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Results for “Maturation-Promoting Factor”

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At least 19 recordsLinked to original sources
PubMed1978

Progesterone-induced meiosis in Xenopus laevis oocytes: a role for cAMP at the "maturation-promoting factor" level.

Cholera toxin inhibition of progesterone-induced meiosis of Xenopus laevis oocytes in vitro has been correlated with increased cAMP levels. Inhibition of germinal vesicle breakdown (Gvbd) and cAMP increase occurred after a lag period of 2 hr, when cholera toxin was injected, or 4--5 hr, when applied externally. The ability of the maturation-promoting factor (Mpf) to provoke Gvbd when injected into recipient oocytes was found to be dependent upon whether the oocytes had been exposed to cholera toxin alone or to toxin and progesterone. With the former, cAMP levels were elevated and Mpf activity was abolished, whereas with the latter, the increase in cAMP was less pronounced and Mpf activity was observed. Injection of cAMP or its 8-thio derivatives shortly before the appearance of progesterone-induced Mpf abolished Gvbd. If injected earlier or later, no inhibition was observed. In contrast, cholera toxin inhibited maturation even when added several hours before progesterone, suggesting a sustained accumulation of cAMP. No Gvbd occurred when 8-thio-methyl-cAMP was injected together with Mpf. These data suggest that cAMP is involved in the control of the formation/amplification and/or activity of Mpf-a result which may be of general significance in cell division mechanisms.

Animals↗
PubMed1977

Discussion paper: induction by progesterone and a "maturation-promoting factor" of soluble proteins in Xenopus laevis oocytes in vitro.

In vitro incubation of isolated X. laevis oocytes with progesterone induces the specific labeling of proteins during maturation. They are detected as several discrete peaks with a double-labeling technique that involves the injection of [3H] leucine into hormone-treated and [14C] leucine into control cells. The peaks are separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Identical proteins are obtained with two other steroids that provoke maturation and with p-chloromercuribenzoate. The same progesterone-induced proteins are observed even if oocytes have been treated with actinomycin D or ethidium bromide or have been enucleated. Cycloheximide suppresses the labeling of proteins that are induced by progesterone, if administered either at the time of radioactive amino acid exposure or after prelabeling the oocytes to determine whether the hormone-induced peaks are due to compartmental changes of the proteins. A cytoplasmic fraction, obtained from maturing oocytes before GVBD, promotes maturation when injected into resting oocytes and therefore contains an MPF. After this injection, GVBD occurs earlier than during incubation with progesterone and is accompanied by the formation of the same proteins and of MPF, both suppressed by cycloheximide. The possibility that MPF is an induced protein is then discussed. It has been also found that there is a decrease in membrane permeability, as evidenced by decreased leucine uptake into proteins of oocytes incubated in radioactive amino acids and exposed to progesterone. The same decrease occurs after injection of MPF-containing cytoplasm into oocytes.

4-Chloromercuribenzenesulfonate↗
PubMed1992

Regulation of M-phase progression in Chaetopterus oocytes by protein kinase C.

We have examined the presence of protein kinase C in oocytes of Chaetopterus pergamentaceus and its role in the initiation of germinal vesicle breakdown (GVBD). First, we demonstrated that the oocytes contain a phospholipid- and calcium-dependent protein kinase, protein kinase C (PKC). Since PKC is the primary intracellular receptor for phorbol esters, we tested the ability of phorbol 12,13-dibutyrate (PDBu) to induce GVBD and compared several critical events and processes involved in GVBD induced by PDBu to those induced normally (by seawater). Seawater and 100-200 nM PDBu induced chromosome condensation, spindle formation, and spindle migration over a similar time course. Both treatments induced similar alterations in the SDS-PAGE pattern of newly synthesized proteins. The synthesis of polypeptides of approximately 46 and 54 kDa increased specifically. Both treatments increased oocyte protein phosphorylation, especially of proteins of 22, 32, 46, 55, 64, and 84 kDa. Both treatments resulted in the activation of an M-phase-specific histone H1 kinase activity, which demonstrates the appearance of maturation-promoting factor. Staurosporine, a potent protein kinase C inhibitor, blocked GVBD and the activation of M-phase-specific H1 kinase, whereas HA1004, which preferentially antagonizes protein kinase A, had no effect. The results of this study demonstrate that protein kinase C can activate a wide spectrum of essential biochemical and morphological processes involved in GVBD. Further, these studies suggest that protein kinase C elicits GVBD by activating maturation-promoting factor and support the hypothesis that protein kinase C plays an essential role in oocyte maturation in this species.

Alkaloids↗
PubMed1979

Subunits of RNA polymerase in function and structure. 8. Catalytic properties of self-reactivated core enzyme.

