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DNA synthesis from unbalanced nucleotide pools causes limited DNA damage that triggers ATR-CHK1-dependent p53 activation
Pharmaceutical Science and Research
  • Kedar Hastak
  • Rajib K. Paul
  • Mukesh K. Agarwal
  • Vijay S. Thakur
  • Arm R. Amin, Marshall University
  • Sudesh Agrawal
  • R. Michael Sramkoski
  • James W. Jacobberger
  • Mark W. Jackson
  • George R. Stark
  • Munna L. Agarwal
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p53-dependent G1 and G2 cell cycle checkpoints are activated in response DNA damage that help to maintain genomic stability. p53 also helps to protect cells from damage that occurs during S phase, for example, when the cells are starved for DNA precursors or irradiated with a low dose of UV. p53 is activated in normal cells starved for pyrimidine nucleotides by treatment with N-(phosphonacetyl)-L-aspartate (PALA). The treated cells progress through a first S phase with kinetics similar to those of untreated cells. However, the DNA of the treated cells begins to become damaged rapidly, within 12 h, as revealed by a comet assay, which detects broken DNA, and by staining for phosphorylated histone H2AX, which accumulates at sites of DNA damage. Because the cells survive, the damage must be reversible, suggesting single-strand breaks or gaps as the most likely possibility. The transiently damaged DNA stimulates activation of ATR and CHK1, which in turn catalyze the phosphorylation and accumulation of p53. Although PALA-induced DNA damage occurs only in dividing cells, the p53 that is activated is only competent to transcribe genes such as p21 and macrophage inhibitory cytokine 1 (whose products regulate G2 and G1 or S phase checkpoints, respectively) after the cells have exited the S phase during which damage occurs. We propose that p53 is activated by stimulation of mismatch repair in response to the misincorporation of deoxynucleotides into newly synthesized DNA, long before the lack of pyrimidine nucleoside triphosphates causes the rate of DNA synthesis to slow appreciably.


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Copyright © 2008 by The National Academy of Sciences of the USA.

Citation Information
Hastak K, Paul RK, Agarwal MK, Thakur VS, Amin AR, Agrawal S, Sramkoski RM, Jacobberger JW, Jackson MW, Stark GR, Agarwal ML. DNA synthesis from unbalanced nucleotide pools causes limited DNA damage that triggers ATR-CHK1-dependent p53 activation. Proceedings of the National Academy of Sciences. 2008 Apr 29;105(17):6314-9.