A novel pocket in 14-3-3epsilon is required to mediate specific complex formation with cdc25C and to inhibit cell cycle progression upon activation of checkpoint pathways

Exp Cell Res. 2009 May 1;315(8):1448-57. doi: 10.1016/j.yexcr.2009.01.018. Epub 2009 Jan 31.

Abstract

Mitotic progression requires the activity of the dual specificity phosphatase, cdc25C. Cdc25C function is inhibited by complex formation with two 14-3-3 isoforms, 14-3-3epsilon and 14-3-3gamma. To understand the molecular basis of specific complex formation between 14-3-3 proteins and their ligands, chimeric 14-3-3 proteins were tested for their ability to form a complex with cdc25C in vivo. Specific complex formation between cdc25C and 14-3-3epsilon in vivo requires a phenylalanine residue at position 135 (F135) in 14-3-3epsilon. Mutation of this residue to the corresponding residue present in other 14-3-3 isoforms (F135V) leads to reduced binding to cdc25C and a decrease in the ability to inhibit cdc25C function in vivo. Similarly, F135V failed to rescue the incomplete S phase and the G2 DNA damage checkpoint defects observed in cells lacking 14-3-3epsilon. A comparative analysis of the 14-3-3 structures present in the database suggested that the F135 in 14-3-3epsilon was required to maintain the integrity of a pocket that might be involved in secondary interactions with cdc25C. These results suggest that the specificity of the 14-3-3 ligand interaction may be dependent on structural motifs present in the individual 14-3-3 isoforms.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 14-3-3 Proteins / chemistry
  • 14-3-3 Proteins / genetics*
  • 14-3-3 Proteins / metabolism
  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Cell Cycle Proteins / metabolism*
  • Humans
  • Mice
  • Models, Molecular
  • Molecular Sequence Data
  • Phylogeny
  • Protein Binding
  • Protein Structure, Tertiary
  • Rats
  • Sequence Alignment
  • Signal Transduction
  • cdc25 Phosphatases / metabolism*

Substances

  • 14-3-3 Proteins
  • Cell Cycle Proteins
  • cdc25 Phosphatases