Changes in protein phosphorylation accompanying maturation of Xenopus laevis oocytes

doi:10.1016/0012-1606(77)90093-8

Developmental Biology

Volume 58, Issue 2, 15 July 1977, Pages 295-312

https://doi.org/10.1016/0012-1606(77)90093-8 Get rights and content

Abstract

Protein phosphorylation has been measured after injection of [³²P]phosphate into oocytes of Xenopus laevis undergoing progesterone-induced meiotic maturation. As oocytes mature, there is a burst of nonyolk protein phosphorylation several hours after progesterone exposure and shortly before germinal vesicle breakdown (GVBD). This burst is not due to changes in the specific activity of the phosphate or ATP pool. Enucleated oocytes exposed to progesterone also experience the burst, indicating the cytoplasmic location of phosphoprotein formation. When an oocyte receives an injection of cytoplasm containing the maturation-promoting factor (MPF), a burst of protein phosphorylation occurs immediately, and GVBD occurs shortly thereafter, even in the presence of cycloheximide. Under a variety of conditions promoting or blocking maturation, oocytes which undergo GVBD are the only ones to have experienced the phosphorylation burst. The results suggest that the protein phosphorylation burst is a necessary step in the mechanism by which MPF promotes GVBD.

References (55)

S.E. Allerton et al.
Chemical characterization of the phosphoprotein phosvitin
J. Biol. Chem
(1965)
E. Baltus et al.
Cytochemical and biochemical studies on progesterone-induced maturation in amphibian oocytes. I. Ribonucleic acid and protein synthesis (effects of inhibitors and of a “maturation promoting factor”)
Differentiation
(1973)
E.W. Bergink et al.
Precursor-product relationship between amphibian vitellogenin and the yolk proteins, lipovitellin and phosvitin
J. Biol. Chem
(1974)
D.B. Bylund et al.
Decomposition of phosphoserine and phosphothreonine during acid hydrolysis
Anal. Biochem
(1976)
J.A. Cutting et al.
Staining of phosphoproteins on acrylamide gel electropherograms
Anal. Biochem
(1973)
R. Delange et al.
Activation of skeletal muscle phosphorylase kinase by adenosine triphosphate and adenosine 3′5′-monophosphate
J. Biol. Chem
(1968)
R.E. Ecker et al.
Protein synthesis in amphibian oocytes and early embryos
Develop. Biol
(1968)
C.M. Feldherr
The uptake of endogenous proteins by oocyte nuclei
Exp. Cell Res
(1975)
L.R. Gurley et al.
Sequential phosphorylation of histone subfractions in the Chinese hamster cell cycle
J. Biol. Chem
(1975)
U.E. Handschin et al.
Spectrophotometric determination of fluorophor, protein, and fluorophor/protein ratios in fluorescamine and MDPF fluorescent antibody conjugates
Anal. Biochem
(1976)

D.W. Jared et al.

Distribution of incorporated and synthesized protein among cell fractions of Xenopus oocytes

Develop. Biol

(1973)

E.G. Krebs et al.

Phosphorylase b kinase from rabbit skeletal muscle

O.H. Lowry et al.

Protein measurement with the Folin phenol reagent

J. Biol. Chem

(1951)

J. Maller et al.

Spindle formation and cleavage in Xenopus eggs injected with centriole-containing fractions from sperm

Exp. Cell Res

(1976)

R.W. Merriam

Progesterone-induced maturational events in oocytes of Xenopus laevis. I. Continuous necessity for diffusable calcium and magnesium

Exp. Cell Res

(1971)

R.W. Merriam

Progesterone-induced maturational events in oocytes of Xenopus laevis. II. Changes in intracellular calcium and magnesium distribution at germinal vesicle breakdown

Exp. Cell Res

(1971)

P. Pennequin et al.

Decreased uptake of [³H] leucine during progesterone-induced maturation of Xenopus laevis oocytes

FEBS Lett

(1975)

E. Randerath et al.

Resolution of complex nucleotide mixtures by two-dimensional anion-exchange thin-layer chromatography

J. Chromatogr

(1964)

R. Rangel-Aldao et al.

Dissociation and reassociation of phosphorylated and non-phosphorylated forms of the adenosine 3′5′-monophosphate-dependent protein kinase from bovine cardiac muscle

J. Biol. Chem

(1976)

J.K. Reynhout et al.

Studies on the appearance and nature of a maturation-inducing factor in the cytoplasm of amphibian oocytes exposed to progesterone

Develop. Biol

(1974)

O.M. Rosen et al.

Reversible autophosphorylation of a cyclic 3′-5′ AMP-dependent protein kinase from bovine cardiac muscle

J. Biol. Chem

(1975)

H. Sanui

Measurement of inorganic orthophosphate in biological materials: Extraction properties of butyl acetate

Anal. Biochem

(1974)

N.K. Schaffer et al.

