Coupling of mitosis to the completion of S phase in Xenopus occurs via modulation of the tyrosine kinase that phosphorylates p34cdc2

Smythe, Carl; Newport, John W.

doi:10.1016/0092-8674(92)90153-4

Cited by 218 publications

(160 citation statements)

References 42 publications

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“…In contrast to the results of Kumagai and Dunphy [10], Smythe and Newport [26] found an elevated tyrosine kinase activity when unreplicated DNA was added to a cycling extract (not arrested by cycloheximide, as in the experiments of Kumagai and Dunphy [10]). Smythe and Newport [26] assayed Wee1 activity by adding a large excess of GST-cyclin (a nondegradable recombinant protein consisting of the glutathione-binding domain of glutathione S-transferase fused to residues 13-401 of sea urchin cyclin B1) together with 0.5 mM vanadate (an inhibitor of tyrosine phosphatases); after a 10 min incubation, Cdc2/ GST-cyclin dimers were recovered by immunoprecipitation and the phosphotyrosine content of Cdc2 was measured.…”

Section: Conservation Equationsmentioning

confidence: 57%

“…However Cdc2/cycB dimers remain tyrosine phosphorylated (inactive), suggesting that the checkpoint signal may work through the tyrosine modifying enzymes, Wee1 and Cdc25. Smythe and Newport [26] found that tyrosine kinase activity increases 5-10-fold in the presence of unreplicated DNA. Other studies [27,28] established that activities of both type 1 and type 2A phosphatases remain elevated in the presence of unreplicated DNA.…”

Section: Modelmentioning

confidence: 99%

“…Smythe and Newport [26] assayed Wee1 activity by adding a large excess of GST-cyclin (a nondegradable recombinant protein consisting of the glutathione-binding domain of glutathione S-transferase fused to residues 13-401 of sea urchin cyclin B1) together with 0.5 mM vanadate (an inhibitor of tyrosine phosphatases); after a 10 min incubation, Cdc2/ GST-cyclin dimers were recovered by immunoprecipitation and the phosphotyrosine content of Cdc2 was measured. Table 1, with parameter values in Table 2, except [CKI] = 0 (no inhibitor) and [10]: 32 nM nondegradable DcycB was added to M-phase extracts (S) and to interphase extracts with no nuclei (A, −aph) and with 1000 nuclei/ml plus 100 mg/ml aphidicolin (X, +aph), and tyrosine-phosphorylated complexes were quantified by anti-phosphotyrosine antibody (arbitrary units).…”

Section: Conservation Equationsmentioning

confidence: 99%

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Modeling M-phase control in Xenopus oocyte extracts: the surveillance mechanism for unreplicated DNA

Marlovits

Tyson

Novák

et al. 1998

Biophysical Chemistry

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Alternating phases of DNA synthesis and mitosis, during the first 12 cell divisions of frog embryos, are driven by autonomous cytoplasmic oscillations of M-phase promoting factor (MPF). Cell-free extracts of frog eggs provide a convenient preparation for studying the molecular machinery that generates MPF oscillations and the surveillance mechanism that normally prevents entry into mitosis until chromosomal DNA is fully replicated. Early experiments suggested that unreplicated DNA blocks MPF activity by inducing phosphorylation of a crucial tyrosine residue, but recent evidence implicates a stoichiometric inhibitor (an MPF binding protein) as the 'braking' agent. Using a realistic mathematical model of the mitotic control system in frog egg extracts, we suggest that both tyrosine phosphorylation and a stoichiometric inhibitors are involved in the block of MPF by unreplicated DNA. Both pathways operate by raising the cyclin threshold for MPF activation. As a bonus, in the process of analyzing these experiments, we obtain more direct and reliable estimates of the rate constants in the model.

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Section: Conservation Equationsmentioning

confidence: 57%

Section: Modelmentioning

confidence: 99%

Section: Conservation Equationsmentioning

confidence: 99%

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Modeling M-phase control in Xenopus oocyte extracts: the surveillance mechanism for unreplicated DNA

Marlovits

Tyson

Novák

et al. 1998

Biophysical Chemistry

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“…We therefore determined whether ATR kinase was a ected by ca eine and LY294002. We found that levels of ca eine which overcome checkpoint arrest inhibited both basal and DNA-activated ATR kinase activity (Figure 4a) but did not have an e ect on the mitotic cdc2 kinase (data not shown (Crompton et al, 1993;Smythe and Newport, 1992). Similarly, LY294002 was found to be an e ective inhibitor of both basal and DNA-activated ATR kinase activity (Figure 4b), although the concentration of LY294002 required for inhibition is substantially higher than that required for PI-3 kinase (Cheatham et al, 1994).…”

