Mitotic chromosomes are constrained by topoisomerase II-sensitive DNA entanglements

J Cell Biol. 2010 Mar 8;188(5):653-63. doi: 10.1083/jcb.200910085. Epub 2010 Mar 1.


We have analyzed the topological organization of chromatin inside mitotic chromosomes. We show that mitotic chromatin is heavily self-entangled through experiments in which topoisomerase (topo) II is observed to reduce mitotic chromosome elastic stiffness. Single chromosomes were relaxed by 35% by exogenously added topo II in a manner that depends on hydrolysable adenosine triphosphate (ATP), whereas an inactive topo II cleavage mutant did not change chromosome stiffness. Moreover, experiments using type I topos produced much smaller relaxation effects than topo II, indicating that chromosome relaxation by topo II is caused by decatenation and/or unknotting of double-stranded DNA. In further experiments in which chromosomes are first exposed to protease to partially release protein constraints on chromatin, ATP alone relaxes mitotic chromosomes. The topo II-specific inhibitor ICRF-187 blocks this effect, indicating that it is caused by endogenous topo II bound to the chromosome. Our experiments show that DNA entanglements act in concert with protein-mediated compaction to fold chromatin into mitotic chromosomes.

Publication types

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

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Animals
  • Antigens, Neoplasm / genetics
  • Antigens, Neoplasm / metabolism*
  • Chromosomes / chemistry*
  • Chromosomes / genetics
  • Chromosomes / metabolism*
  • DNA / chemistry
  • DNA / genetics
  • DNA / metabolism*
  • DNA Topoisomerases, Type II / genetics
  • DNA Topoisomerases, Type II / metabolism*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Humans
  • Isoenzymes / genetics
  • Isoenzymes / metabolism
  • Mitosis / physiology*
  • Nucleic Acid Conformation*
  • Stress, Mechanical
  • Trypsin / metabolism


  • Antigens, Neoplasm
  • DNA-Binding Proteins
  • Isoenzymes
  • Adenosine Triphosphate
  • DNA
  • Trypsin
  • DNA Topoisomerases, Type II