Biology of the oocyte

The last step of oogenesis also called meiotic maturation is a key event for embryonic development and ensures the formation of a haploid germ cell ready to be fertilized.

In the ovary, oocytes are arrested in prophase of the first meiotic division. At time of ovulation, oocytes reenter meiosis upon hormonal stimulation and reach the second meiotic division without an intervening S phase. Oocytes then become arrested a second time in Metaphase II awaiting for fertilization. This process relies on the activation of MPF (Cdk1-CyclinB complexes) and recapitulates all the biochemical mechanisms regulating the G2-M transition of the cell cycle.

Our team is interested on studying three steps of meiotic maturation using Xenopus oocyte:

  • What regulate the activation of MPF in response to steroid hormones? Our researches are mainly focused on the respective contribution of kinases (PKA, Cdk1 and MAPK) and phosphatases (PP1 and PP2A)
  • What is the molecular mechanism inhibiting DNA replication during the meiosis I-meiosis II transition; this inhibition being essential for the formation of a haploid gamete?
  • Which mechanisms arrest the oocyte in metaphase II?

Understanding the cell cycle regulation in meiosis is not only critical for reproduction and early development, it also helps us to decipher the process of tumoregenesis as cancer often results from the misregulation of cell division.

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The ultimate step of oogenesis, meiotic maturation, is a key event for embryonic development. It occurs at time of ovulation and ensures the formation of a haploid gamete able to be fertilized. During this step, oocytes that are arrested in prophase of the 1st meiotic division, reenter meiosis upon hormonal stimulation: they complete the 1st meiotic division and reach the metaphase of the 2nd one, where they arrest, awaiting fertilization. This process relies on the activation of MPF (M-phase promoting factor, Cdk1-Cyclin B complex). Hence, meiotic maturation has been long proven to be a powerful model system to study the regulation of the M-phase of the cell cycle.

Our team focuses on three steps of meiotic maturation, using Xenopus oocytes as a model:

  • What does regulate the activation of MPF in response to steroid hormones? We investigate the respective contribution of kinases (PKA, Cdk1 and MAPK) and phosphatases (PP1 and PP2A) in the transduction pathway culminating in MPF activation.
  • What is the molecular mechanism inhibiting DNA replication during the meiosis I-meiosis II transition, an essential process for the formation of a haploid genome?
  • What are the molecular mechanisms arresting the oocyte at metaphase II?

Our studies contribute to the understanding of the regulation of a crucial step of cell cycle control, the G2-M transition, as well as hormonal regulation and signal transduction, increasing our knowledge of fertility, reproduction and development.

Highlights

The prophase arrest is controlled by a high activity of PKA. Meiosis resumption relies on a drop in PKA activity, hence the dephosphorylation of a critical PKA substrate. This step leads to the further activation of MPF and the Mos/MAPK cascade that induce the completion of meiotic division. Our recent results revealed that:

  • the mechanisms of steroid production controlling the induction of meiosis (Haccard et al. Mol. Cell Endocr. 2012).
  • the critical event in meiotic resumption is a change in the balance between Myt1 activity and Cyclin B synthesis (Gaffre et al. Development 2011).
  • Greatwall positively regulates meiotic divisions through the inhibition of PP2A (Dupre et al. J. Cell Sci. 2013).
  • the critical PKA substrate is ARPP19. Once phosphorylated by PKA, it inhibits MPF activation and maintains the prophase block. Its dephosphorylation is needed to overcome this blockade. More, ARPP19 is later phosphorylated by Greatwall and converted into an essential positive regulator of MPF (Dupre et al. Nature Commun. 2014).
  • the default fate of an unfertilized oocyte is to die by apoptosis (Du Pasquier et al. PLoS ONE 2011).

Future directions

Information about the cell cycle molecular machinery came from pioneer experiments combining genetic and biochemical studies performed in yeast and oocytes, providing the identification of MPF, responsible for driving cell division. However, important points, such as the mechanisms underlying entry into M-phase, remain ill-defined. Based on the conserved nature of cell cycle components and since meiotic and mitotic divisions share identical molecular players, we chose a powerful physiological model system: meiotic maturation of Xenopus oocyte, to cover this lack of knowledge.

Our project focuses on two questions:

  • Deciphering the molecular connections between two major players controlling meiotic divisions of all vertebrate oocytes, PKA and phosphatase 2A. Specific emphasis will be paid to how ARPP19 phosphorylated by PKA prevents the initial activation of MPF and the interplay between PKA- and Greatwall-phosphorylations that shift the protein from an inhibitor towards an activator of M-phase.
  • Unraveling the molecular mechanisms repressing DNA replication between the two meiotic divisions. We will investigate the implications of the MAPK pathway, Cdc6 protein and the ATM/ATR kinases in this process.

Collaborations

  • Dr. Evelyn Houliston, UPMC-CNRS UMR7009-Biologie du Développement, 06230 Villefranche sur mer, France
  • Dr. Tim. Hunt, Prix nobel de Medecine (2001), Cancer Research UK, Clare Hall Laboratories,South Mimms,Herts EN6 3LD, England.
  • Dr. Michael Goldberg, Department of Molecular Biology and Genetics, Biotechnology Building, Cornell University, Ithaca, NY 14853-2703
  • Dr. Angu Nairn, Department of Psychiatry, Yale University School of Medicine, New Haven, Connecticut 06508, USA.
  • Dr. Thierry Lorca, Universités Montpellier 2 et 1, Centre de Recherche de Biochimie Macromoléculaire, CNRS UMR 5237, IFR 122, 1919 Route de Mende, 34293 Montpellier cedex 5, France
  • Dr. Jean Gautier Columbia University, Department of Genetics and Development, 1130 St. Nicholas Avenue, ICRC Room 603A, New York NY 10032.
  • Dr. Mathieu Bollen, Laboratory of Biosignaling & Therapeutics, Department of Cellular and Molecular Medicine, University of Leuven, B-3000 Leuven, Belgium