The mammalian mismatch repair pathway removes DNA 8-oxodGMP incorporated from the oxidized dNTP pool
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Updated: Jul 1, 2026
Proofreading and DNA Repair Assay Using Single Nucleotide Extension and MALDI-TOF Mass Spectrometry Analysis
Published on: June 19, 2018
Claudia Colussi1, Eleonora Parlanti, Paolo Degan
1Laboratory of Comparative Toxicology and Ecotoxicology, Istituto Superiore di Sanita', Rome, Italy.
Abstract:
Mismatch repair (MMR) corrects replication errors. It requires the MSH2, MSH6, MLH1, and PMS2 proteins which comprise the MutSalpha and MutLalpha heterodimers. Inactivation of MSH2 or MLH1 in human tumors greatly increases spontaneous mutation rates. Oxidation produces many detrimental DNA alterations against which cells deploy multiple protective strategies. The Ogg-1 DNA glycosylase initiates base excision repair (BER) of 8-oxoguanine (8-oxoG) from 8-oxoG:C pairs. The Myh DNA glycosylase removes mismatched adenines incorporated opposite 8-oxoG during replication. Subsequent BER generates 8-oxoG:C pairs, a substrate for excision by Ogg-1. MTH1-an 8-oxodGTPase which eliminates 8-oxodGTP from the dNTP pool-affords additional protection by minimizing 8-oxodGMP incorporation during replication. Here we show that the dNTP pool is, nevertheless, an important source of DNA 8-oxoG and that MMR provides supplementary protection by excising incorporated 8-oxodGMP. Incorporated 8-oxodGMP contributes significantly to the mutator phenotype of MMR-deficient cells. Thus, although BER of 8-oxoG is independent of Msh2, both steady-state and H(2)O(2)-induced DNA 8-oxoG levels are higher in Msh2-defective cells than in their repair-proficient counterparts. Increased expression of MTH1 in MMR-defective cells significantly reduces steady-state and H(2)O(2)-induced DNA 8-oxoG levels. This reduction dramatically diminishes the spontaneous mutation rate of Msh2(-/-) MEFs.
Insights
Mismatch repair (MMR) corrects DNA replication errors. MMR deficiency increases 8-oxoguanine (8-oxoG) DNA damage, but MTH1 enzyme reduces this, lowering mutation rates in cells.
Area of Science:
- Molecular Biology
- Genetics
- DNA Repair Mechanisms
Background:
- Mismatch repair (MMR) system, involving MSH2, MSH6, MLH1, and PMS2 proteins, corrects DNA replication errors.
- Oxidative stress generates DNA alterations, countered by base excision repair (BER) pathways involving Ogg-1, Myh, and MTH1 enzymes.
- The dNTP pool can be a source of DNA damage, specifically 8-oxoguanine (8-oxoG).
Purpose of the Study:
- To investigate the role of MMR in protecting against DNA 8-oxoG damage originating from the dNTP pool.
- To determine if MMR deficiency impacts DNA 8-oxoG levels and mutation rates.
- To assess the protective effect of MTH1 against 8-oxoG accumulation in MMR-deficient cells.
Main Methods:
- Analysis of DNA 8-oxoG levels in MMR-proficient and MMR-deficient cells (Msh2(-/-) MEFs).
- Assessment of spontaneous and H(2)O(2)-induced DNA 8-oxoG levels.
- Evaluation of MTH1 expression and its impact on DNA 8-oxoG and mutation rates.
Main Results:
- MMR provides supplementary protection against incorporated 8-oxoguanine (8-oxodGMP) from the dNTP pool.
- Msh2-defective cells exhibit higher steady-state and H(2)O(2)-induced DNA 8-oxoG levels.
- Increased MTH1 expression significantly reduces DNA 8-oxoG levels and the spontaneous mutation rate in Msh2(-/-) MEFs.
Conclusions:
- MMR plays a crucial role in mitigating DNA 8-oxoG damage originating from the dNTP pool.
- MTH1 is a key enzyme in protecting MMR-defective cells from accumulating 8-oxoG and developing a mutator phenotype.
- Combined action of MMR and MTH1 is essential for maintaining genomic stability.
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