Serine phosphoric acid from diisopropylphosphoryl chymotrypsin

J. Biol. Chem

(1953)

G. Schmidt et al.

A method for the determination of deoxyribonucleic acid, ribonucleic acid and phosphoproteins in animal tissues

J. Biol. Chem

(1945)

S. Schorderet-Slatkine et al.

Progesterone-induced maturation in oocytes of Xenopus laevis

Appearance of a “maturation promoting factor” in enucleated oocytes

Cell Differ

(1973)

L.D. Smith

Molecular events during oocyte maturation

L.D. Smith et al.

Role of the oocyte nucleus in physiological maturation in Rana pipiens

(1969)

Cited by (175)

Cell cycle control in the early embryonic development of aquatic animal species
2015, Comparative Biochemistry and Physiology Part - C: Toxicology and Pharmacology
Citation Excerpt :
This result was repeated in the frog X. laevis, and through microinjection experiments in these two species, it was demonstrated that MPF activity is also present in the cytoplasm of mitotic cells, and it oscillates during the cell cycle, peaking in metaphase and disappearing with the completion of mitosis (Sunkara et al., 1979; Gerhart et al., 1984). MPF was shown to be an enzymatic activity that causes an increase in overall protein phosphorylation (Maller et al., 1977). We now know that MPF is a Cyclin-dependent kinase (CDK), which is composed of the Cdc2 catalytic subunit and Cyclin B regulatory subunit.
The cell cycle is integrated with many aspects of embryonic development. Not only is proper control over the pace of cell proliferation important, but also the timing of cell cycle progression is coordinated with transcription, cell migration, and cell differentiation. Due to the ease with which the embryos of aquatic organisms can be observed and manipulated, they have been a popular choice for embryologists throughout history. In the cell cycle field, aquatic organisms have been extremely important because they have played a major role in the discovery and analysis of key regulators of the cell cycle. In particular, the frog Xenopus laevis has been instrumental for understanding how the basic embryonic cell cycle is regulated. More recently, the zebrafish has been used to understand how the cell cycle is remodeled during vertebrate development and how it is regulated during morphogenesis. This review describes how some of the unique strengths of aquatic species have been leveraged for cell cycle research and suggests how species such as Xenopus and zebrafish will continue to reveal the roles of the cell cycle in human biology and disease.
Pioneering the xenopus oocyte and egg extract system
2012, Journal of Biological Chemistry
Citation Excerpt :
The tight correlation between increased phosphorylation and MPF and M phase was confirmed in a variety of other systems by several laboratories. Therefore, the main conclusion of this work and of my thesis was that MPF might be a protein kinase or an activator of a protein kinase (6). In considering where to undertake postdoctoral training, I believed it was important to continue pioneering the Xenopus oocyte and egg system and to develop further the possible link between protein phosphorylation and MPF.
Phosphatases driving mitosis: Pushing the gas and lifting the brakes
2012, Progress in Molecular Biology and Translational Science
Citation Excerpt :
These proteins were shown to be phosphorylated in response to an activity known as M-phase promoting factor (MPF), leading researchers to hypothesize that MPF-driven protein phosphorylation was controlling the key events of mitosis. It appeared from this work that MPF might contain a kinase activity, but whether MPF was directly phosphorylating mitotic phosphoproteins or might be activating secondary kinases was not fully settled until MPF was characterized molecularly; purified, active MPF was shown to contain a protein kinase catalytic subunit, now known as Cdk1, and an activating subunit, Cyclin B.2–9 It was determined that active MPF could both directly and indirectly (through the activation of mitotic kinases) phosphorylate proteins involved in mitotic progression, and it became clear that the activation of Cdk1 kinase activity was a central event in the initiation and maintenance of mitosis. Work over the next decade aimed to understand the precise mechanisms regulating MPF activation and focused primarily on kinases and phosphatases acting upstream of the Cdk1/Cyclin B complex.
Entry into and progression through mitosis depends critically on the establishment and maintenance of protein phosphorylation. For this reason, studies on mitotic progression have focused heavily on the activation of MPF (M phase promoting factor), a cyclin-dependent kinase responsible for phosphorylating proteins that execute the dynamic events of mitosis. Recent work, however, has significantly expanded our understanding of mechanisms that allow accumulation of phosphoproteins at M phase, suggesting that mitotic entry relies not only on MPF activation but also on the inhibition of antimitotic phosphatases. It is now clear that there exists a separate, albeit equally important, signaling pathway for the inactivation of protein phosphatases at the G2/M transition. This pathway, which is governed by the kinase Greatwall is essential for both entry into and maintenance of M phase. This chapter will outline the molecular events regulating entry into mitosis, specifically highlighting the role that protein phosphorylation plays in triggering both MPF activation and the inhibition of phosphatase activity that would otherwise prevent accumulation of mitotic phosphoproteins. These intricate regulatory pathways are essential for maintaining normal cell division and preventing inappropriate cell proliferation, a central hallmark of cancer cells.
