scholarly journals Cyclin A2 Regulates Nuclear-Envelope Breakdown and the Nuclear Accumulation of Cyclin B1

2007 ◽  
Vol 17 (1) ◽  
pp. 85-91 ◽  
Author(s):  
Delquin Gong ◽  
Joseph R. Pomerening ◽  
Jason W. Myers ◽  
Christer Gustavsson ◽  
Joshua T. Jones ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Cyclin B1 ◽  
Nuclear Accumulation ◽  
Nuclear Envelope Breakdown ◽  
Cyclin A2

scholarly journals Nucleo-cytoplasmic interactions that control nuclear envelope breakdown and entry into mitosis in the sea urchin zygote

1999 ◽  
Vol 112 (8) ◽  
pp. 1139-1148 ◽  
Author(s):  
E.H. Hinchcliffe ◽  
E.A. Thompson ◽  
F.J. Miller ◽  
J. Yang ◽  
G. Sluder
Keyword(s):  
Nuclear Envelope ◽  
Dna Synthesis ◽  
Sea Urchin ◽  
Mammalian Cells ◽  
Cyclin B1 ◽  
Nuclear Accumulation ◽  
Dna Breaks ◽  
Male Pronucleus ◽  
Nuclear Envelope Breakdown ◽  
Female Pronucleus

In sea urchin zygotes and mammalian cells nuclear envelope breakdown (NEB) is not driven simply by a rise in cytoplasmic cyclin dependent kinase 1-cyclin B (Cdk1-B) activity; the checkpoint monitoring DNA synthesis can prevent NEB in the face of mitotic levels of Cdk1-B. Using sea urchin zygotes we investigated whether this checkpoint prevents NEB by restricting import of regulatory proteins into the nucleus. We find that cyclin B1-GFP accumulates in nuclei that cannot complete DNA synthesis and do not break down. Thus, this checkpoint limits NEB downstream of both the cytoplasmic activation and nuclear accumulation of Cdk1-B1. In separate experiments we fertilize sea urchin eggs with sperm whose DNA has been covalently cross-linked to inhibit replication. When the pronuclei fuse, the resulting zygote nucleus does not break down for >180 minutes (equivalent to three cell cycles), even though Cdk1-B activity rises to greater than mitotic levels. If pronuclear fusion is prevented, then the female pronucleus breaks down at the normal time (average 68 minutes) and the male pronucleus with cross-linked DNA breaks down 16 minutes later. This male pronucleus has a functional checkpoint because it does not break down for >120 minutes if the female pronucleus is removed just prior to NEB. These results reveal the existence of an activity released by the female pronucleus upon its breakdown, that overrides the checkpoint in the male pronucleus and induces NEB. Microinjecting wheat germ agglutinin into binucleate zygotes reveals that this activity involves molecules that must be actively translocated into the male pronucleus.

scholarly journals Nuclear-cytoplasmic compartmentalization promotes robust timing of mitotic events by cyclin B1-Cdk1

2021 ◽  
Author(s):  
Gembu Maryu ◽  
Qiong Yang
Keyword(s):  
Phase Plane ◽  
Nuclear Translocation ◽  
Spatial Models ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Cyclin B1 ◽  
Nuclear Accumulation ◽  
Nuclear Envelope Breakdown ◽  
Synthetic Cells ◽  
Tunable Frequency

Studies applying well-mixed cytosolic extracts found the mitotic network centered on cyclin-dependent kinase (Cdk1) performs robust relaxation oscillations with tunable frequency. However, recent work also highlighted the importance of cyclin B1-Cdk1 nuclear translocation in mitotic timing. How nuclear compartmentalization affects the oscillator properties and the accurate ordering of mitotic events, especially in embryos lacking checkpoints, remains elusive. Here we developed a Forster resonance energy transfer (FRET) biosensor for analyzing Cdk1 spatiotemporal dynamics in synthetic cells containing nuclei compared to those without. We found cellular compartmentalization significantly impacts clock behaviors. While the amplitude-frequency dependency measured in the homogeneous cytoplasm showed highly tunable frequency for a fixed amplitude, confirming predictions by non-spatial models, the frequency remains constant against cyclin variations when nuclei are present, suggesting a possible buffering mechanism of nuclear compartments to ensure robust timing. We also found all cyclin degrades within similar mitotic durations despite variable interphase cyclin expression. This scalable degradation of cyclin may further promote the precise mitotic duration. Simultaneous measurements revealed Cdk1 and cyclin B1 cycle rigorously out of phase, producing a wide orbit on their phase plane, essential for robust oscillations. We further mapped mitotic events on the phase-plane orbits. Unlike cytoplasmic-only cells showing delayed Cdk1 activation, nucleus-containing cells exhibit steady cyclin B1-Cdk1 nuclear accumulation until nuclear envelope breakdown (NEB) followed by an abrupt cyclin-independent activation to trigger anaphase. Thus, both biphasic activation and subcellular localization of Cdk1 ensure accurate ordering of substrates.

