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.

REFERENCES

Adachi
Y.
,
Laemmli
U. K.
(
1994
).
Study of the cell cycle-dependent assembly of the DNA pre-replication centres in xenopus egg extracts.
EMBO J
13
,
4153
4164
Alfa
C. E.
,
Hyams
J. S.
,
Ducommun
B.
,
Beach
D.
(
1990
).
Distinct nuclear and spindle pole body populations of cyclin-cdc2 in fission yeast.
Nature
347
,
680
682
Amon
A.
,
Surana
U.
,
Muroff
I.
,
Nasmyth
K.
(
1992
).
Regulation ofp34CDC28tyrosine phosphorylation is not required to entry in mitosis in S.
Cerevisiae. Nature
355
,
368
371
Booher
R. N.
,
Alfa
C. E.
,
Hyams
J. S.
,
Beach
D. H.
(
1989
).
The fission yeast cdc2/cdc13/suc1 protein kinase.
Cell
58
,
485
497
Broek
D.
,
Bartlett
R.
,
Crawford
K.
,
Nurse
P.
(
1991
).
Involvement of p34cdc2in establishing the dependency of S phase on mitosis.
Nature
349
,
388
393
Dasso
M.
,
Newport
J. W.
(
1990
).
Completion of DNA replication is monitored by a feedback system that controls the initiation of mitosis in vitro: studies in Xenopus.
Cell
61
,
811
823
Dunphy
W. G.
,
Newport
J. W.
(
1989
).
Fission yeast p13 blocks mitotic activation and tyrosine dephosphorylation of the Xenopus cdc2 protein kinase.
Cell
58
,
181
191
Dunphy
W. G.
,
Kumagai
A.
(
1991
).
The cdc25 protein contains an intrinsic phosphatase activity.
Cell
67
,
189
196
Edgar
B. A.
,
O'Farrell
P. H.
(
1990
).
The three postblastoderm cell cycles of Drosophila embryogenesis are regulated in G2by string.
Cell
62
,
469
480
Edgecombe
M.
,
Patel
R.
,
Whitaker
M.
(
1991
).
A cyclin-abundance cycle-independent p34cdc2tyrosine phosphorylation cycle in early sea urchin embryos.
EMBO J
10
,
3769
3775
Enoch
T.
,
Nurse
P.
(
1990
).
Mutation of fission yeast cell cycle control genes abolishes dependence of mitosis on DNA replication.
Cell
60
,
665
673
Featherstone
C.
,
Russell
P.
(
1991
).
Fission yeast p107wee1 mitotic inhibitor is a tyrosine/serine kinase.
Nature
349
,
808
811
Ferrel
J. E.
,
Wu
M.
,
Gerhart
J. C.
,
Martin
G. S.
(
1991
).
Cell cycle tyrosine phosphorylation of p34cdc2 and a MAP kinase homolog in Xenopus oocytes and eggs.
Mol. Cell Biol
11
,
1965
1971
Gautier
J.
,
Solomon
M. J.
,
Booher
R. N.
,
Bazan
J. F.
,
Kirshner
M. W.
(
1991
).
Cdc25 is a specific tyrosine phosphatase that directly activates p34cdc2.
Cell
67
,
197
211
Gerhart
J. C.
,
Wu
M.
,
Kirshner
M.
(
1984
).
Cell cycle dynamics of an M-phase specific cytoplasmic factor in Xenopus laevis oocytes and eggs.
J. Cell Biol
98
,
1247
1255
Gould
K. L.
,
Nurse
P.
(
1989
).
Tyrosine phosphorylation of the fission yeast cdc2+ protein kinase regulates entry into mitosis.
Nature
342
,
39
45
Harland
R. M.
,
Laskey
R. A.
(
1980
).
Regulated replication of DNA microinjected into eggs of X. laevis.
Cell
21
,
761
771
Hartwell
L. H.
,
Weinert
T. A.
(
1989
).
Checkpoints: controls that ensure the order of cell cycle events.
Science
246
,
629
634
Hayles
J.
,
Fisher
D.
,
Woollard
A.
,
Nurse
P.
(
1994
).