Cdc2-cyclin B triggers H3 kinase activation of Aurora-A in Xenopus oocytes
2003, Journal of Biological Chemistry
Citation Excerpt :
Aurora-A Phosphorylation and Activation Are under the Control of cAMP and Protein Synthesis—Aurora-A phosphorylation and H3 kinase activation take place in response to progesterone at GVBD time. It is well established that progesterone induces an early drop in cAMP and the synthesis of new proteins, both events being required for Cdc2 activation and GVBD (42, 43). To ascertain whether both events are necessary for Aurora-A activation, prophase-arrested oocytes were incubated in the presence of either cycloheximide (CHX), a protein synthesis inhibitor, or IBMX, a phosphodiesterase inhibitor that maintains a high level of cAMP in the cell.
Xenopus oocytes are arrested in meiotic prophase I and resume meiotic divisions in response to progesterone. Progesterone triggers activation of M-phase promoting factor (MPF) or Cdc2-cyclin B complex and neosynthesis of Mos kinase, responsible for MAPK activation. Both Cdc2 and MAPK activities are required for the success of meiotic maturation. However, the signaling pathway induced by progesterone and leading to MPF activation is poorly understood, and most of the targets of both Cdc2 and MAPK in the oocyte remain to be determined. Aurora-A is a Ser/Thr kinase involved in separation of centrosomes and in spindle assembly during mitosis. It has been proposed that in Xenopus oocytes Aurora-A could be an early component of the progesterone-transduction pathway, acting through the regulation of Mos synthesis upstream Cdc2 activation. We addressed here the question of Aurora-A regulation during meiotic maturation by using new in vitro and in vivo experimental approaches. We demonstrate that Cdc2 kinase activity is necessary and sufficient to trigger both Aurora-A phosphorylation and kinase activation in Xenopus oocyte. In contrast, these events are independent of the Mos/MAPK pathway. Aurora-A is phosphorylated in vivo at least on three residues that regulate differentially its kinase activity. Therefore, Aurora-A is under the control of Cdc2 in the Xenopus oocyte and could be involved in meiotic spindle establishment.
From oocyte maturation to the in vitro cell cycle: The history of discoveries of Maturation-Promoting Factor (MPF) and Cytostatic Factor (CSF)
2001, Differentiation
This article briefly reviews the classical cell cycle studies using oocytes and zygotes of mainly amphibians in the past century. The discussions are focused on the investigations into the cytoplasmic factors that regulate meiosis during oocyte maturation and the initiation of mitosis during fertilisation, which were carried out in the author's lab between 1967 and 1987. This chronicle traces the development of the problems and the direction in which their solutions were attempted in the course of these investigations. The author tries to answer the following questions: why he decided to study oocyte maturation, how he discovered progesterone as a maturation-inducing hormone, how he discovered and characterised the cytoplasmic regulators of the cell cycle, Maturation-Promoting Factor (MPF) and Cyto-Static Factor (CSF), and how he invented the method of observing cell cycle processes in a cytoplasmic extract in vitro.
Recurring themes in oocyte maturation
1998, Biology of the Cell
I am pleased to contribute to this special issue of Biology of the Cell in honor of Yoshio Masui. Oocyte maturation remains a small enough and young enough field that the authors assembled for this issue can trace the entire development of the field over the last 30 years, beginning with the early demonstration by Masui (1967) and others that steroids can induce complete maturation of denuded oocytes in vitro. Not very long after that Masui and Markert (1971) published the seminal paper that identified MPF and CSF activity. In this article I intend to highlight recurring themes in oocyte maturation that continue to be actively investigated, almost all of which derive from studies pursued at some point in time by Masui and his colleagues.

View all citing articles on Scopus

^☆: Portions of this work were submitted by J. Maller in partial fulfillment of the requirements for the Ph.D. degree, University of California, Berkeley, California, 1974.

^☆☆: This research was supported by United States Public Health Service Grant No. GM19363.

View full text

Full paperChanges in protein phosphorylation accompanying maturation of Xenopus laevis oocytes☆,☆☆

Abstract

J. Biol. Chem

Differentiation

J. Biol. Chem

Anal. Biochem

Anal. Biochem

J. Biol. Chem

Develop. Biol

Exp. Cell Res

J. Biol. Chem

Anal. Biochem

Develop. Biol

J. Biol. Chem

Exp. Cell Res

Exp. Cell Res

Exp. Cell Res

FEBS Lett

J. Chromatogr

J. Biol. Chem

Develop. Biol

J. Biol. Chem

Anal. Biochem

J. Biol. Chem

J. Biol. Chem

Cell Differ

Role of the oocyte nucleus in physiological maturation in Rana pipiens

Full paper
Changes in protein phosphorylation accompanying maturation of Xenopus laevis oocytes☆,☆☆