before mitosis. Cyclin B plateaus before peak CDK1 activation, and proteasome inhibition caused minimal accumulation during mitosis. Inhibiting CDK1 permitted continual cyclin B synthesis, whereas adding nondegradable cyclin stalled it. Cycloheximide treatment before mitosis affected neither cyclin levels nor mitotic entry, corroborating this repression. Attenuated cyclin production collaborates with its destruction, since excess cyclin B1 mRNA accelerated cyclin synthesis and caused incomplete proteolysis and mitotic arrest. This repression involved neither adenylation nor the 3′ untranslated region, but it corresponded with a shift in cyclin B1 mRNA from polysome to nonpolysome fractions. A pulse-driven CDK1–anaphase-promoting complex (APC) model corroborated these results, revealing reduced cyclin levels during an oscillation and permitting more effective removal. This design also increased the robustness of the oscillator, with lessened sensitivity to changes in cyclin synthesis rate. Taken together, the results of this study underscore that attenuating cyclin synthesis late in interphase improves both the efficiency and robustness of the CDK1-APC oscillator.

The cytoskeleton-dependent localization of cdc2/cyclin B in blastomere cortex duringXenopus embryonic cell cycle

2005 ◽  
Vol 72 (3) ◽  
pp. 336-345 ◽  
Author(s):  
Norihiko Nakamura ◽  
Toshinobu Tokumoto ◽  
Shuichi Ueno ◽  
Yasuhiro Iwao
Keyword(s):  
Cell Cycle ◽  
Embryonic Cell ◽  
Cyclin B ◽  
Embryonic Cell Cycle

Cyclin B mRNA depletion only transiently inhibits the Xenopus embryonic cell cycle

Development ◽  
1991 ◽  
Vol 111 (4) ◽  
pp. 1173-1178 ◽  
Author(s):  
D.L. Weeks ◽  
J.A. Walder ◽  
J.M. Dagle
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Antisense Oligonucleotides ◽  
Normal Development ◽  
Embryonic Cell ◽  
Cyclin B ◽  
Xenopus Embryos ◽  
Embryonic Cleavage ◽  
Embryonic Cell Cycle

The control of the cell cycle is dependent on the ability to synthesize and degrade proteins called cyclins. When antisense oligonucleotides are used to deplete Xenopus embryos of mRNA encoding cyclin B protein, embryonic cleavage is inhibited. Surprisingly, after missing several rounds of cleavage, the cell cycle and cell division resumes. These studies indicate that the early embryonic cell cycle can proceed with undetectable levels of cyclin B encoding mRNA. In contrast, other events of normal development, including the activation of embryonic transcription and gastrulation, are inhibited.

scholarly journals Characterization of Vertebrate Cohesin Complexes and Their Regulation in Prophase

2000 ◽  
Vol 151 (4) ◽  
pp. 749-762 ◽  
Author(s):  
Izabela Sumara ◽  
Elisabeth Vorlaufer ◽  
Christian Gieffers ◽  
Beate H. Peters ◽  
Jan-Michael Peters
Keyword(s):  
Budding Yeast ◽  
Human Cells ◽  
Cyclin B ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Sister Chromatids ◽  
A Subunit ◽  
Mitotic Regulation

In eukaryotes, sister chromatids remain connected from the time of their synthesis until they are separated in anaphase. This cohesion depends on a complex of proteins called cohesins. In budding yeast, the anaphase-promoting complex (APC) pathway initiates anaphase by removing cohesins from chromosomes. In vertebrates, cohesins dissociate from chromosomes already in prophase. To study their mitotic regulation we have purified two 14S cohesin complexes from human cells. Both complexes contain SMC1, SMC3, SCC1, and either one of the yeast Scc3p orthologs SA1 and SA2. SA1 is also a subunit of 14S cohesin in Xenopus. These complexes interact with PDS5, a protein whose fungal orthologs have been implicated in chromosome cohesion, condensation, and recombination. The bulk of SA1- and SA2-containing complexes and PDS5 are chromatin-associated until they become soluble from prophase to telophase. Reconstitution of this process in mitotic Xenopus extracts shows that cohesin dissociation does neither depend on cyclin B proteolysis nor on the presence of the APC. Cohesins can also dissociate from chromatin in the absence of cyclin-dependent kinase 1 activity. These results suggest that vertebrate cohesins are regulated by a novel prophase pathway which is distinct from the APC pathway that controls cohesins in yeast.

