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Nitrosative stress suppresses checkpoint activation after DNA synthesis inhibition

Robert J Tomko1, Ndang N Azang-Njaah, John S Lazo

  • 1Department of Pharmacology and Chemical Biology and University of Pittsburgh Drug Discovery Institute, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, USA.

Cell Cycle (Georgetown, Tex.)
|January 23, 2009
PubMed

Insights

Nitrosative stress inhibits DNA synthesis by suppressing Cdc25A and preventing cell cycle checkpoints. This failure to activate S-phase checkpoints may increase genome damage transmission in precancerous cells.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • DNA synthesis relies on cyclin-dependent kinase 2 (Cdk2) activation by Cdc25A.
  • Nitrosative stress impacts Cdk2 dephosphorylation and Cdc25A translation.
  • The effect of nitrosative stress on S-phase progression and checkpoint activation is unclear.

Purpose of the Study:

  • To investigate the impact of nitrosative stress on DNA synthesis and S-phase progression.
  • To determine if nitrosative stress activates or suppresses cell cycle checkpoints.
  • To understand the mechanism by which nitrosative stress inhibits DNA synthesis.

Main Methods:

  • Inducible nitric oxide synthase (iNOS) and S-nitrosocysteine ethyl ester (SNCEE) were used to induce nitrosative stress.
  • Ectopic expression of Cdc25A and a Cdc25-independent Cdk2 mutant were employed.
  • Levels of phosphorylated checkpoint proteins (pSer345-Chk1, pThr68-Chk2) and gammaH2AX were assessed.

Main Results:

  • Nitrosative stress rapidly inhibited DNA synthesis and led to Cdc25A loss.
  • Inhibition of DNA synthesis was observed despite ectopic Cdc25A or Cdk2 mutant expression.
  • Nitrosative stress suppressed S-phase checkpoint activation, including the accumulation of key phosphorylated checkpoint proteins.

Conclusions:

  • Nitrosative stress inhibits DNA synthesis independently of canonical checkpoint activation.
  • The suppression of S-phase checkpoints by nitrosative stress may facilitate genome instability.
  • Failure to activate checkpoints under nitrosative stress could increase the risk of transmitting damaged genomes in precancerous cells.

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