Molecular clues to alkylating agent‐induced cell death and drug resistance

O 6 ‐methyl‐guanine (O 6 ‐meG) is the major carcinogenic lesion in DNA induced by methylating agents. This DNA adduct can be recognized and processed by both the mismatch repair (MMR) system and the repair protein, O 6 ‐methylguanine methyltransferase (MGMT), also called alkylguanine alkyltransferase (AGT). While the processing of O 6 ‐meG by AGT is to remove the methyl group from O 6 ‐meG, the recognition by MMR recognition proteins, MutSα and MutSβ, initiates a signaling cascade to induce programmed cell death. Although both systems play important roles in maintaining genome stability, they appear to cause an opposite outcome in cancer chemoherapy, i.e., apoptosis mediated by the MMR system and drug‐resistance induced by AGT. However, little is known how these two systems partition O 6 meG adducts in vivo. Using gel‐shift analysis, we determined specific binding affinities of MutSα, MutSβ, and AGT for DNA duplexes containing an O 6 ‐meG adduct. We demonstrate here that under the equal molar condition, the order of binding affinities for these proteins is: MutSβ > AGT > MutSα. These results were confirmed by competitive binding assays. Our data therefore provide useful molecular insights into the mechanism by which alkylating agents induce cell death or drug resistance.