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ATM is activated in response to N-methyl-N'-nitro-N-nitrosoguanidine-induced DNA alkylation

Aaron W Adamson1, Wan-Ju Kim, Sanjeev Shangary

  • 1Department of Biochemistry and Molecular Biology and the Stanley S. Scott Cancer Center, Louisiana State University Health Sciences Center, New Orleans, Louisiana 70112, USA.

The Journal of Biological Chemistry
|August 2, 2002
PubMed

Insights

The ATM kinase, crucial for DNA damage response, is activated by the DNA alkylating agent MNNG, not just radiation. This activation involves DNA repair processes and p53 phosphorylation, independent of apoptosis.

Area of Science:

  • Molecular Biology
  • Cellular Biology
  • DNA Damage Response

Background:

  • The tumor suppressor p53 is vital for cellular responses to DNA damage, including cell cycle arrest and apoptosis.
  • Phosphorylation of p53 by ATM (Ataxia Telangiectasia Mutated) kinase is a key regulatory mechanism in response to ionizing radiation.
  • The precise mechanisms by which DNA alkylating agents like MNNG induce p53 up-regulation and phosphorylation remain unclear.

Purpose of the Study:

  • To investigate the role of ATM kinase in the cellular response to MNNG, specifically p53 up-regulation and phosphorylation.
  • To elucidate the signaling pathways involved in MNNG-induced p53 activation.
  • To determine if MNNG-induced apoptosis is required for ATM activation.

Main Methods:

  • Treatment of normal human fibroblasts and ATM-deficient fibroblasts with MNNG.
  • Use of ATM kinase inhibitor wortmannin to assess ATM's role.
  • Analysis of p53 up-regulation and phosphorylation (Serine 15) in response to MNNG.
  • In vitro kinase assays to measure ATM activation.
  • Comet assays to detect DNA strand breaks.
  • Morphological and biochemical assessments for apoptosis.

Main Results:

  • MNNG-induced p53 up-regulation and phosphorylation in normal fibroblasts were sensitive to wortmannin, indicating ATM involvement.
  • ATM-deficient cells showed delayed p53 up-regulation after MNNG treatment.
  • MMR-deficient cells exhibited rapid p53 phosphorylation at Serine 15 and rapid ATM activation in response to MNNG, despite delayed p53 up-regulation.
  • MNNG did not induce apoptosis in normal fibroblasts, and ATM activation occurred rapidly (within 30 min) alongside DNA strand breaks during repair.
  • DNA strand breaks arising during DNA repair, not apoptosis-induced breaks, appear to activate ATM in response to MNNG.

Conclusions:

  • ATM kinase plays a significant role in the cellular response to the DNA alkylating agent MNNG, mediating p53 up-regulation and phosphorylation.
  • ATM activation by MNNG is linked to DNA repair processes and the accumulation of DNA strand breaks, rather than MNNG-induced apoptosis.
  • The findings expand the known roles of ATM activation beyond ionizing radiation, highlighting its importance in response to DNA alkylation.

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