Nek9 regulates spindle organization and cell cycle progression during mouse oocyte meiosis and its location in early embryo mitosis

Cell Cycle. 2012 Dec 1;11(23):4366-77. doi: 10.4161/cc.22690. Epub 2012 Nov 16.

Abstract

Nek9 (also known as Nercc1), a member of the NIMA (never in mitosis A) family of protein kinases, regulates spindle formation, chromosome alignment and segregation in mitosis. Here, we showed that Nek9 protein was expressed from germinal vesicle (GV) to metaphase II (MII) stages in mouse oocytes with no detectable changes. Confocal microscopy identified that Nek9 was localized to the spindle poles at the metaphase stages and associated with the midbody at anaphase or telophase stage in both meiotic oocytes and the first mitotic embyros. Depletion of Nek9 by specific morpholino injection resulted in severely defective spindles and misaligned chromosomes with significant pro-MI/MI arrest and failure of first polar body (PB1) extrusion. Knockdown of Nek9 also impaired the spindle-pole localization of γ-tubulin and resulted in retention of the spindle assembly checkpoint protein Bub3 at the kinetochores even after 10 h of culture. Live-cell imaging analysis also confirmed that knockdown of Nek9 resulted in oocyte arrest at the pro-MI/MI stage with abnormal spindles, misaligned chromosomes and failed polar body emission. Taken together, our results suggest that Nek9 may act as a MTOC-associated protein regulating microtubule nucleation, spindle organization and, thus, cell cycle progression during mouse oocyte meiotic maturation, fertilization and early embryo cleavage.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antineoplastic Agents, Phytogenic / pharmacology
  • Cell Cycle Proteins / metabolism
  • Chromosomal Proteins, Non-Histone
  • Chromosome Segregation
  • Chromosomes / metabolism
  • Female
  • Kinetochores / metabolism
  • M Phase Cell Cycle Checkpoints
  • Meiosis
  • Mice
  • Mice, Inbred ICR
  • Mitosis
  • Morpholinos / pharmacology
  • NIMA-Related Kinases
  • Nocodazole / pharmacology
  • Oocytes / drug effects
  • Oocytes / growth & development
  • Oocytes / metabolism*
  • Paclitaxel / pharmacology
  • Poly-ADP-Ribose Binding Proteins
  • Protein-Serine-Threonine Kinases / antagonists & inhibitors
  • Protein-Serine-Threonine Kinases / genetics
  • Protein-Serine-Threonine Kinases / metabolism*
  • Spindle Apparatus / metabolism*
  • Tubulin / metabolism

Substances

  • Antineoplastic Agents, Phytogenic
  • Bub3 protein, mouse
  • Cell Cycle Proteins
  • Chromosomal Proteins, Non-Histone
  • Morpholinos
  • Poly-ADP-Ribose Binding Proteins
  • Tubulin
  • NIMA-Related Kinases
  • Nek9 protein, mouse
  • Protein-Serine-Threonine Kinases
  • Paclitaxel
  • Nocodazole