Cell cycle-regulated phosphorylation of the Xenopus polo-like kinase Plx1

J Biol Chem. 2002 Jul 12;277(28):25247-56. doi: 10.1074/jbc.M202855200. Epub 2002 May 6.

Abstract

Polo-like kinases (Plks) control multiple important events during M phase progression, but little is known about their activation during the cell cycle. The activities of both mammalian Plk1 and Xenopus Plx1 peak during M phase, and this activation has been attributed to phosphorylation. However, no phosphorylation sites have previously been identified in any member of the Plk family. Here we have combined tryptic phosphopeptide mapping with mass spectrometry to identify four major phosphorylation sites in Xenopus Plx1. All four sites appear to be phosphorylated in a cell cycle-dependent manner. Phosphorylations at two sites (Ser-260 and Ser-326) most likely represent autophosphorylation events, whereas two other sites (Thr-201 and Ser-340) are targeted by upstream kinases. Several recombinant kinases were tested for their ability to phosphorylate Plx1 in vitro. Whereas xPlkk1 phosphorylated primarily Thr-10, Thr-201 was readily phosphorylated by protein kinase A, and Cdk1/cyclin B was identified as a likely kinase acting on Ser-340. Phosphorylation of Ser-340 was shown to be responsible for the retarded electrophoretic mobility of Plx1 during M phase, and phosphorylation of Thr-201 was identified as a major activating event.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Cycle Proteins
  • Cell Cycle*
  • Evolution, Molecular
  • Mass Spectrometry
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Peptide Mapping
  • Phosphorylation
  • Protein Serine-Threonine Kinases / chemistry
  • Protein Serine-Threonine Kinases / genetics
  • Protein Serine-Threonine Kinases / metabolism*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Serine / metabolism
  • Spodoptera
  • Threonine / metabolism
  • Xenopus
  • Xenopus Proteins*

Substances

  • Cell Cycle Proteins
  • Recombinant Proteins
  • Xenopus Proteins
  • Threonine
  • Serine
  • Plk1 protein, Xenopus
  • Protein Serine-Threonine Kinases