Sea urchin zygote chromatin exhibit an unfolded nucleosomal array during the first S phase

The proteasome has been shown to be involved in exit from mitosis by bringing about destruction of mitotic cyclins. Here, we present evidence that the proteasome is also required for proper completion of S phase and for entry into mitosis in the sea urchin embryonic cleavage cycle. A series of structurally related peptide-aldehydes prevent nuclear envelope breakdown in their order of inhibitory efficacies against the proteasome. Their efficacies in blocking exit from S phase and exit from mitosis correlate well, indicating that the proteasome is involved at both these steps. Mitotic histone HI kinase activation and tyrosine dephosphorylation of p34(cdc2) kinase are blocked by inhibition of the proteasome, indicating that the proteasome plays an important role in the pathway that leads to embryonic p34(cdc2)kinase activation. Arrested embryos continued to incorporate [(3)H]thymidine and characteristically developed large nuclei. Pre-mitotic arrest can be overcome by treatment with caffeine, a manoeuvre that is known to override the DNA replication checkpoint. These data demonstrate that the proteasome is involved in the control of termination of S phase and consequently in the initiation of M phase of the first embryonic cell cycle.

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ROLE OF CYCLIN DEPENDENT KINASE IN G1→S TRANSITION AND IN S PHASE OF THE FIRST SEA URCHIN EMBRYONIC CYCLE

Biology of the Cell ◽

10.1016/s0248-4900(97)86848-7 ◽

1996 ◽

Vol 88 (1-2) ◽

pp. 71-71

Author(s):

Jean-Luc Moreau ◽

Gérard Peaucellier ◽

André Picard ◽

Anne Marie Genevière

Keyword(s):

Sea Urchin ◽

S Phase ◽

Cyclin Dependent Kinase

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Nuclear AP4A-binding activity of sea urchin embryos changes in relation to the initiation of S phase

Experimental Cell Research ◽

10.1016/0014-4827(86)90596-3 ◽

1986 ◽

Vol 165 (2) ◽

pp. 429-440 ◽

Cited By ~ 3

Author(s):

Mizue Morioka ◽

Hiraku Shimada

Keyword(s):

Sea Urchin ◽

Binding Activity ◽

S Phase ◽

Sea Urchin Embryos

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Differential replication capacities of G1 and S-phase extracts from sea urchin eggs

Journal of Cell Science ◽

10.1242/jcs.104.2.565 ◽

1993 ◽

Vol 104 (2) ◽

pp. 565-572

Author(s):

H. Zhang ◽

J.V. Ruderman

Keyword(s):

Dna Synthesis ◽

Sea Urchin ◽

Nuclear Dna ◽

Specific Pattern ◽

S Phase ◽

Sperm Chromatin ◽

G1 Arrest ◽

Sea Urchin Eggs ◽

Egg Extracts ◽

Fertilized Egg

Sea urchin eggs are arrested in G1 of the first mitotic cell cycle. Fertilization triggers release from G1 arrest and the onset of DNA synthesis about 20 minutes later, even when protein synthesis is blocked. Here we describe extracts from eggs and S-phase embryos that reproduce this stage-specific pattern of DNA synthesis. Fertilized egg extracts formed nuclear membranes around decondensed Xenopus sperm chromatin whereas unfertilized egg extracts did not. Aphidicolin-sensitive deoxynucleotide incorporation was high in extracts of fertilized S-phase eggs and low in those of unfertilized eggs. In contrast, single-stranded DNA templates directed high rates of incorporation in both unfertilized and fertilized egg extracts, suggesting that the stage-specific activities in nuclear DNA synthesis is restricted to initiation on double-stranded DNA. Mixing experiments showed that unfertilized eggs do not contain a dominant inhibitor of replication, nor does fertilization induce the appearance of a soluble, dominant activator.

