Related Experiment Video
Updated: Jan 7, 2026
Visualizing and Quantifying Endonuclease-Based Site-Specific DNA Damage
Published on: August 21, 2021
Human MutLα activates methylpurine DNA glycosylase to induce alkylation damage cytotoxicity
Mohamed E Ashour1, Ellissa Krekeler1, Monika Chandan Bhowmik1
1Department of Pathology & Immunology, Division of Laboratory and Genomic Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA.
Abstract:
Alkylation chemotherapy is commonly used against tumors such as glioblastoma, yet resistance often develops through downregulation of mismatch repair (MMR). Previous work has established that loss of MMR prevents the excision of the thymine-containing strand across O 6meG-T mismatches. Thus, MMR dysfunction is advantageous because it prevents a vicious cycle of attempted repair that leads to cell death. Here, we provide an alternative explanation to this prevailing mechanism of alkylation chemoresistance by MMR loss. We find that the MMR protein MutLα physically and functionally interacts with the base excision repair (BER) enzyme methylpurine DNA glycosylase (MPG), which processes common alkylation adducts, such as 7meG and 3meA. Biochemical reconstitution demonstrates that MutLα activates MPG glycosylase activity at least partly by promoting MPG substrate binding. This glycosylase stimulation requires ATP hydrolysis as well as the MLH1-interacting region on MPG. Both MutLα or its ability to interact with MPG promote the generation of alkylation-induced abasic sites in cells, which contribute to the cytotoxicity of methyl methanesulfonate (MMS), an SN2 alkylating agent which does not produce O 6meG. Our results provide new insight into the mechanism of alkylation chemoresistance and uncover an unappreciated crosstalk between MMR and base excision repair.
Insights
Mismatch repair (MMR) loss confers chemoresistance by activating base excision repair (BER) enzymes. This interaction generates DNA damage, impacting cancer treatment strategies.
Area of Science:
- Molecular Biology
- Cancer Research
- DNA Repair Mechanisms
Background:
- Alkylation chemotherapy is vital for treating glioblastoma, but resistance frequently emerges.
- Mismatch repair (MMR) deficiency is a known mechanism of chemoresistance, preventing futile repair cycles.
- The prevailing view links MMR loss to resistance by preventing excision of specific DNA mismatches.
Purpose of the Study:
- To investigate an alternative mechanism for MMR loss-mediated alkylation chemoresistance.
- To explore the interaction between MMR proteins and base excision repair (BER) enzymes.
- To elucidate the functional consequences of MMR-BER crosstalk in cellular response to alkylating agents.
Main Methods:
- Biochemical reconstitution assays to study protein-protein interactions and enzyme activity.
- Cellular assays to assess DNA damage generation and cytotoxicity.
- Investigating the interaction between MutLα and methylpurine DNA glycosylase (MPG).
Main Results:
- MutLα (MMR) physically and functionally interacts with MPG (BER).
- MutLα activates MPG glycosylase activity, dependent on ATP hydrolysis and MPG's MLH1-interacting region.
- This interaction promotes the generation of alkylation-induced abasic sites, contributing to methyl methanesulfonate (MMS) cytotoxicity.
Conclusions:
- MMR loss provides an alternative chemoresistance mechanism by activating MPG's DNA glycosylase activity.
- Uncovered crosstalk between MMR and BER pathways offers new insights into DNA repair.
- This interaction influences cellular response to alkylating chemotherapy, particularly agents like MMS.
More Related Videos
10:59CometChip: A High-throughput 96-Well Platform for Measuring DNA Damage in Microarrayed Human Cells
Published on: October 18, 2014
12:19Tools to Study the Role of Architectural Protein HMGB1 in the Processing of Helix Distorting, Site-specific DNA Interstrand Crosslinks
Published on: November 10, 2016
Related Concept Videos
01:20Mismatch Repair
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...
01:36Mismatch Repair
01:38Nucleotide Excision Repair
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...
01:08Nucleotide Excision Repair
01:30Spontaneous and Induced Mutations
02:25Overview of DNA Repair
Chemically...