133. SPATIAL REGULATION OF APCCdh1 INDUCED CYCLIN B1 DEGRADATION MAINTAINS GV ARREST IN MOUSE OOCYTES

2009 ◽  
Vol 21 (9) ◽  
pp. 52
Author(s):  
J. E. Holt ◽  
J. L. Weaver ◽  
K. T. Jones
Keyword(s):  
Cyclin B1 ◽  
26S Proteasome ◽  
Nuclear Accumulation ◽  
Regulatory Subunit ◽  
Anaphase Promoting Complex ◽  
Intracellular Location ◽  
Nuclear Localisation ◽  
Nuclear Envelope Breakdown ◽  
Mouse Oocytes ◽  
Nuclear Levels

Within the mammalian ovary, oocytes remain prophase I arrested until a hormonal cue triggers meiotic resumption. The E3 ubiquitin ligase, Anaphase-Promoting Complex with its co-activator Cdh1 (APCCdh1) is known to be essential for this process by promoting cyclin B1 degradation via the 26S proteasome. Cyclin B1 is the regulatory subunit of Maturation-Promoting Factor, forming a heterodimer with CDK1, which is essential for nuclear envelope breakdown (NEB). Here we describe the intracellular partitioning that would explain how Cdh1 activity is fine-tuned, such that cyclin B1 is maintained at sufficient levels to allow oocytes to resume meiosis but not so high as to cause premature meiosis re-entry. Using RT-PCR we detected only one splice variant in mouse oocytes, Cdh1α, which possessed a nuclear localisation signal. By immunofluorescence we confirmed the nuclear location of Cdh1 and degradation machinery components including essential subunits of the APC and 26S proteasome. In all systems studied, cyclin B1 shuttles between the cytoplasm and nucleus, with nuclear localisation occurring just before NEB. We reasoned therefore that the nuclear localisation of the APCCdh1 and 26S proteasome would aid in maintaining low nuclear levels of cyclin B1. Using two GFP-coupled cyclin B1 mutants, which differed in their intracellular location we found that nuclear accumulation of cyclin B1 was necessary in order for it to promote meiotic resumption because over-expression of nuclear-cyclin B1 accelerated entry into meiosis, whereas cytoplasmic-cyclin B1 did not. However, in milrinone-arrested GV oocytes rates of nuclear-cyclin B1 degradation were 5 fold higher than cytoplasmic-cyclin B1. Therefore we conclude that in oocytes, an increase in the nuclear-cytoplasmic ratio of cyclin B1 is an essential step in meiotic resumption, and that nuclear APCCdh1 activity guards against early meiotic resumption, until the degradation machinery is overwhelmed by cyclin B1 translocation.

scholarly journals Activation of cyclin B1–Cdk1 synchronizes events in the nucleus and the cytoplasm at mitosis

2010 ◽  
Vol 189 (2) ◽  
pp. 247-259 ◽  
Author(s):  
Olivier Gavet ◽  
Jonathon Pines
Keyword(s):  
Nuclear Import ◽  
Nuclear Export ◽  
Nuclear Lamina ◽  
Fold Increase ◽  
Cyclin B1 ◽  
Cyclin B ◽  
Nuclear Accumulation ◽  
Animal Cells ◽  
Nuclear Envelope Breakdown ◽  
Prevailing View

The cyclin B–Cdk1 kinase triggers mitosis in most eukaryotes. In animal cells, cyclin B shuttles between the nucleus and cytoplasm in interphase before rapidly accumulating in the nucleus at prophase, which promotes disassembly of the nuclear lamina and nuclear envelope breakdown (NEBD). What triggers the nuclear accumulation of cyclin B1 is presently unclear, although the prevailing view is that the Plk1 kinase inhibits its nuclear export. In this study, we use a biosensor specific for cyclin B1–Cdk1 activity to show that activating cyclin B1–Cdk1 immediately triggers its rapid accumulation in the nucleus through a 40-fold increase in nuclear import that remains dependent on Cdk1 activity until NEBD. Nevertheless, a substantial proportion of cyclin B1–Cdk1 remains in the cytoplasm. The increase in nuclear import is driven by changes in the nuclear import machinery that require neither Plk1 nor inhibition of nuclear export. Thus, the intrinsic link between cyclin B1–Cdk1 activation and its rapid nuclear import inherently coordinates the reorganization of the nucleus and the cytoplasm at mitotic entry.