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The Coordination of Centrosome Reproduction with Nuclear Events of the Cell Cycle in the Sea Urchin Zygote

The Journal of Cell Biology ◽

10.1083/jcb.140.6.1417 ◽

1998 ◽

Vol 140 (6) ◽

pp. 1417-1426 ◽

Cited By ~ 64

Author(s):

Edward H. Hinchcliffe ◽

Grizzel O. Cassels ◽

Conly L. Rieder ◽

Greenfield Sluder

Keyword(s):

Cell Cycle ◽

Sea Urchin ◽

S Phase ◽

Cyclin B ◽

Cyclin Dependent Kinase ◽

Phase Arrest ◽

Stage Specificity ◽

S Phase Arrest ◽

Nuclear Events ◽

Block Protein Synthesis

Centrosomes repeatedly reproduce in sea urchin zygotes arrested in S phase, whether cyclin-dependent kinase 1–cyclin B (Cdk1-B) activity remains at prefertilization levels or rises to mitotic values. In contrast, when zygotes are arrested in mitosis using cyclin B Δ-90, anaphase occurs at the normal time, yet centrosomes do not reproduce. Together, these results reveal the cell cycle stage specificity for centrosome reproduction and demonstrate that neither the level nor the cycling of Cdk1-B activity coordinate centrosome reproduction with nuclear events. In addition, the proteolytic events of the metaphase–anaphase transition do not control when centrosomes duplicate. When we block protein synthesis at first prophase, the zygotes divide and arrest before second S phase. Both blastomeres contain just two complete centrosomes, which indicates that the cytoplasmic conditions between mitosis and S phase support centrosome reproduction. However, the fact that these daughter centrosomes do not reproduce again under such supportive conditions suggests that they are lacking a component required for reproduction. The repeated reproduction of centrosomes during S phase arrest points to the existence of a necessary “licensing” event that restores this component to daughter centrosomes during S phase, preparing them to reproduce in the next cell cycle.

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ERK1 activation is required for S-phase onset and cell cycle progression after fertilization in sea urchin embryos

Development ◽

10.1242/dev.01607 ◽

2005 ◽

Vol 132 (3) ◽

pp. 579-589 ◽

Cited By ~ 17

Author(s):

R. Philipova

Keyword(s):

Cell Cycle ◽

Sea Urchin ◽

Cell Cycle Progression ◽

S Phase ◽

Cycle Progression ◽

Sea Urchin Embryos

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Caffeine overrides the S-phase cell cycle block in sea urchin embryos

Zygote ◽

10.1017/s0967199400003804 ◽

1997 ◽

Vol 5 (2) ◽

pp. 127-138 ◽

Cited By ~ 7

Author(s):

Rajnikant Patel ◽

Elizabeth M. Wright ◽

Michael Whitaker

Keyword(s):

Protein Kinase ◽

Sea Urchin ◽

S Phase ◽

Phase Cell ◽

Protein Kinase Activity ◽

Checkpoint Control ◽

Cell Cycles ◽

Sea Urchin Embryos ◽

Sea Urchin Embryo ◽

M Phase

SummaryDuring the early mitotic cell cycles of the sea urchin embryo, the cell oscillates between S-phase and M-phase. In the presence of aphidicolin, a DNA synthesis inhibitor, a checkpoint control blocks the activation of the p34cdc2 protein kinase, by keeping it in the inactive, tyrosine phosphorylated form, and the embryos do not enter mitosis. Caffeine has been shown to bypass the G2/M-phase checkpoint in mammalian cells and in cycling Xenopus extracts and to induce mitosis despite the presence of damaged or unreplicated DNA. In this study we show that caffeine also induces mitosis and cell division in sea urchin embryos, in the presence of unreplicated DNA, by stimulating the tyrosine dephosphorylation of p34cdc2 and switching on its protein kinase activity. We also show that the caffeine-induced activation of the p34cdc2 protein kinase is not mediated by either of the two second messengers, calcium and cAMP, or by inhibition of the p34cdc2 tyrosine kinase. Thus, none of the mechanisms proposed for caffeine's action can explain how it overrides the S-phase checkpoint in the early cell cycles of the sea urchin embryo.