The Ubiquitin-Ligase APC-Cdh1 Is Required for Maintaining Genomic Stability and Is Transcriptional Repressed by AML1/ETO.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1609-1609
Author(s):  
Ralph Waesch ◽  
Dirk Engelbert ◽  
Dominik Schnerch
Keyword(s):  
Ubiquitin Ligase ◽  
Transcriptional Repression ◽  
Genetic Instability ◽  
Kinase Inhibitor ◽  
Cell Cycle Control ◽  
Genomic Stability ◽  
Cyclin B1 ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Cyclin Dependent Kinase Inhibitor

Abstract Accurate DNA replication and chromosome segregation is essential during cell division in order to provide genomic stability and avoid malignant growth. We found that ubiquitin-dependent proteolytic cell cycle control by the E3-ubiquitin-ligase anaphase-promoting complex (APC) activated by Cdh1 (APC-Cdh1) is required for maintaining genomic integrity and viability in proliferating human cells. Lentiviral-delivered stable expression of short hairpins targeted against Cdh1 causes an apoptotic phenotype associated with p53-stabilization and p53-dependent transcriptional up-regulation of the cyclin-dependent kinase inhibitor p21. Cdh1-depleted cells enter mitosis delayed compared to controls suggesting the activation of a DNA-damage checkpoint in S- and G2-phase. Depletion of Cdh1 leads to premature accumulation of cyclin A2 and cyclin B1 in G1- and S-phase. This may interfere with loading of pre-replication-complexes onto origins of replication in G1 and cause chromosomal instability when cells progress into mitosis. In addition, stabilization of the Cdh1 target Aurora A at physiological levels is sufficient to cause centrosome overduplication and subsequent polyploidization in Cdh1 and p53 deficient cells. Genetic instability is a hallmark of cancer cells and deregulation of APC-Cdh1 may be involved, since we observe downregulation of Cdh1 in t(8;21) acute myeloid leukemia. Moreover, expression of AML1/ETO in an inducible system leads to downregulation of Cdh1 indicating transcriptional repression of Cdh1 by AML1/ETO. Consistent with these data AML1/ETO positive cell lines show numerical chromosomal aberrations. Thus, disruption of the central mitotic control machinery leads to genetic instability by several mechanisms and the APC may be an important suppressor of tumor progression in AML1/ETO positive leukemia.

scholarly journals Mitosis persists in the absence of Cdk1 activity when proteolysis or protein phosphatase activity is suppressed

2007 ◽  
Vol 179 (4) ◽  
pp. 671-685 ◽  
Author(s):  
Dimitrios A. Skoufias ◽  
Rose-Laure Indorato ◽  
Françoise Lacroix ◽  
Andreas Panopoulos ◽  
Robert L. Margolis
Keyword(s):  
Phosphatase Activity ◽  
Protein Phosphatase ◽  
Kinase Activity ◽  
Proteasome Inhibitors ◽  
Cyclin B1 ◽  
Mitotic Arrest ◽  
Mitotic Exit ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Specific Degradation

