Ambiguous coding is required for the lethal interaction between methylated DNA bases and DNA mismatch repair

Abstract

The thiopurine 6-thioguanine (S 6G) is used to treat acute leukaemia. Its cytotoxic effect requires an active DNA mismatch repair (MMR) system. S 6G is incorporated into DNA where a small fraction undergoes in situ conversion to S 6-thiomethylguanine (S 6meG). After replication, S 6meG-containing base pairs interact with MMR. This interaction is ultimately lethal and MMR-defective cells are resistant to S 6G. Here, we report that growing human cells extensively incorporate the thiopyrimidine nucleoside 4-thiothymidine (S 4TdR) into their DNA. The incorporated thiopyrimidine (S 4T) can also undergo facile S-methylation to 4-thiomethylthymine (S 4meT). The rate of methylation of S 4TdR in model substrates is similar to that for the conversion of S 6G to S 6meG indicating that the DNA of cells grown in S 4TdR will contain significant levels of S 4meT. Despite this, S 4TdR is not associated with MMR-related cell death. We demonstrate that, in contrast to S 6meG, neither DNA S 4T nor S 4meT codes ambiguously. S 4T retains the coding properties of unmodified T, whereas S 4meT behaves like a normal cytosine and exclusively directs the incorporation of guanine. The preferred S 4meT:G base pair is also a poor substrate for binding by the hMutSα mismatch recognition factor. We suggest that the ability of S 4meT to produce a structurally acceptable base pair during replication underlies the absence of MMR-related death in cells treated with S 4TdR.