scholarly journals The Roles of Cyclin A2, B1, and B2 in Early and Late Mitotic Events

2010 ◽  
Vol 21 (18) ◽  
pp. 3149-3161 ◽  
Author(s):  
Delquin Gong ◽  
James E. Ferrell
Keyword(s):  
Chromatin Condensation ◽  
Time Lapse ◽  
Cyclin B1 ◽  
S Phase ◽  
Epifluorescence Microscopy ◽  
Nuclear Accumulation ◽  
H3 Phosphorylation ◽  
Resistant Form ◽  
Cyclin A2 ◽  
Histone H3 Phosphorylation

Here we have used siRNAs and time-lapse epifluorescence microscopy to examine the roles of various candidate mitotic cyclins in chromatin condensation in HeLa cells. Knocking down cyclin A2 resulted in a substantial (∼7 h) delay in chromatin condensation and histone H3 phosphorylation, and expressing an siRNA-resistant form of cyclin A2 partially rescued chromatin condensation. There was no detectable delay in DNA replication in the cyclin A2 knockdowns, arguing that the delay in chromatin condensation is not secondary to a delay in S-phase completion. Cyclin A2 is required for the activation and nuclear accumulation of cyclin B1-Cdk1, raising the possibility that cyclin B1-Cdk1 mediates the effects of cyclin A2. Consistent with this possibility, we found that chromatin condensation was tightly associated temporally with the redistribution of cyclin B1 to the nucleus. Moreover, a constitutively nuclear cyclin B1 rescued chromatin condensation in cyclin A2 knockdown cells. On the other hand, knocking down cyclin B1 delayed chromatin condensation by only about one hour. Our working hypothesis is that active, nuclear cyclin B1-Cdk1 normally cooperates with cyclin A2 to bring about early mitotic events. Because cyclin A2 is present only during the early stages of mitosis, we asked whether cyclin B knockdown might have more dramatic defects on late mitotic events. Consistent with this possibility, we found that cyclin B1- and cyclin B1/B2-knockdown cells had difficulty in maintaining a mitotic arrest in the presence of nocodazole. Taken together, these data suggest that cyclin A2 helps initiate mitosis, in part through its effects on cyclin B1, and that cyclins B1 and B2 are particularly critical for the maintenance of the mitotic state.

Cyclin A and Nek2A: APC/C–Cdc20 substrates invisible to the mitotic spindle checkpoint

2010 ◽  
Vol 38 (1) ◽  
pp. 72-77 ◽  
Author(s):  
Wouter van Zon ◽  
Rob M.F. Wolthuis
Keyword(s):  
Nuclear Envelope ◽  
Spindle Checkpoint ◽  
Cyclin A ◽  
Cyclin B1 ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Nuclear Envelope Breakdown ◽  
Anaphase Onset ◽  
Sister Chromatids ◽  
Spindle Microtubules

Active cyclin B1–Cdk1 (cyclin-dependent kinase 1) keeps cells in mitosis, allowing time for spindle microtubules to capture the chromosomes and for incorrect chromosome-spindle attachments to be repaired. Meanwhile, securin, an inhibitor of separase, secures cohesion between sister chromatids, preventing anaphase onset. The spindle checkpoint is a signalling pathway emerging from improperly attached chromosomes that inhibits Cdc20, the mitotic activator of the APC/C (anaphase-promoting complex/cyclosome) ubiquitin ligase. Blocking Cdc20 stabilizes cyclin B1 and securin to delay mitotic exit and anaphase until all chromosomes reach bipolar spindle attachments. Cells entering mitosis in the absence of a functional spindle checkpoint degrade cyclin B1 and securin right after nuclear-envelope breakdown, in prometaphase. Interestingly, two APC/C substrates, cyclin A and Nek2A, are normally degraded at nuclear-envelope breakdown, even when the spindle checkpoint is active. This indicates that the APC/C is activated early in mitosis, whereas cyclin B1 and securin are protected as long as the spindle checkpoint inhibits Cdc20. Remarkably, destruction of cyclin A and Nek2A also depends on Cdc20. The paradox of Cdc20 being both active and inhibited in prometaphase could be explained if cyclin A and Nek2A are either exceptionally efficient Cdc20 substrates, or if they are equipped with ‘stealth’ mechanisms to effectively escape detection by the spindle checkpoint. In the present paper, we discuss recently emerging models for spindle-checkpoint-independent APC/C–Cdc20 activity, which might even have implications for cancer therapy.

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