Cellular transition to anaphase and mitotic exit has been linked to the loss of cyclin-dependent kinase 1 (Cdk1) kinase activity as a result of anaphase-promoting complex/cyclosome (APC/C)–dependent specific degradation of its cyclin B1 subunit. Cdk1 inhibition by roscovitine is known to induce premature mitotic exit, whereas inhibition of the APC/C-dependent degradation of cyclin B1 by MG132 induces mitotic arrest. In this study, we find that combining both drugs causes prolonged mitotic arrest in the absence of Cdk1 activity. Different Cdk1 and proteasome inhibitors produce similar results, indicating that the effect is not drug specific. We verify mitotic status by the retention of mitosis-specific markers and Cdk1 phosphorylation substrates, although cells can undergo late mitotic furrowing while still in mitosis. Overall, we conclude that continuous Cdk1 activity is not essential to maintain the mitotic state and that phosphatase activity directed at Cdk1 substrates is largely quiescent during mitosis. Furthermore, the degradation of a protein other than cyclin B1 is essential to activate a phosphatase that, in turn, enables mitotic exit.

scholarly journals Mechanisms and regulation of the degradation of cyclin B

1999 ◽  
Vol 354 (1389) ◽  
pp. 1571-1576 ◽  
Author(s):  
A. Hershko
Keyword(s):  
Protein Kinase ◽  
Maximal Activity ◽  
Embryonic Cell ◽  
Cyclin B ◽  
Anaphase Promoting Complex ◽  
Ubiquitin Pathway ◽  
Cell Cycles ◽  
Ubiquitin Protein Ligase ◽  
B Subunit ◽  
Kinetics Of

The degradation of the cyclin B subunit of protein kinase Cdk1/cyclin B is required for inactivation of the kinase and exit from mitosis. Cyclin B is degraded by the ubiquitin pathway, a system involved in most selective protein degradation in eukaryotic cells. In this pathway, proteins are targeted for degradation by ligation to ubiquitin, a process carried out by the sequential action of three enzymes: the ubiquitin–activating enzyme E1, a ubiquitin–carrier protein E2 and a ubiquitin–protein ligase E3. In the system responsible for cyclin B degradation, the E3–like function is carried out by a large complex called cyclosome or anaphase–promoting complex (APC). In the early embryonic cell cycles, the cyclosome is inactive in the interphase, but becomes active at the end of mitosis. Activation requires phosphorylation of the cyclosome/APC by protein kinase Cdk1/cyclin B. The lag kinetics of cyclosome activation may be explained by Suc1–assisted multiple phosphorylations of partly phosphorylated complex. The presence of a Fizzy/Cdc20–like protein is necessary for maximal activity of the mitotic form of cyclosome/APC in cyclin–ubiquitin ligation.

scholarly journals Coupling of T161 and T14 phosphorylations protects cyclin B–CDK1 from premature activation

2011 ◽  
Vol 22 (21) ◽  
pp. 3971-3985 ◽  
Author(s):  
Katia Coulonval ◽  
Hugues Kooken ◽  
Pierre P. Roger
Keyword(s):  
Substantial Reduction ◽  
Cyclin B1 ◽  
Cyclin B ◽  
Activation Loop ◽  
Coupling Mechanism ◽  
Cyclin Dependent Kinase ◽  
Leptomycin B ◽  
Two Dimensional Gel Electrophoresis ◽  
Sequence Of Events ◽  
Tightly Coupled

Mitosis is triggered by the abrupt dephosphorylation of inhibitory Y15 and T14 residues of cyclin B1–bound cyclin-dependent kinase (CDK)1 that is also phosphorylated at T161 in its activation loop. The sequence of events leading to the accumulation of fully phosphorylated cyclin B1–CDK1 complexes remains unclear. Two-dimensional gel electrophoresis allowed us to determine whether T14, Y15, and T161 phosphorylations occur on same CDK1 molecules and to characterize the physiological occurrence of their seven phosphorylation combinations. Intriguingly, in cyclin B1–CDK1, the activating T161 phosphorylation never occurred without the T14 phosphorylation. This strict association could not be uncoupled by a substantial reduction of T14 phosphorylation in response to Myt1 knockdown, suggesting some causal relationship. However, T14 phosphorylation was not directly required for T161 phosphorylation, because Myt1 knockdown did uncouple these phosphorylations when leptomycin B prevented cyclin B1–CDK1 complexes from accumulating in cytoplasm. The coupling mechanism therefore depended on unperturbed cyclin B1–CDK1 traffic. The unexpected observation that the activating phosphorylation of cyclin B1–CDK1 was tightly coupled to its T14 phosphorylation, but not Y15 phosphorylation, suggests a mechanism that prevents premature activation by constitutively active CDK-activating kinase. This explained the opposite effects of reduced expression of Myt1 and Wee1, with only the latter inducing catastrophic mitoses.

