ATM mediates repression of DNA end-degradation in an ATP-dependent manner

Elias A. Rahal, Leigh A. Henricksen, Yuling Li, John Turchi, Katherine S. Pawelczak, Kathleen Dixon

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Ataxia telangiectasia mutated (ATM) is a PI3-kinase-like kinase (PIKK) associated with DNA double-strand break (DSB) repair and cell cycle control. We have previously reported comparable efficiencies of DSB repair in nuclear extracts from both ATM deficient (A-T) and control (ATM+) cells; however, the repair products from the A-T nuclear extracts contained deletions encompassing longer stretches of DNA compared to controls. These deletions appeared to result from end-joining at sites of microhomology. These data suggest that ATM hinders error-prone repair pathways that depend on degradation of DNA ends at a break. Such degradation may account for the longer deletions we formerly observed in A-T cell extracts. To address this possibility we assessed the degradation of DNA duplex substrates in A-T and control nuclear extracts under DSB repair conditions. We observed a marked shift in signal intensity from full-length products to shorter products in A-T nuclear extracts, and addition of purified ATM to A-T nuclear extracts restored full-length product detection. This repression of degradation by ATM was both ATP-dependent and inhibited by the PIKK inhibitors wortmannin and caffeine. Addition of pre-phosphorylated ATM to an A-T nuclear extract in the presence of PIKK inhibitors was insufficient in repressing degradation, indicating that kinase activities are required. These results demonstrate a role for ATM in preventing the degradation of DNA ends possibly through repressing nucleases implicated in microhomology-mediated end-joining.

Original languageEnglish
Pages (from-to)464-475
Number of pages12
JournalDNA Repair
Volume7
Issue number3
DOIs
StatePublished - Mar 1 2008

Fingerprint

Ataxia Telangiectasia
Adenosine Triphosphate
Repair
Degradation
DNA
Phosphotransferases
Phosphatidylinositol 3-Kinases
Joining
T-cells
Corrosion inhibitors
Caffeine
Double-Stranded DNA Breaks
Cell Cycle Checkpoints
Cell Extracts
Cells
Substrates
T-Lymphocytes

Keywords

  • ATM
  • DNA degradation
  • Double-strand break repair
  • Microhomology-mediated end-joining
  • PI-3-kinase-like kinases

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

Rahal, E. A., Henricksen, L. A., Li, Y., Turchi, J., Pawelczak, K. S., & Dixon, K. (2008). ATM mediates repression of DNA end-degradation in an ATP-dependent manner. DNA Repair, 7(3), 464-475. https://doi.org/10.1016/j.dnarep.2007.12.003

ATM mediates repression of DNA end-degradation in an ATP-dependent manner. / Rahal, Elias A.; Henricksen, Leigh A.; Li, Yuling; Turchi, John; Pawelczak, Katherine S.; Dixon, Kathleen.

In: DNA Repair, Vol. 7, No. 3, 01.03.2008, p. 464-475.

Research output: Contribution to journalArticle

Rahal, EA, Henricksen, LA, Li, Y, Turchi, J, Pawelczak, KS & Dixon, K 2008, 'ATM mediates repression of DNA end-degradation in an ATP-dependent manner', DNA Repair, vol. 7, no. 3, pp. 464-475. https://doi.org/10.1016/j.dnarep.2007.12.003
Rahal, Elias A. ; Henricksen, Leigh A. ; Li, Yuling ; Turchi, John ; Pawelczak, Katherine S. ; Dixon, Kathleen. / ATM mediates repression of DNA end-degradation in an ATP-dependent manner. In: DNA Repair. 2008 ; Vol. 7, No. 3. pp. 464-475.
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AB - Ataxia telangiectasia mutated (ATM) is a PI3-kinase-like kinase (PIKK) associated with DNA double-strand break (DSB) repair and cell cycle control. We have previously reported comparable efficiencies of DSB repair in nuclear extracts from both ATM deficient (A-T) and control (ATM+) cells; however, the repair products from the A-T nuclear extracts contained deletions encompassing longer stretches of DNA compared to controls. These deletions appeared to result from end-joining at sites of microhomology. These data suggest that ATM hinders error-prone repair pathways that depend on degradation of DNA ends at a break. Such degradation may account for the longer deletions we formerly observed in A-T cell extracts. To address this possibility we assessed the degradation of DNA duplex substrates in A-T and control nuclear extracts under DSB repair conditions. We observed a marked shift in signal intensity from full-length products to shorter products in A-T nuclear extracts, and addition of purified ATM to A-T nuclear extracts restored full-length product detection. This repression of degradation by ATM was both ATP-dependent and inhibited by the PIKK inhibitors wortmannin and caffeine. Addition of pre-phosphorylated ATM to an A-T nuclear extract in the presence of PIKK inhibitors was insufficient in repressing degradation, indicating that kinase activities are required. These results demonstrate a role for ATM in preventing the degradation of DNA ends possibly through repressing nucleases implicated in microhomology-mediated end-joining.

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