ATR kinase activation mediated by MutSalpha and MutLalpha in response to cytotoxic O6-methylguanine adducts

Mol Cell. 2006 May 19;22(4):501-10. doi: 10.1016/j.molcel.2006.04.023.

Abstract

S(N)1-type alkylating agents that produce cytotoxic O(6)-methyl-G (O(6)-meG) DNA adducts induce cell cycle arrest and apoptosis in a manner requiring the DNA mismatch repair (MMR) proteins MutSalpha and MutLalpha. Here, we show that checkpoint signaling in response to DNA methylation occurs during S phase and requires DNA replication that gives rise to O(6)-meG/T mispairs. DNA binding studies reveal that MutSalpha specifically recognizes O(6)-meG/T mispairs, but not O(6)-meG/C. In an in vitro assay, ATR-ATRIP, but not RPA, is preferentially recruited to O(6)-meG/T mismatches in a MutSalpha- and MutLalpha-dependent manner. Furthermore, ATR kinase is activated to phosphorylate Chk1 in the presence of O(6)-meG/T mispairs and MMR proteins. These results suggest that MMR proteins can act as direct sensors of methylation damage and help recruit ATR-ATRIP to sites of cytotoxic O(6)-meG adducts to initiate ATR checkpoint signaling.

Publication types

  • Research Support, N.I.H., Intramural

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Animals
  • Ataxia Telangiectasia Mutated Proteins
  • Base Pair Mismatch
  • Base Sequence
  • Carrier Proteins / metabolism
  • Cell Cycle Proteins / metabolism*
  • Cell Line
  • DNA / genetics
  • DNA / metabolism
  • DNA Adducts / metabolism*
  • DNA Adducts / toxicity
  • DNA Damage
  • DNA Repair / physiology*
  • DNA-Binding Proteins
  • Enzyme Activation / drug effects
  • Exodeoxyribonucleases / metabolism
  • Guanine / analogs & derivatives*
  • Guanine / metabolism
  • HeLa Cells
  • Humans
  • Mice
  • MutS DNA Mismatch-Binding Protein / metabolism*
  • Nuclear Proteins / metabolism
  • Phosphoproteins / metabolism
  • Protein-Serine-Threonine Kinases / metabolism*
  • S Phase
  • Signal Transduction

Substances

  • ATRIP protein, human
  • Adaptor Proteins, Signal Transducing
  • Carrier Proteins
  • Cell Cycle Proteins
  • DNA Adducts
  • DNA-Binding Proteins
  • Nuclear Proteins
  • Phosphoproteins
  • Guanine
  • DNA
  • O-(6)-methylguanine
  • ATR protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • Protein-Serine-Threonine Kinases
  • Exodeoxyribonucleases
  • three prime repair exonuclease 1
  • MutS DNA Mismatch-Binding Protein