As an attempt to identify the maturation pathway of Escherichia coli RNA polymerase, the catalytic properties of core enzyme reactivated in the absence of maturation-promoting factors (sigma subunit or DNA) (that is, of self-reactivated core enzyme) were compared with those of native core enzyme. Differences have been found in the intrinsic activities such as in the template specificity, Km value of DNA template for the polymerase, activation energy for RNA synthesis, and increment of enzyme activity by sigma subunit. These observations imply that the transcription initiation by self-reactivated core enzyme is inaccurate and, therefore, more strict conditions including the presence of maturation-promoting factors are required for premature core be activated to the genuine function with the transcription specificity of native core enzyme.

DNA-Directed RNA Polymerases↗
PubMed1992

In vitro cell cycle arrest induced by using artificial DNA templates.

In cell extracts of Xenopus eggs which oscillate between S and M phases of the cell cycle, the onset of mitosis is blocked by the presence of incompletely replicated DNA. In this report, we show that several artificial DNA templates (M13 single-stranded DNA and double-stranded plasmid DNA) can trigger this feedback pathway, which inhibits mitosis. Single-stranded M13 DNA is much more effective than double-stranded plasmid DNA at inhibiting the onset of mitosis. Furthermore, we have shown that low levels of M13 single-stranded DNA and high levels of double-stranded plasmid DNA can elevate the tyrosine kinase activity responsible for phosphorylating p34cdc2, thereby inactivating maturation-promoting factor and inhibiting entry into mitosis. This constitutes a simplified system with which to study the signal transduction pathway from the DNA template to the tyrosine kinase responsible for inhibiting p34cdc2 activity.

Animals↗
PubMed1992

Oscillation of MPF is accompanied by periodic association between cdc25 and cdc2-cyclin B.

Activation of maturation-promoting factor at the onset of mitosis requires the tyrosine dephosphorylation of one of its components, the cdc2 protein kinase. cdc25 is the specific tyrosine phosphatase that activates cdc2. We find that Xenopus oocytes contain a relative of cdc25, p72. In Xenopus embryos the abundance of p72 does not oscillate during the cell cycle. However, p72 directly associates with cdc2-cyclin B in a cell cycle-dependent manner, reaching a peak at M phase. The M phase kinase that associates with p72 is catalytically active. These results suggest that the mechanism by which cdc25 triggers cdc2 activation involves a periodic physical association between cdc25 and the cyclin B-cdc2 complex and also that mitotic control can be affected by mechanisms other than transcriptional regulation of the cdc25 gene.

Animals↗
PubMed1992

Mitotic checkpoints.

Entry into mitosis is triggered by activation of maturation promoting factor and a complex of p34cdc2 kinase and cyclin B. Activation induces nuclear lamina breakdown, chromosome condensation and mitotic spindle assembly. Exit from mitosis is initiated by the degradation of cyclin B and the subsequent inactivation of maturation-promoting factor. A more thorough understanding of the checkpoints for initiation of and exit from mitosis has evolved during the past few years.

Animals↗
PubMed1992

Evidence for the regulation of exocytic transport by protein phosphorylation.

We investigated the effects of the protein phosphatase inhibitors okadaic acid and microcystin-LR upon transport of newly synthesized proteins through the exocytic pathway. Treatment of CHO cells with 1 microM okadaic acid rapidly inhibited movement of a marker protein (vesicular stomatitis virus G protein) from the endoplasmic reticulum to the Golgi compartment. Both okadaic acid and microcystin-LR also inhibited transport in an in vitro assay reconstituting movement to the Golgi compartment, at concentrations equivalent to those required to inhibit phosphorylase phosphatase activity. Inhibition both in vivo and in vitro could be antagonized by protein kinase inhibitors, suggesting that protein phosphorylation was directly responsible for this effect. An early stage in the transport reaction associated with vesicle formation or targeting was inhibited by protein phosphorylation, which could be reversed by fractions enriched in protein phosphatase 2A. Protein kinase antagonists did not inhibit transport between sequential compartments of the exocytic pathway in vitro, suggesting that protein phosphorylation is not itself required for vesicular transport. During mitosis, vesicular transport is inhibited simultaneous to the activation of maturation-promoting factor. It is proposed that the inhibition caused by okadaic acid and microcystin-LR involves a similar mechanism to that responsible for the mitotic arrest of vesicular transport.

Adenosine Triphosphate↗
PubMed1992

M-phase-specific histone H1 kinase in fish oocytes. Purification, components and biochemical properties.

