Identification of a new APC/C recognition domain, the A box, which is required for the Cdh1-dependent destruction of the kinase Aurora-A during mitotic exit

Genes Dev. 2002 Sep 1;16(17):2274-85. doi: 10.1101/gad.1007302.

Abstract

The mitotic kinase Aurora A (Aur-A) is required for formation of a bipolar mitotic spindle and accurate chromosome segregation. In somatic cells, Aur-A protein and kinase activity levels peak during mitosis, and Aur-A is degraded during mitotic exit. Here, we investigated how Aur-A protein and kinase activity levels are regulated, taking advantage of the rapid synchronous cell division cycles of Xenopus eggs and cell-free systems derived from them. Aur-A kinase activity oscillates in the early embryonic cell cycles, just as in somatic cells, but Aur-A protein levels are constant, indicating that regulated activation and inactivation, instead of periodic proteolysis, is the dominant mode of Aur-A regulation in these cell cycles. Cdh1, the APC/C activator that targets many mitotic proteins for ubiquitin-dependent proteolysis during late mitosis and G1 in somatic cells, is missing in Xenopus eggs and early embryos. We find that addition of Cdh1 to egg extracts undergoing M phase exit is sufficient to induce rapid degradation of Aur-A. Aur-A contains both of the two known APC/C recognition signals, (1) a C-terminal D box similar to those required for ubiquitin-dependent destruction of cyclin B and several other mitotic proteins, and (2) an N-terminal KEN box similar to that found on cdc20, which is ubiquitinated in response to APC/C(Cdh1). The D box is required for Cdh1-induced destruction of Aur-A but the KEN box is not. Destruction also requires a short region in the N terminus, which contains a newly identified recognition signal, the A box. The A box is conserved in vertebrate Aur-As and contains serine 53, which is phosphorylated during M phase. Mutation of serine 53 to aspartic acid, which can mimic the effect of phosphorylation, completely blocks Cdh1-dependent destruction of Aur-A. These results suggest that dephosphorylation of serine 53 during mitotic exit could control the timing of Aur-A destruction, allowing recognition of both the A box and D box by Cdh1-activated APC/C.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 3T3 Cells
  • Amino Acid Sequence
  • Anaphase-Promoting Complex-Cyclosome
  • Animals
  • Aurora Kinase A
  • Aurora Kinases
  • Binding Sites
  • Cell Cycle Proteins
  • Conserved Sequence
  • Female
  • Fertilization / physiology
  • In Vitro Techniques
  • Ligases / metabolism*
  • Male
  • Mice
  • Mitosis / physiology*
  • Molecular Sequence Data
  • Mutation
  • Oocytes / cytology
  • Oocytes / metabolism
  • Protein Kinases / chemistry
  • Protein Kinases / genetics
  • Protein Kinases / metabolism*
  • Protein Structure, Tertiary
  • Protein-Serine-Threonine Kinases
  • Sequence Deletion
  • Sequence Homology, Amino Acid
  • Ubiquitin-Protein Ligase Complexes*
  • Xenopus
  • Xenopus Proteins

Substances

  • Cell Cycle Proteins
  • Xenopus Proteins
  • Ubiquitin-Protein Ligase Complexes
  • Anaphase-Promoting Complex-Cyclosome
  • Protein Kinases
  • AURKA protein, Xenopus
  • Aurka protein, mouse
  • Aurora Kinase A
  • Aurora Kinases
  • Protein-Serine-Threonine Kinases
  • Ligases