Temporal order of S phase and mitosis in fission yeast is determined by the state of the p34cdc2-mitotic B cyclin complex.
Cell
78
,
813
822
Hogan
B.
,
Gross
P. R.
(
1971
).
The effect of protein synthesis inhibition on the entry of messenger RNA into the cytoplasm of sea urchin embryos.
J. Cell Biol
49
,
692
701
Kelly
T. J.
,
Martin
G. S.
,
Forsburg
S. L.
,
Stephen
R. J.
,
Russo
A.
,
Nurse
P.
(
1993
).
The fission yeast cdc18+gene product couples S phase to start and mitosis.
Cell
74
,
371
382
King
R. W.
,
Jackson
P. K.
,
Kirshner
M. W.
(
1994
).
Mitosis in transition.
Cell
79
,
563
571
Krek
W.
,
Nigg
E. A.
(
1991
).
Differential phosphorylation of vertebrate p34cdc2kinase at the G1/S and G2/M transitions of the cell cycle: identification of major phosphorylation sites.
EMBO J
10
,
3331
3341
Kumagai
A.
,
Dunphy
W. G.
(
1991
).
The cdc25 protein controls tyrosine dephosphorylation of the cdc2 protein in a cell-free system.
Cell
64
,
903
914
Labbe
J. C.
,
Capony
J. P.
,
Caput
D.
,
Cavadore
J. C.
,
Derancourt
J.
,
Kaghad
M.
,
Lelias
J. M.
,
Picard
A.
,
Doree
M.
(
1989
).
MPF from starfish oocytes at first meiotic metaphase is a heterodimer containing one molecule of cdc2 and one molecule of cyclin B.
EMBO J
8
,
3053
3058
Lowry
O. H.
,
Rosebrough
N. J.
,
Farr
A. L.
,
Randall
R. J.
(
1951
).
Protein measurement with the Folin phenol reagent.
J. Biol. Chem
193
,
265
275
Lundgren
K.
,
Walworth
N.
,
Booher
R.
,
Dembski
M.
,
Kirshner
M.
,
Beach
D.
(
1991
).
mik1 and wee1 cooperate in the tyrosine phosphorylation of cdc2.
EMBO J
10
,
865
875
McGowan
C. H.
,
Russell
P.
(
1993
).
Human wee1 kinase inhibits celldivision by phosphorylating p34cdc2exclusively on tyr 15.
EMBO J
12
,
75
85
Masui
Y.
(
1992
).
Toward understanding the control of the division cycle in animal cells.
Biochem. Cell Biol
70
,
920
945
Meijer
L.
,
Arion
D.
,
Golsteyn
R.
,
Pines
J.
,
Brizuela
L.
,
Hunt
T.
,
Beach
D.
(
1989
).
Cyclin is a component of the sea urchin egg M-phase specific histone H1 kinase.
EMBO J
8
,
2275
2282
Meijer
L.
,
Azzi
L.
,
Wang
J. Y.
(
1991
).
Cyclin B targets p34cdc2for tyrosine phosphorylation.
EMBO J
10
,
1545
1554
Morla
A. O.
,
Draetta
G.
,
Beach
D.
,
Wang
J. Y.
(
1989
).
Reversible tyrosine phosphorylation of cdc2: dephosphorylation accompanies activation during entry into mitosis.
Cell
58
,
193
203
Newport
J. W.
,
Kirshner
M. W.
(
1984
).
Regulation of the cell cycle during early Xenopus development.
Cell
37
,
731
742
Nurse
P.
(
1990
).
Universal control mechanism regulating onset of M-phase.
Nature
344
,
503
508
Nurse
P.
(
1994
).
Ordering S phase and M phase in the cell cycle.
Cell
79
,
547
550
Ookata
K.
,
Hisanaga
Sh.
,
Okano
T.
,
Tachibana
K.
,
Kishimoto
T.
(
1992
).
Relocation and distinct subcellular localization of p34cdc2-cyclinB complex at meiosis reinitiation in starfish oocytes.
EMBO J
11
,
1763
1772
Osmani
A. H.
,
Mc Guire
S. L.
,
Osmani
S. A.
(
1991
).
Parallel activation of the nimA, a gene required for the G2/M transition in Aspergillus nidulans.
