Difficulty: Intermediate | Prerequisites: DNA structure, DNA replication basics
DNA is constantly being damaged by UV light, chemical reactions, and replication errors. Cells have several repair systems, each specialised for a different kind of damage: nucleotide excision repair for bulky lesions, base excision repair for small base modifications, mismatch repair for replication slip-ups, and two pathways for double-strand breaks (homologous recombination and non-homologous end joining). When these systems fail, the consequences range from point mutations to cancer.
Nucleotide excision repair (NER)
A repair pathway that removes bulky, helix-distorting DNA lesions (such as pyrimidine dimers caused by UV light) by excising a short stretch of the damaged strand and resynthesising it using the undamaged strand as a template.
Think of it as cutting out a damaged section of pipe and replacing it with a new piece.
Base excision repair (BER)
A repair pathway that corrects small, non-helix-distorting base modifications (such as oxidised or deaminated bases) by removing the damaged base alone, then cutting the backbone, filling the gap, and sealing it.
In simple terms, this is a single-letter correction rather than removing a whole chunk.
Mismatch repair (MMR)
A post-replication quality control system that detects and corrects base pairs that were incorrectly matched during DNA replication but escaped the polymerase's own proofreading.
Think of it as spell-check that runs after the document has been typed.
Pyrimidine dimer
An abnormal covalent bond between two adjacent pyrimidine bases (typically thymines) on the same DNA strand, caused by UV radiation. Distorts the helix and blocks replication and transcription if not repaired.
DNA glycosylase
An enzyme that recognises a specific type of damaged base and cleaves the glycosidic bond linking it to the sugar-phosphate backbone, leaving an abasic site (AP site). This is the first step of base excision repair.
Transcription-coupled repair (TC-NER)
A sub-pathway of NER that preferentially repairs damage on the template strand of actively transcribed genes. When RNA polymerase stalls at a lesion, it signals repair factors to that site.
In simple terms, genes the cell is actively using get priority repair service.
Global genomic repair (GG-NER)
The other sub-pathway of NER, which scans the entire genome for bulky lesions regardless of whether the gene is being transcribed.
Homologous recombination (HR)
A high-fidelity repair pathway for double-strand breaks that uses the undamaged sister chromatid as a template to restore the broken sequence accurately. Available mainly in S and G2 phases of the cell cycle, when a sister chromatid is present.
Non-homologous end joining (NHEJ)
A repair pathway for double-strand breaks that directly ligates the broken ends together without a template. Faster than HR but error-prone: it can cause small insertions or deletions at the break site.
Think of it as gluing two broken ends of a rope together, which may lose or gain a few fibres at the join.
Xeroderma pigmentosum (XP)
A rare inherited disorder caused by defects in nucleotide excision repair genes. Patients cannot repair UV-induced DNA damage effectively and have an extremely high risk of skin cancer with even minimal sun exposure.
DNA is not chemically inert. It is damaged thousands of times per cell per day by spontaneous hydrolysis, reactive oxygen species, UV radiation, and replication errors. Without repair, mutations accumulate, proteins malfunction, and cells either die or become cancerous.
Targets bulky, helix-distorting lesions, most notably pyrimidine dimers (thymine dimers) caused by UV light.
Two sub-pathways:
Transcription-coupled NER (TC-NER): triggered when RNA polymerase stalls at a lesion on the template strand of an active gene. Repairs actively transcribed DNA first.
Global genomic NER (GG-NER): damage-sensing proteins scan the entire genome for helix distortions, regardless of transcription status.
General steps:
Recognition of the lesion.
Dual incisions flanking the damage (typically cutting out a stretch of roughly 24 to 32 nucleotides in eukaryotes).
Removal of the damaged fragment.
Gap filling by a DNA polymerase using the undamaged strand as template.
Ligation to seal the nick.
Targets small, non-helix-distorting lesions: oxidised bases, deaminated bases (e.g. uracil arising from cytosine deamination), alkylated bases.
Steps:
A specific DNA glycosylase recognises the damaged base and cleaves the glycosidic bond, releasing the base and leaving an abasic (AP) site.
AP endonuclease cuts the backbone at the AP site.
DNA polymerase fills the gap (short-patch: one nucleotide; long-patch: several nucleotides).
DNA ligase seals the nick.
Different glycosylases exist for different types of damage, so BER is really a family of related pathways sharing the same downstream steps.
