Dan Yarosh
MENDING LIFE:
WHY AND HOW CELLS PROTECT THEIR DNA
Essays on Genome Stability, Disease, Aging and Therapy
6. Proofreading the Genome: How Mismatch Repair Prevents Mutations
DNA repair pathways provide layer upon layer of proof reading for the transcription and replication of the three billion bases in the genome. These enzymes are built to recognize gross distortions of the double helix caused by breaks or base modifications. But there are more subtle mistakes that slip by these systems: base mismatches and loops formed by small insertions or deletions, all of which are comprised of normal DNA. If left uncorrected, these small errors accumulate rapidly and upon further replication are fixed as mutations.
One further molecular proofreading system is mismatch repair (MMR). This pathway scans newly synthesized DNA, detects c
opying errors that result in bases mismatched with their opposite base, removes them, and restores the correct sequence. When mismatch repair fails, mutation rates increase dramatically. The consequences include cancer predisposition, changes in therapy responses, and altered immune recognition. Mismatch repair is one of the genome’s most important quality-control systems.
The Big Picture: Why Proofreading Is Essential
DNA replication is highly accurate but not perfect. The enzyme system incorporates the wrong base approximately once in every one hundred thousand to one million nucleotides that it replicates. Proofreading activity reduces this error rate further, but residual mismatches remain, primarily base substitutions and frameshift mutations. These changes are difficult to detect among the 3 billion DNA bases, since the mismatched sites contain normal DNA bases, and the DNA is normally packaged in histones and chromosomes.
Imagine that DNA is a giant zipper and each tooth is a DNA base. The human genome zipper would stretch from Miami to Quebec and back. Now imagine all this way the zipper was wrapped around people, under their arms, through their legs, and around their necks. Now, try to find the one tooth of the zipper that is not zipped in! Even if you find it, you need to figure out which base is wrong, or if there is a small duplication or deletion that causes the mispairing. Oh, you are under some time pressure because someone is going to want to unzip soon. This makes looking for a needle in a haystack child’s play.
If not located in the DNA in time, unrepaired mismatches lead to mutations, causing microsatellite instability, protein truncation and altered gene expression.
The system of mismatch repair improves replication fidelity by another one hundred to one thousand-fold, preserving the integrity of the genome across generations of cells. This is not a dispensable luxury for humans. Those with a defect in one of the genes that codes for mismatch repair proteins have Lynch Syndrome, which predisposes to colon and other cancers.
How Mismatch Repair Works
Mismatch repair is a highly coordinated three step process.
(1) Error recognition: Specialized proteins scan newly synthesized DNA for distortions caused by incorrect base pairing. In human cells, the MutS homolog complexes (MSH proteins) recognize single-base mismatches and small insertion or deletion loops in DNA, bind to the error and initiate repair.
(2) Recruitment of repair machinery: After detection, MutL homolog (MLH) proteins are recruited to coordinate downstream repair steps, identify the newly synthesized strand and activate cutting activity. The system must distinguish between the original template strand and the newly synthesized strand to avoid removing the correct sequence.
(3) Excision and Resynthesis: A segment of the mismatched DNA containing the error is removed and then replaced with the correct sequence using the original strand as a template. Finally, the break is resealed.
Even when the mismatch is detected, the fix is not always easy. Several environmental toxins produce an insidious modification of one of the DNA bases, causing it to mispair with the wrong base. When it’s time to copy the pair, the wrong base is used and the sequence is mutated. The mismatch repair system recognizes that something is not right and incises the DNA at the wrong base. The MMR systems continue to mis-read and mis-insert the wrong base, and the repeated cycles of incision, mis-repair and incision eventually kills the cell. This quirk in repair is exploited by the anti-cancer chemotherapeutic agent temozolomide, which produces just such an insidious modification in tumors until the cancer cell kills itself.
Microsatellites: Sensitive Indicators of Repair Failure
Microsatellites are short repetitive DNA sequences scattered throughout the genome. Because of their repetitive structure, they are especially prone to slippage of the template during replication, which can cause their length to change. This is like one side of the zipper having too many teeth across from the other. In cells with defective mismatch repair, microsatellites frequently expand or contract in length. This phenomenon, called microsatellite instability (MSI), serves as a hallmark of MMR deficiency. MSI is now widely used as a diagnostic marker in oncology to identify tumors with flawed MMR.
When It Goes Wrong: Lynch Syndrome and Sporadic Cancer
Mismatch repair defects have some of the clearest clinical consequences in DNA repair biology. In Lynch Syndrome, inherited mutations in key MMR genes (predominantly MSH2 and MLH1) cause a hereditary cancer predisposition disorder. Because replication errors accumulate rapidly in these patients, tumors arise earlier and more frequently. Affected individuals have a markedly increased risk of colorectal, endometrial, ovarian and gastric cancer, but most patients don’t even know they have the disease until an unexpected cancer crops up. Dozens of different mutations in MMR genes have been found in Lynch Syndrome patients and the detection of MSI is invaluable in confirming the diagnosis. Early detection and surveillance dramatically improve the life expectancy of these patients, but they are never out of the woods.
