Mitotic phosphorylation of DNA topoisomerase II alpha by protein kinase CK2 creates the MPM-2 phosphoepitope on Ser-1469

J Biol Chem. 2000 Nov 3;275(44):34710-8. doi: 10.1074/jbc.M005179200.

Abstract

DNA topoisomerase II alpha is required for chromatin condensation during prophase. This process is temporally linked with the appearance of mitosis-specific phosphorylation sites on topoisomerase IIalpha including one recognized by the MPM-2 monoclonal antibody. We now report that the ability of mitotic extracts to create the MPM-2 epitope on human topoisomerase II alpha is abolished by immunodepletion of protein kinase CK2. Furthermore, the MPM-2 phosphoepitope on topoisomerase II alpha can be generated by purified CK2. Phosphorylation of C-truncated topoisomerase II alpha mutant proteins conclusively shows, that the MPM-2 epitope is present in the last 163 amino acids. Use of peptides containing all conserved CK2 consensus sites in this region indicates that only the peptide containing Arg-1466 to Ala-1485 is able to compete with topoisomerase II alpha for binding of the MPM-2 antibody. Replacement of Ser-1469 with Ala abolishes the ability of the phosphorylated peptide to bind to the MPM-2 antibody while a peptide containing phosphorylated Ser-1469 binds tightly. Surprisingly, the MPM-2 phosphoepitope influences neither the catalytic activity of topoisomerase II alpha nor its ability to form molecular complexes with CK2 in vitro. In conclusion, we have identified protein kinase CK2 as a new MPM-2 kinase able to phosphorylate an important mitotic protein, topoisomerase II alpha, on Ser-1469.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antigens, Neoplasm
  • Casein Kinase II
  • Catalysis
  • Cell Cycle Proteins*
  • Cell Extracts
  • Chromosomes, Human
  • DNA Topoisomerases, Type II* / chemistry
  • DNA Topoisomerases, Type II* / metabolism*
  • DNA-Binding Proteins
  • Guanosine Triphosphate / metabolism
  • HeLa Cells
  • Heparin / metabolism
  • Humans
  • Isoenzymes / antagonists & inhibitors
  • Isoenzymes / chemistry
  • Isoenzymes / metabolism*
  • Kinesin
  • Mitosis*
  • Molecular Sequence Data
  • Phosphoproteins / metabolism*
  • Phosphorylation
  • Protein-Serine-Threonine Kinases / metabolism*
  • Sequence Homology, Amino Acid
  • Serine / metabolism*
  • Topoisomerase II Inhibitors

Substances

  • Antigens, Neoplasm
  • Cell Cycle Proteins
  • Cell Extracts
  • DNA-Binding Proteins
  • Isoenzymes
  • Phosphoproteins
  • Topoisomerase II Inhibitors
  • Serine
  • Guanosine Triphosphate
  • Heparin
  • Casein Kinase II
  • Protein-Serine-Threonine Kinases
  • KIF20B protein, human
  • Kinesin
  • DNA Topoisomerases, Type II