DNA Replication Checkpoint Prevents Precocious Chromosome Segregation by Regulating Spindle Behavior

Mol Cell. 2004 Dec 3;16(5):687-700. doi: 10.1016/j.molcel.2004.11.001.

Abstract

The DNA replication checkpoint maintains replication fork integrity and prevents chromosome segregation during replication stresses. Mec1 and Rad53 (human ATM/ATR- and Chk2-like kinases, respectively) are critical effectors of this pathway in yeast. When treated with replication inhibitors, checkpoint-deficient mec1 or rad53 mutant fails to maintain replication fork integrity and proceeds to partition unreplicated chromosomes. We show that this unnatural chromosome segregation requires neither the onset of mitosis nor APC activation, cohesin cleavage, or biorientation of kinetochores. Instead, the checkpoint deficiency leads to deregulation of microtubule-associated proteins Cin8 and Stu2, which, in the absence of both chromosome cohesion and bipolar attachment of kinetochores to microtubules, induce untimely spindle elongation, causing premature chromosome separation. The checkpoint's ability to prevent nuclear division is abolished by combined deficiency of microtubule-destabilizing motor Kip3 and Mad2 functions. Thus, the DNA replication checkpoint prevents precocious chromosome segregation, not by inhibiting entry into mitosis as widely believed, but by directly regulating spindle dynamics.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blotting, Northern
  • Blotting, Southern
  • Blotting, Western
  • Carrier Proteins / metabolism
  • Cell Cycle Proteins / metabolism
  • Cell Nucleus / metabolism
  • Checkpoint Kinase 2
  • Chromosome Segregation
  • Chromosomes / ultrastructure*
  • DNA / metabolism
  • DNA Replication*
  • Flow Cytometry
  • G1 Phase
  • Hydroxyurea / pharmacology
  • Intracellular Signaling Peptides and Proteins
  • Kinesin
  • Kinetochores / metabolism
  • Mad2 Proteins
  • Microscopy, Fluorescence
  • Microtubule-Associated Proteins / metabolism
  • Microtubules / metabolism
  • Mitosis
  • Models, Biological
  • Mutation
  • Nuclear Proteins
  • Plasmids / metabolism
  • Protein-Serine-Threonine Kinases / metabolism
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae / physiology*
  • Saccharomyces cerevisiae Proteins / metabolism
  • Spindle Apparatus*
  • Temperature
  • Time Factors
  • Up-Regulation

Substances

  • CIN8 protein, S cerevisiae
  • Carrier Proteins
  • Cell Cycle Proteins
  • Intracellular Signaling Peptides and Proteins
  • KIP3 protein, S cerevisiae
  • MAD2 protein, S cerevisiae
  • Mad2 Proteins
  • Microtubule-Associated Proteins
  • Nuclear Proteins
  • STU2 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • DNA
  • Checkpoint Kinase 2
  • MEC1 protein, S cerevisiae
  • Protein-Serine-Threonine Kinases
  • RAD53 protein, S cerevisiae
  • Kinesin
  • Hydroxyurea