scholarly journals Anaphase-Promoting Complex/Cyclosome–Dependent Proteolysis of Human Cyclin a Starts at the Beginning of Mitosis and Is Not Subject to the Spindle Assembly Checkpoint

2001 ◽  
Vol 153 (1) ◽  
pp. 137-148 ◽  
Author(s):  
Stephan Geley ◽  
Edgar Kramer ◽  
Christian Gieffers ◽  
Julian Gannon ◽  
Jan-Michael Peters ◽  
...  
Keyword(s):  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Cyclin A ◽  
Cyclin B1 ◽  
Cyclin B ◽  
Anaphase Promoting Complex ◽  
Wild Type ◽  
Destruction Box

Cyclin A is a stable protein in S and G2 phases, but is destabilized when cells enter mitosis and is almost completely degraded before the metaphase to anaphase transition. Microinjection of antibodies against subunits of the anaphase-promoting complex/cyclosome (APC/C) or against human Cdc20 (fizzy) arrested cells at metaphase and stabilized both cyclins A and B1. Cyclin A was efficiently polyubiquitylated by Cdc20 or Cdh1-activated APC/C in vitro, but in contrast to cyclin B1, the proteolysis of cyclin A was not delayed by the spindle assembly checkpoint. The degradation of cyclin B1 was accelerated by inhibition of the spindle assembly checkpoint. These data suggest that the APC/C is activated as cells enter mitosis and immediately targets cyclin A for degradation, whereas the spindle assembly checkpoint delays the degradation of cyclin B1 until the metaphase to anaphase transition. The “destruction box” (D-box) of cyclin A is 10–20 residues longer than that of cyclin B. Overexpression of wild-type cyclin A delayed the metaphase to anaphase transition, whereas expression of cyclin A mutants lacking a D-box arrested cells in anaphase.

scholarly journals Study of Cyclin Proteolysis in Anaphase-Promoting Complex (APC) Mutant Cells Reveals the Requirement for APC Function in the Final Steps of the Fission Yeast Septation Initiation Network

2001 ◽  
Vol 21 (19) ◽  
pp. 6681-6694 ◽  
Author(s):  
Louise Chang ◽  
Jennifer L. Morrell ◽  
Anna Feoktistova ◽  
Kathleen L. Gould
Keyword(s):  
Kinase Activity ◽  
Nuclear Division ◽  
Cyclin B ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Wild Type ◽  
Mitotic Cyclin ◽  
Septation Initiation Network ◽  
Mutant Cells ◽  
Null Mutants

ABSTRACT Cytokinesis in eukaryotic cells requires the inactivation of mitotic cyclin-dependent kinase complexes. An apparent exception to this relationship is found in Schizosaccharomyces pombemutants with mutations of the anaphase-promoting complex (APC). These conditional lethal mutants arrest with unsegregated chromosomes because they cannot degrade the securin, Cut2p. Although failing at nuclear division, these mutants septate and divide. Since septation requires Cdc2p inactivation in wild-type S. pombe, it has been suggested that Cdc2p inactivation occurs in these mutants by a mechanism independent of cyclin degradation. In contrast to this prediction, we show that Cdc2p kinase activity fluctuates in APCcut mutants due to Cdc13/cyclin B destruction. In APC-null mutants, however, septation and cutting do not occur and Cdc13p is stable. We conclude that APC cut mutants are hypomorphic with respect to Cdc13p degradation. Indeed, overproduction of nondestructible Cdc13p prevents septation in APC cutmutants and the normal reorganization of septation initiation network components during anaphase.

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