Mismatch repair proteins collaborate with methyltransferases in the repair of O6-methylguanine

Abstract

DNA repair is essential for combatting the adverse effects of damage to the genome. One example of base damage is O 6 -methylguanine (O 6 mG), which stably pairs with thymine during replication and thereby creates a promutagenic O 6 mG:T mismatch. This mismatch has also been linked with cellular toxicity. Therefore, in the absence of repair, O 6 mG:T mismatches can lead to cell death or result in G:C→A:T transition mutations upon the next round of replication. Cysteine thiolate residues on the Ada and Ogt methyltransferase (MTase) proteins directly reverse the O 6 mG base damage to yield guanine. When a cytosine is opposite the lesion, MTase repair restores a normal G:C pairing. However, if replication past the lesion has produced an O 6 mG:T mismatch, MTase conversion to a G:T mispair must still undergo correction to avoid mutation. Two mismatch repair pathways in E. coli that convert G:T mispairs to native G:C pairings are methyl-directed mismatch repair (MMR) and very short patch repair (VSPR). This work examined the possible roles that proteins in these pathways play in coordination with the canonical MTase repair of O 6 mG:T mismatches. The possibility of this repair network was analyzed by probing the efficiency of MTase repair of a single O 6 mG residue in cells deficient in individual mismatch repair proteins (Dam, MutH, MutS, MutL, or Vsr). We found that MTase repair in cells deficient in Dam or MutH showed wild-type levels of MTase repair. In contrast, cells lacking any of the VSPR proteins MutS, MutL, or Vsr showed a decrease in repair of O 6 mG by the Ada and Ogt MTases. Evidence is presented that the VSPR pathway positively influences MTase repair of O 6 mG:T mismatches, and assists the efficiency of restoring these mismatches to native G:C base pairs.

Cytotoxic and mutagenic properties of O6-alkyl-2′-deoxyguanosine lesions in Escherichia coli cells

“…6B). Intriguingly, G 3 A muta- Replication studies of O 6 -alkylguanine lesions tion frequency for O 6 -Me-dG was increased to 100% only in cells lacking both Ogt and Ada, which is consistent with previous findings (16,24).…”

Cytotoxic and mutagenic properties ofO4-alkylthymidine lesions inEscherichia colicells

“…M13mp7(L2), wild-type AB1157 and C215 E. coli strains, and Ada- and Ogt-deficient E. coli strains (as FC215 derivatives), including C216 (Δ ogt::kan ), C217 (Δ ada Δ alkB::cam ) and C218 (Δ ogt::kan Δ ada Δ alkB::cam ), were kindly provided by Prof. John M. Essigmann ( 35 ). Polymerase-deficient AB1157 strains [Δ pol B1 ::spec (Pol II-deficient), Δ dinB (Pol IV-deficient), Δ umuC ::kan (Pol V-deficient) and Δ pol B1 ::spec ΔdinB Δ umuC ::kan(Pol II, Pol IV, Pol V-triple knockout)] were generously provided by Prof. Graham C. Walker ( 36 ).…”

Prochlorococcus: Advantages and Limits of Minimalism

“…One of them, ybaZ, is related to the ada and ogt genes, with which it was confounded in previous genome analyses (Dufresne et al 2005, Marais et al 2008. Ada and Ogt methyltransferases, which are known to prevent G:C to A:T tranversions in E. coli by reverting potentially mutagenic 6-O-methylguanine adducts into guanine (Rye et al 2008), are in fact seemingly missing in all marine picocyanobacteria. The alkyltransferase-like protein YbaZ does not have any alkyltransferase activity per se in E. coli but was suggested to tag nucleotides affected by large 6-O-alkylguanine adducts (such as ethyl-or propylguanine) before their excision by the nucleotide excision repair (NER) pathway (Mazon et al 2009).…”