Corrects base-base mismatches and small insertion/deletion loops that escape the proofreading exonuclease of the replicative polymerase.
The system must distinguish the newly synthesised (error-containing) strand from the parental (correct) strand. In bacteria, this is done through methylation patterns (the parental strand is methylated; the new strand is not yet methylated). The mechanism in eukaryotes involves strand-specific nicks associated with the replication machinery.
Steps:
Mismatch recognition (MutS homologues in eukaryotes).
Recruitment of additional factors (MutL homologues).
Excision of a stretch of the new strand containing the error.
Resynthesis and ligation.
Defects in MMR genes (e.g. MSH2, MLH1) are linked to Lynch syndrome (hereditary nonpolyposis colorectal cancer, HNPCC).
Double-strand breaks (DSBs) are the most dangerous form of DNA damage. A single unrepaired DSB can kill a cell. Two main pathways handle them:
Uses the intact sister chromatid as a template for accurate repair.
Available primarily during S and G2 phases, when a sister chromatid exists.
Steps (simplified):
The broken ends are processed to generate 3' single-stranded overhangs.
One overhang invades the homologous duplex on the sister chromatid.
DNA synthesis copies the missing information from the template.
The strands are resolved and ligated.
High fidelity: the original sequence is restored.
Directly ligates the two broken ends without a template.
Active throughout the cell cycle, and is the predominant DSB repair pathway in G1 (when no sister chromatid is available).
Ku proteins bind the broken ends and recruit a ligase complex.
Error-prone: small deletions or insertions frequently occur at the junction because the ends may be trimmed or filled before ligation.
A rare autosomal recessive disorder caused by mutations in any of several NER genes (XPA through XPG, plus XPV).
Patients cannot efficiently repair UV-induced pyrimidine dimers.
Even minimal sun exposure leads to severe sunburn, freckling, and a dramatically elevated risk of skin cancers (melanoma, squamous cell carcinoma, basal cell carcinoma), often in childhood.
XP illustrates the direct link between a failure in a specific DNA repair pathway and cancer predisposition.
"NER and BER do the same thing." They do not. NER handles bulky, helix-distorting damage (like pyrimidine dimers) by cutting out a multi-nucleotide stretch. BER handles small, non-distorting base lesions by removing a single damaged base. Different damage, different enzymes, different scale of excision.
"NHEJ is just a backup for homologous recombination." NHEJ is actually the predominant double-strand break repair pathway in G1 phase, when no sister chromatid is available. It is not a lesser version of HR; it is the cell's main option for much of the cell cycle.
"Mismatch repair fixes the parental strand." MMR specifically targets the newly synthesised strand (the one containing the error). The whole point of strand discrimination is to ensure the parental strand, which carries the correct sequence, is preserved.
"Xeroderma pigmentosum patients have weak immune systems." XP is a DNA repair deficiency, not an immune deficiency. The problem is an inability to repair UV-induced DNA damage in skin cells, leading to mutations and cancer.
⚠️ Be able to match each repair pathway to its type of damage. A classic exam format: "Which pathway repairs pyrimidine dimers?" (NER), "Which pathway corrects deaminated bases?" (BER), "Which pathway fixes replication mismatches?" (MMR).
⚠️ Know the difference between HR and NHEJ: HR is accurate but needs a sister chromatid (S/G2 phase); NHEJ is fast and available all cycle but error-prone. Exams love asking when each is used and why.
⚠️ Xeroderma pigmentosum is the textbook clinical example of NER deficiency. If the exam asks for a disease linked to a DNA repair defect, XP is the answer they are looking for. Lynch syndrome (HNPCC) is the corresponding example for mismatch repair deficiency.
⚠️ Understand why strand discrimination matters in MMR. The repair system must know which strand is wrong, otherwise it would have a 50/50 chance of removing the correct base and making the mutation permanent.
Many cancer therapies exploit DNA repair. PARP inhibitors (such as olaparib) block a backup repair pathway in cells that already lack HR due to BRCA1 or BRCA2 mutations, causing those cancer cells to accumulate lethal DNA damage while sparing normal cells. Sunscreen works by absorbing UV radiation before it can form pyrimidine dimers, reducing the load on NER.
True or false: Base excision repair removes a large stretch of nucleotides around the damaged site. (False: BER typically removes a single damaged base. NER is the pathway that excises a multi-nucleotide stretch.)