We demonstrate, for the first time in fish, that a Ca(2+)-independent and cyclic-nucleotide-independent histone H1 kinase activity oscillates according to the cell cycle of the oocyte, peaking at the first and the second meiotic metaphase with a transient drop between them. The kinase, M-phase-specific histone H1 kinase (M-H1K), was purified from mature carp oocytes by using two exogenous substrates for assaying its activity: histone H1 and a synthetic peptide (SP peptide, KKAAKSPKKAKK) containing the sequence KSPKK, which includes the consensus sequence of the site phosphorylated by a serine/threonine-specific protein kinase encoded by the fission yeast cdc2+ gene (cdc 2 kinase). The M-H1K and maturation-promoting factor (MPF) activities coincided closely throughout four steps of purification, strongly suggesting the identity of M-H1K and MPF. The final preparation was purified 5000-fold with a recovery of 4%, when histone H1 was used for the kinase assay, and 10,000-fold with a recovery of 7% when SP peptide was used. The purified molecular mass of the kinase was estimated to be 100 kDa by gel filtration and contained four proteins of 33, 34, 46 and 48 kDa. Anti-PSTAIR antibody recognizing cdc2 kinase cross-reacted with the 33-kDa and 34-kDa proteins, while the 46-kDa and 48-kDa bands cross-reacted with monoclonal antibodies raised against cyclin B. The 33-kDa protein was also recognized by an antibody against a goldfish cdk2 (Eg1) kinase, a cdc2-related kinase which has the PSTAIR sequence and binds to p13suc1 but does not form a complex with cyclin B. M-H1K activity corresponded well to the 34-kDa, 46-kDa and 48-kDa proteins but not to the 33-kDa protein. These results strongly suggest that M-H1K consists of cdc2 kinase forming a complex with cyclin B, and that cdk2 kinase is not a component of M-H1K, although it is found in the highly purified M-H1K. The purified M-H1K utilized Mg2+, Mn2+, ATP and GTP, and had a wide pH optimum ranging over 8.0-10.5. The kinase was thermolabile and sensitive to freezing/thawing.

Amino Acid Sequence↗
PubMedDate not supplied

Genistein inhibits DNA synthesis but has no effect on levels of DAG and IP3, cell rounding and alkalinization in sulphate-treated Chang liver cells.

Agonist-activated phosphoinositide (PI)-specific phospholipase C initiates PI hydrolysis to produce signals implicated in mitogenic signaling in which the cyclin-dependent cdc2-protein kinase of the maturation-promoting factor is a major protein-tyrosine kinase (PTK) substrate. It has been suggested that PI mitogenic signals are separable into PTK-dependent and non-PTK-dependent by genistein, a tyrosine-specific protein kinase inhibitor. However, we show here that DNA synthesis was abolished in human Chang liver cells although the sulphate-induced PI second messengers, i.e. inositol 1,4,5-trisphosphate and sn-1,2,diacylglycerol, were at equivalent dose-response levels with or without genistein (0.5 mM, 135 microgram/ml). This genistein dosage had been demonstrated to be effective in suppressing tyrosyl phosphorylation in cells. There was no increase in the trypan blue dead cell index. We have shown previously that human Chang cells stimulated by this 'non-growth-factor' agonist, i.e. sulphate, as well as extracellular ATP, became rounded with raised intracellular pH. ATP-induced cell rounding and intracellular alkalinization were not affected by the presence of genistein (0.5 mM). In the present investigation, that genistein dosage had also no effect on these cellular responses when initiated by added sulphate. It seems that the mitogenic signaling function of PI second messengers is dissociable and requires unsuppressed PTK activity.

Adenosine Triphosphate↗
PubMedDate not supplied

Towards understanding the control of the division cycle in animal cells.

The author reviewed the historical process by which classical knowledge of cell division accumulated, to give rise to the molecular biology of the cell cycle, and discussed the perspective of this field of research. The study of the control of cell division began at the turn of the century. It was hypothesized that cell division was a physiological regulation necessary for growing cells to maintain a proper nucleocytoplasmic ratio to survive, which was later substantiated by the finding that amoeba cells could be prevented from dividing by repeated excision of the cytoplasm. However, the observation in Tetrahymena that heat-shocked cells grow exceedingly, but fail to divide, suggested that the cell required the accumulation of a labile "division protein" to initiate division. Mechanisms that control the cell cycle were studied in oocytes by nuclear transplantation and cytoplasmic transfer, and in cultured mammalian cells, protozoa, and Physarum plasmodia by cell fusion. These experiments demonstrated the existence of cytoplasmic factors that control the cell cycle. Maturation promoting factor (MPF) thus discovered in frog oocytes became known to be an ubiquitous cytoplasmic factor that causes the transition from interphase to metaphase in all organisms. The insight into the molecular control of cell growth and division was gained from yeast cell genetics. For biochemical analysis of the cell cycle control, the method to observe the cell cycle in vitro was developed using frog egg extracts. Thus, MPF was identified as a cdc2--cyclin protein complex. Its activity was found to depend on synthesis and phosphorylation of these proteins. However, recently it was found that there were cell cycle phenomena that were difficult to explain in these terms. Various other cellular factors, including nucleocytoplasmic ratio and microtubule assembly, were also found to control MPF, as well as the cell cycle. It remained open to future how these factors control MPF to alter the pattern of the cell cycle.