Section: Atr Is a Dna-activated P53 Kinasementioning

confidence: 92%

“…ATR and its homologue in S. pombe, rad3, have been implicated in both the S/M and G2/M checkpoints. In human (Schlegel and Pardee, 1986), yeast (Osman and McCready, 1998) and Xenopus cell free systems (Smythe and Newport, 1992), cell cycle arrest induced by these checkpoints has been shown to be suppressed in the presence of ca eine. Given the similarity between ATR and PI-3 kinase, we also wished to establish whether inhibitors of the latter protein blocked ATR function.…”

Section: Atr Is a Dna-activated P53 Kinasementioning

confidence: 99%

ATR is a caffeine-sensitive, DNA-activated protein kinase with a substrate specificity distinct from DNA-PK

et al. 1999

Self Cite

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ATR is a large, 4300 kDa protein containing a carboxy-terminus kinase domain related to PI-3 kinase, and is homologous to the ATM gene product in human cells and the rad3/MEC1 proteins in yeast. These proteins, together with the DNA-PK, are part of a new family of PI-3 kinase related proteins. All members of this family play important roles in checkpoints which operate to permit cell survival following many forms of DNA damage. We have expressed ATR protein in HEK293 cells and puri®ed the protein to near-homogeneity. We show that pure ATR is a protein kinase which is activated by circular single-stranded, doublestranded or linear DNA. Thus ATR is a new member of a sub-family of PIK related kinases, founded by the DNA-PK, which are activated in the presence of DNA. Unlike DNA-PK, ATR does not appear to require Ku proteins for its activation by DNA. We show directly that, like ATM and DNA-PK, ATR phosphorylates the genome surveillance protein p53 on serine 15, a site which is up-regulated in response to DNA damage. In addition, we ®nd that ATR has a substrate speci®city similar to, but unique from, the DNA-PK in vitro, suggesting that these proteins have overlapping but distinct functions in vivo. Finally, we ®nd that the kinase activity of ATR in the presence and absence of DNA is suppressed by ca eine, a compound which is known to induce loss of checkpoint control. Our results are consistent with the notion that ATR plays a role in monitoring DNA structure and phosphorylation of proteins involved in the DNA damage response pathways.

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Maitotoxin, a calcium channel activator, inhibits cell cycle progression through the G1/S and G2/M transitions and prevents CDC2 kinase activation in GH4C1 cells

Dolah

Ramsdell

1996

J. Cell. Physiol.

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Calcium regulates progression through several checkpoints in the cell cycle, including the G1/S-phase transition, G2/M-phase transition, and exit from mitosis. In the GH4C1 rat pituitary cell line, calcium mobilizing polypeptides and calcium channel activation inhibit cell proliferation. This report examines the effects of maitotoxin (MTX), an activator of type L voltage-dependent calcium channels (L-VDCC), on calcium influx and cell cycle progression in GH4C1 cells. MTX causes both a block from G1 to S-phase and a concentration-dependent accumulation of cells in G2+M. MTX does not increase the mitotic index; thus, sustained calcium channel activation by MTX results in an accumulation of cells in G2. In order to temporally localize the MTX-induced G2 block relative to cell cycle regulatory events at the G2/M transition, we assessed the relative activity of two cell cycle regulatory protein kinases, CDC2 and CDK2, in MTX-treated cells. CDC2-specific histone kinase activity in MTX-treated cells is lower than either in cells blocked in mitosis with the microtubule destabilizing agent demecolcine or in randomly cycling cells. In contrast, the activity of CDK2 is highest in MTX-treated cells, consistent with a G2 block prior to CDC2 activation. Together, these results implicate with a G2 block prior to CDC2 activation. Together, these results implicate calcium as an intracellular signal required for progression through G2 phase of the cell cycle prior to CDC2 kinase activation.

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Coupling of mitosis to the completion of S phase in Xenopus occurs via modulation of the tyrosine kinase that phosphorylates p34cdc2

Cited by 218 publications

References 42 publications

Modeling M-phase control in Xenopus oocyte extracts: the surveillance mechanism for unreplicated DNA

Modeling M-phase control in Xenopus oocyte extracts: the surveillance mechanism for unreplicated DNA

ATR is a caffeine-sensitive, DNA-activated protein kinase with a substrate specificity distinct from DNA-PK

Maitotoxin, a calcium channel activator, inhibits cell cycle progression through the G1/S and G2/M transitions and prevents CDC2 kinase activation in GH4C1 cells

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