Mismatch repair deficiency also occurs in non-inherited cancers. Sporadic silencing of MMR genes leads to MSI in tumors without a family history of such cancers. These tumors exhibit extremely high mutation burdens, abundant neoantigens, and strong immune infiltration. They are now routinely identified by MSI testing.
Ironically, this instability can make them more responsive to immunotherapy. Tumors with MMR deficiency accumulate large numbers of mutations, producing abnormal proteins that the immune system can recognize. This has led to the approval of immune checkpoint inhibitors for tumors characterized by MSI, regardless of tissue origin. This relationship is so strong that MMR deficiency has become a biomarker for predicting a patient’s response to these drugs.
Beyond Cancer: Neurological Disease and Antibody Development
Although most attention focuses on cancer, mismatch repair also influences the brain. Errors in repetitive DNA sequences can lead to expansion disorders and neurodegenerative conditions. In Huntington’s disease (HD), inherited longer-than-normal repeats in DNA confuse the MMR system, which tries to fix the repeats but instead leads to their amplification. The dramatic expansion of these repeats seems to be at the core of HD neurological symptoms. The more active the MMR complex is, the earlier symptoms begin. One therapeutic approach currently under study is to inhibit the MMR system from binding to these long repeats and thereby block amplification.
In the immune system, MMR is used to overcome genome stability. It is hijacked by B cells to introduce mutations into antibody genes during maturation, allowing the host to adapt its antibody production to the invading challenger. The paradoxical use of a DNA repair system to create mutations is crucial for increasing the affinity of antibodies for pathogens. Deficiencies in the MMR components in B cells lead to impaired antibody maturation and greater susceptibility to infections.
What We’re Learning Now: Precision Gene Editing
Mismatch repair also affects the latest gene editing technologies, which we will cover in the last essay. In precision editing (PE), specific single base changes are introduced in DNA to repair mutant sequences. However, the MMR system works against PE efficiency by reversing the correction. New generations of PE have devised methods to temporarily inhibit the specific MMR complexes during the therapeutic correction to preserve the new edits.
This illustrates a recurring theme: DNA repair pathways are both guardians of the healthy genome and instigators of cancer, genetic disease and neurological disorders. We are just now learning how to control them for better health.
Key Takeaways
- DNA replication errors are inevitable, and dangerous even at low levels
- Mismatch repair dramatically increases replication fidelity
- Failure leads to microsatellite instability, mutations, and cell death
- Mismatch repair deficiency drives Lynch syndrome and many cancers
- Mismatch repair status now guides immunotherapy decisions
- Overcoming mismatch repair is essential to the success of precision gene editing
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References & Further Reading
1. Jiricny, J. (2013). Postreplicative mismatch repair. Cold Spring Harb Perspect Biol, 5, a012633.
Comprehensive overview of MMR mechanisms
2. Li, G. M. (2008). Mechanisms and functions of DNA mismatch repair. Cell Res, 18, 85–98.
Molecular steps of repair
3. Lynch, H. T., & de la Chapelle, A. (2003). Hereditary colorectal cancer. New Engl J Med, 348, 919–932.
Lynch Syndrome overview
4. Hause, R. J., et al. (2016). Classification and characterization of microsatellite instability across 18 cancer types. Nat Med 22, 1342–1350.
MSI as a pan-cancer marker
5. Le, D. T., et al. (2015). PD-1 blockade in tumors with mismatch-repair deficiency. New Engl J Med, 373, 1979.
Landmark immunotherapy trial
6. Baretti, M., & Le, D. T. (2018). DNA mismatch repair in cancer. Pharmacol Ther, 189, 45-62.
Clinical implications of MMR
7. Kunkel, T. A., & Erie, D. A. (2015). Eukaryotic mismatch repair in relation to replication fidelity. Annu Rev Genet 49, 291-313.
Repair and replication accuracy
8. Peña-Diaz, J., & Jiricny, J. (2012). Mammalian mismatch repair: error-free or error-prone? Trends Bioch Sci, 37, 206-214.
Molecular mechanisms of MMR
9. Da Silva, F., et al. (2009). Mismatch repair genes in Lynch syndrome: a review. Sao Paulo Med J, 127, 46-51.
Review of Lynch Syndrome
10. Feng, Y., & Martin, A. (2022). Mutagenic repair during antibody diversification: emerging insights. Trends Immunol 43, 604-607.
Review of MMR in antibody formation