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Temporal Correlations between Sensitivity to Radiation-induced Mitotic Delay and the S Phase of the Sea Urchin Egg

International Journal of Radiation Biology and Related Studies in Physics Chemistry and Medicine ◽

10.1080/09553007914550431 ◽

1979 ◽

Vol 35 (4) ◽

pp. 361-364

Author(s):

R.C. Rustad ◽

G. Viswanathan ◽

B.C. Antonellis

Keyword(s):

Sea Urchin ◽

S Phase ◽

Temporal Correlations ◽

Sea Urchin Egg ◽

Mitotic Delay ◽

Radiation Induced

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Active cyclin B-cdc2 kinase does not inhibit DNA replication and cannot drive prematurely fertilized sea urchin eggs into mitosis

Journal of Cell Science ◽

10.1242/jcs.108.7.2693 ◽

1995 ◽

Vol 108 (7) ◽

pp. 2693-2703 ◽

Cited By ~ 8

Author(s):

A.M. Geneviere-Garrigues ◽

A. Barakat ◽

M. Doree ◽

J.L. Moreau ◽

A. Picard

Keyword(s):

Dna Replication ◽

Sea Urchin ◽

S Phase ◽

Cyclin B ◽

G1 Arrest ◽

Cdc2 Kinase ◽

Kinase Activation ◽

M Phase ◽

Set Up ◽

The One

Feedback mechanisms preventing M phase occurrence before S phase completion are assumed to depend on inhibition of cyclin B-cdc2 kinase activation by unreplicated DNA. In sea urchin, fertilization stimulates protein synthesis and releases eggs from G1 arrest. We found that in the one-cell sea urchin embryo cyclin B-cdc2 kinase undergoes partial activation before S phase, reaching in S phase a level that is sufficient for G2-M phase transition. S phase entry is not inhibited by this level of cyclin B-dependent kinase activity. Inhibition of DNA replication by aphidicolin suppresses nuclear envelope breakdown, yet it does not prevent the microtubule array from being converted from its interphasic to its mitotic state. Moreover, mitotic cytoplasmic events occur at the same time in control and aphidicolin-treated embryos. Thus unreplicated DNA only prevents mitotic nuclear, not cytoplasmic, events from occurring prematurely. These results together show that the inhibition of cyclin B-cdc2 kinase activation is probably not the only mechanism that prevents mitotic nuclear events from occurring as long as DNA replication has not been completed. In contrast, cytoplasmic mitotic events seem to be controlled by a timing mechanism independent of DNA replication, set up at fertilization, that prevents premature opening of a window for mitotic events.

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Cytoskeletal elements link calcium channel activity and the cell cycle in early sea urchin embryos

Development ◽

10.1242/dev.121.6.1827 ◽

1995 ◽

Vol 121 (6) ◽

pp. 1827-1831

Author(s):

I. Yazaki ◽

E. Tosti ◽

B. Dale

Keyword(s):

Cell Cycle ◽

Calcium Channels ◽

Calcium Channel ◽

Sea Urchin ◽

Cytochalasin B ◽

S Phase ◽

Channel Activity ◽

Significant Delay ◽

M Phase ◽

Cytoskeletal Elements

Using the whole-cell clamp technique, we show that L-type calcium channels are activated in early sea urchin blastomeres during M-phase and subsequently inactivated in S-phase. This cyclical channel behaviour occurs in the absence of the nucleus suggesting cytoplasmic regulation independent of the centrosome cycle. Puromycin at 100–400 micromolar does not prevent inactivation of the current showing that this phase, at least, does not require protein synthesis. Cytochalasin B at 2 microgram/ml inhibits the cyclical activity in both M and S phases, while 100 microgram/ml of colchicine inactivates the L-type current in M-phase and activates a large T-type calcium current in S-phase, suggesting that channel behaviour is regulated by cytoskeletal elements. Since, fragmentation experiments show the calcium channels to be clustered in the apical membrane, and some L-type calcium channel inhibitors induced a significant delay in the cell cycle, the channel may play a role in regulating cytokinesis possibly by contributing to local intracellular calcium gradients.

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