Cell
67
,
283
291
Parker
L. L.
,
Atherton-Fessler
S.
,
Lee
M. S.
,
Piwnica-Worms
H.
(
1992
).
p107wee1is a dual-specificity kinase that phosphorylates p34cdc2on tyrosine 15.
Proc. Nat. Acad. Sci. USA
89
,
2917
2921
Picard
A.
,
Labbe
J. C.
,
Peaucellier
G.
,
Le Bouffant
F.
,
Le Peuch
C.
,
Doree
M.
(
1987
).
Changes in the activity of the maturation promoting factor are correlated with those of a major cyclic AMP-and calcium-independent protein kinase during the first meiotic cell cycle in the early starfish embryo.
Dev. Growth Differ
29
,
93
103
Picard
A.
,
Harricane
M. C.
,
Labbe
J. C.
,
Doree
M.
(
1988
).
Germinal vesicle components are not required for the cell-cycle oscillator of the early starfish embryo.
Dev. Biol
128
,
121
128
Picard
A.
,
Labbe
J. C.
,
Barakat
A.
,
Cavadore
J. C.
,
Doree
M.
(
1991
).
Okadaic acid mimics a nuclear component required for cyclin B-cdc2 kinase microinjection to drive starfish oocytes into M phase.
J. Cell Biol
115
,
337
344
Pines
J.
,
Hunter
T.
(
1991
).
Human cyclins A and B1 are differentially located in the cell and undergo cell cycle-dependent nuclear transport.
J. Cell Biol
115
,
1
17
Pines
J.
,
Hunter
T.
(
1994
).
The differential localization of human cyclins A and B is due to a cytoplasmic retention signal in cyclin B.
EMBO J.
,
13
,
3772
3781
Russell
P.
,
Nurse
P.
(
1986
).
cdc 25+functions as an inducer in the mitotic control of fission yeast.
Cell
45
,
145
153
Russell
P.
,
Nurse
P.
(
1987
).
Negative regulation of mitosis by wee1, a gene encoding a protein kinase homolog.
Cell
49
,
559
567
Sakai
M.
,
Kubota
H. Y.
(
1981
).
Cyclic surface changes in the non-nucleate egg fragment of Xenopus laevis.
Dev. Growth Differ
23
,
41
48
Sawai
T.
(
1979
).
Cyclic changes in the cortical layer of non-nucleated fragments of the newt's egg.
J. Embryol. Exp. Morph
51
,
183
189
Schowman
R. M.
,
Foerder
C. A.
(
1979
).
Removal of the fertilisation membrane of sea urchin embryos employing aminotriazole.
Exp. Cell Res
120
,
253
255
Seki
T.
,
Yamashita
K.
,
Nishitani
H.
,
Takagi
T.
,
Russel
P.
,
Nishimoto
T.
(
1992
).
Chromosome condensation caused by loss of RCC1 function requires the cdc25 protein that is located in the cytoplasm.
Mol. Biol. Cell
3
,
1373
1388
Sluder
G.
,
Lewis
K.
(
1987
).
Relationship between nuclear DNA synthesis and centrosome reproduction in sea urchin eggs.
J. Exp. Zool
244
,
89
100
Solomon
M. J.
,
Glotzer
M.
,
Lee
T. H.
,
Philippe
M.
,
Kirshner
M. W.
(
1990
).
Cyclin activation of p34cdc2.
Cell
63
,
1013
1024
Sorger
P.
,
Murray
A. W.
(
1992
).
S-phase feedback control in budding yeast independent of tyrosine phosphorylation of p34cdc28.
Nature
355
,
365
368
Strausfeld
U.
,
Labbe
J. C.
,
Fesquet
D.
,
Cavadore
J. C.
,
Picard
A.
,
Sadhu
K.
,
Russel
P.
,
Doree
M.
(
1991
).
Dephosphorylation and activation of a p34cdc2/cyclin B complex in vitro by human cdc25 protein.
Nature
351
,
242
245
Wagenaar
E. B.
(
1983
).
The timing of synthesis of proteins required for mitosis in the cell cycle of the sea urchin embryo.
Exp. Cell Res
144
,
393
403
This content is only available via PDF.