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Increased PARP-1 association with DNA in alkylation damaged, PARP-inhibited mouse fibroblasts

Padmini S Kedar1, Donna F Stefanick, Julie K Horton

  • 1Laboratory of Structural Biology, National Institute of Environmental Health Sciences, NIH, 111 T.W. Alexander Drive, Research Triangle Park, NC 27709, USA.

Molecular Cancer Research : MCR
|January 17, 2012
PubMed

Insights

Combining methyl methanesulfonate (MMS) with a PARP-1 inhibitor causes significant cell death. This occurs because inhibited PARP-1 binds to DNA, blocking repair and replication, leading to apoptosis.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Genetics

Background:

  • Methyl methanesulfonate (MMS) and PARP-1 inhibitors induce cell death in mouse fibroblasts.
  • This cell death is apoptotic, requires DNA replication, PARP-1 expression, and an intact S-phase checkpoint.
  • It is hypothesized that inhibited PARP-1 immobilizes at DNA repair sites, hindering repair and replication.

Purpose of the Study:

  • To investigate if inhibited PARP-1 remains DNA-associated during alkylation-induced DNA damage repair.
  • To provide mechanistic insight into the hypersensitivity observed with combined MMS and PARP-1 inhibitor treatment.

Main Methods:

  • Chromatin immunoprecipitation (ChIP) using an anti-PARP-1 antibody.
  • Quantitative Polymerase Chain Reaction (qPCR) for DNA quantification.
  • Treatment of mouse fibroblasts with MMS and a PARP-1 inhibitor.

Main Results:

  • Cells treated with MMS plus a PARP-1 inhibitor showed significantly higher levels of DNA associated with PARP-1 compared to untreated cells.
  • This indicates that inhibited PARP-1 remains bound to DNA during the repair of MMS-induced damage.

Conclusions:

  • The study supports the hypothesis that inhibited PARP-1 accumulates on DNA during repair processes.
  • This DNA-bound, inhibited PARP-1 likely contributes to the observed extreme hypersensitivity to the combination of MMS and PARP-1 inhibitors.
  • Findings offer a mechanistic explanation for the potentiation of cell killing by combined MMS and PARP-1 inhibition.

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