Animals↗
PubMedDate not supplied

Regulation of the G2-mitosis transition.

The cell cycle is regulated by pathways composed of a dependent series of steps, by timers, and by checkpoint controls which ensure the completion of one event before the initiation of another. This review focuses on the regulation of the initiation of mitosis, with particular emphasis on the regulation of p34cdc2 activity at this point in the cell cycle. The review draws on data from various organisms, but strongly emphasizes the genetic framework as seen in the fission yeast Schizosaccharomyces pombe and the biology and biochemistry of maturation promoting factor in frog oocytes. An attempt is made to include all known genes and proteins where a link can be made to the initiation event. The nutritional size control and its major known controlling elements, the wee1/mik1 protein kinases, and cdc25 protein tyrosine phosphatase are considered in detail along with their regulation. In addition, the checkpoint control pathways which mediate G2 delay in response to failure of DNA replication or DNA damage are examined.

Animals↗
PubMed1992

Sister chromatid separation in frog egg extracts requires DNA topoisomerase II activity during anaphase.

We have produced metaphase spindles and induced them to enter anaphase in vitro. Sperm nuclei were added to frog egg extracts, allowed to replicate their DNA, and driven into metaphase by the addition of cytoplasm containing active maturation promoting factor (MPF) and cytostatic factor (CSF), an activity that stabilizes MPF. Addition of calcium induces the inactivation of MPF, sister chromatid separation and anaphase chromosome movement. DNA topoisomerase II inhibitors prevent chromosome segregation at anaphase, demonstrating that the chromatids are catenated at metaphase and that decatenation occurs at the start of anaphase. Topoisomerase II activity towards exogenous substrates does not increase at the metaphase to anaphase transition, showing that chromosome separation at anaphase is not triggered by a bulk activation of topoisomerase II.

Anaphase↗
PubMed1992

cdc25+ encodes a protein phosphatase that dephosphorylates p34cdc2.

To determine how the human cdc25 gene product acts to regulate p34cdc2 at the G2 to M transition, we have overproduced the full-length protein (cdc25Hs) as well as several deletion mutants in bacteria as glutathione-S-transferase fusion proteins. The wild-type cdc25Hs gene product was synthesized as an 80-kDa fusion protein (p80GST-cdc25) and was judged to be functional by several criteria: recombinant p80GST-cdc25 induced meiotic maturation of Xenopus oocytes in the presence of cycloheximide; p80GST-cdc25 activated histone H1 kinase activity upon addition to extracts prepared from Xenopus oocytes; p80GST-cdc25 activated p34cdc2/cyclin B complexes (prematuration promoting factor) in immune complex kinase assays performed in vitro; p80GST-cdc25 stimulated the tyrosine dephosphorylation of p34cdc2/cyclin complexes isolated from Xenopus oocyte extracts as well as from overproducing insect cells; and p80GST-cdc25 hydrolyzed p-nitrophenylphosphate. In addition, deletion analysis defined a functional domain residing within the carboxy-terminus of the cdc25Hs protein. Taken together, these results suggest that the cdc25Hs protein is itself a phosphatase and that it may function directly in the tyrosine dephosphorylation and activation of p34cdc2 at the G2 to M transition.

Animals↗
PubMed1992

Relocation and distinct subcellular localization of p34cdc2-cyclin B complex at meiosis reinitiation in starfish oocytes.

M phase promoting factor (MPF) is a major element controlling entry into the M phase of the eukaryotic cell cycle. MPF is composed of two subunits, p34cdc2 and cyclin B. Using indirect immunofluorescence staining with specific antibody against starfish cyclin B, we monitored the dynamics of the subcellular distribution of MPF during meiosis reinitiation in starfish oocytes. We found that all of the cyclin B is already associated with p34cdc2 in immature oocytes arrested at the G2/M border and that this inactive complex is present exclusively in the cytoplasm. After its activation, part of the p34cdc2-cyclin B complex moves into the germinal vesicle before nuclear envelope breakdown, independently of either microtubules or actin filaments. Thereafter, some part of the complex accumulates in the nucleolus and condensed chromosomes. Another portion of the complex accumulates on meiotic asters and spindles, while the rest is still present throughout the cytoplasm. As these patterns of localization are detected in the detergent-extracted oocytes, we propose at least four distinct subcellular states of the p34cdc2-cyclin B complex: freely soluble, microtubule-associated, detergent-resistant cytoskeleton-associated and chromosome-associated. Thus, in addition to the intramolecular modification of p34cdc2-cyclin B complex, its intracellular relocation plays a key role in promoting the M phase.

Animals↗