Fill in the blank: The enzyme that recognises and removes a damaged base in BER is called a ______. (DNA glycosylase.)
True or false: Non-homologous end joining requires a sister chromatid as a template. (False: that describes homologous recombination. NHEJ ligates ends directly without a template.)
Fill in the blank: Patients with xeroderma pigmentosum have defects in the ______ repair pathway. (Nucleotide excision repair / NER.)
True or false: Mismatch repair corrects errors on the parental DNA strand. (False: it corrects the newly synthesised strand, which is the one containing the replication error.)
Q: Compare and contrast nucleotide excision repair (NER) and base excision repair (BER) in terms of the type of damage repaired, the scale of the repair, and the key enzymes involved.
A: NER targets bulky, helix-distorting lesions such as pyrimidine dimers. It excises a stretch of roughly 24 to 32 nucleotides in eukaryotes, and the gap is filled by a DNA polymerase and sealed by ligase. BER targets small, non-helix-distorting base modifications such as oxidised or deaminated bases. A DNA glycosylase removes the single damaged base, AP endonuclease cuts the backbone, and the one-nucleotide (or short) gap is filled and ligated. NER replaces a segment; BER replaces a base.
Q: Explain how the mismatch repair system distinguishes between the parental strand and the newly synthesised strand. Why is this distinction critical?
A: In bacteria, strand discrimination relies on methylation: the parental strand is methylated at GATC sequences by Dam methylase, while the newly synthesised strand is transiently unmethylated. The MMR system targets the unmethylated strand for excision. In eukaryotes, the mechanism involves nicks in the new strand associated with the replication machinery. This distinction is critical because the parental strand carries the correct sequence. If the system could not tell the strands apart, it would have a 50% chance of removing the correct base and permanently fixing the wrong one.
Q: A patient with xeroderma pigmentosum develops multiple skin cancers by age ten. Explain the molecular basis of this disease.
A: XP results from inherited mutations in one of several NER genes (XPA through XPG, or XPV). Without functional NER, the patient's cells cannot repair UV-induced pyrimidine dimers. These unrepaired lesions block normal replication and transcription and, when bypassed by error-prone translesion polymerases, introduce mutations. The accumulation of mutations in oncogenes and tumour suppressor genes drives cancer development, particularly in sun-exposed skin.
Q: Why is homologous recombination considered more accurate than non-homologous end joining for repairing double-strand breaks?
A: HR uses the intact sister chromatid as a template to copy the missing sequence, restoring the original information with high fidelity. NHEJ joins the broken ends directly without a template, often trimming or adding a few nucleotides in the process. This makes NHEJ faster and available throughout the cell cycle, but it comes at the cost of small insertions or deletions at the repair site.
Q: Defects in mismatch repair genes such as MSH2 and MLH1 cause Lynch syndrome. Explain why loss of MMR leads to cancer predisposition.
A: Without functional MMR, replication errors that escape the polymerase's proofreading accumulate at a much higher rate. This is especially evident at microsatellite sequences (short tandem repeats), which are prone to slippage during replication. The resulting microsatellite instability leads to frameshift mutations throughout the genome, including in genes that regulate cell growth and apoptosis, increasing the risk of colorectal and other cancers.
DNA repair connects directly to DNA replication (see companion notes): mismatch repair operates immediately after replication, and the replication fork can stall at unrepaired lesions, triggering repair or tolerance pathways.
Repair deficiencies are central to cancer biology. Tumour suppressor genes such as BRCA1 and BRCA2 encode HR components; their loss is a major driver of breast and ovarian cancers. Understanding repair pathways is also essential for pharmacology, since many chemotherapy drugs work by creating DNA damage that overwhelms the cancer cell's repair capacity.
DNA repair, nucleotide excision repair, NER, base excision repair, BER, mismatch repair, MMR, double-strand break repair, homologous recombination, HR, non-homologous end joining, NHEJ, pyrimidine dimer, thymine dimer, DNA glycosylase, AP site, abasic site, AP endonuclease, transcription-coupled repair, TC-NER, global genomic repair, GG-NER, xeroderma pigmentosum, XP, Lynch syndrome, HNPCC, MSH2, MLH1, microsatellite instability, BRCA1, BRCA2, PARP inhibitor, UV damage, DNA damage response