Nucleotide Excision Repair: How Cells Cut Out UV-Damaged DNA

What NER Is and Where It Happens

Nucleotide excision repair is one of the cell's four major single-strand DNA repair systems, and it is the primary defense against damage that bends or unwinds the double helix. Its signature substrates are ultraviolet photoproducts: cyclobutane pyrimidine dimers (CPDs), in which two adjacent pyrimidines (usually thymines) are covalently fused by a four-membered ring, and 6-4 photoproducts (6-4PPs), which kink the helix more sharply. NER also clears bulky chemical adducts such as benzo[a]pyrene from tobacco smoke and intrastrand crosslinks from the chemotherapy drug cisplatin.

• Where: the nucleus of essentially all cells, operating on genomic DNA in chromatin.
• When: throughout the cell cycle, not restricted to S phase.
• Two subpathways: global genome NER (GG-NER) surveys the entire genome, while transcription-coupled NER (TC-NER) targets lesions that stall RNA polymerase II on actively transcribed strands.

The two branches differ only in how the lesion is first detected; the cutting-and-patching machinery downstream is shared, and it is remarkably conserved from bacteria (the UvrABC system) to humans.

The Mechanism, Step by Step

NER proceeds through a defined relay of protein complexes, often summarized as recognize, verify, unwind, cut, fill, seal:

• 1. Recognition. In GG-NER, the XPC-RAD23B complex binds the distorted, single-stranded DNA opposite the lesion; for poorly-distorting CPDs, the UV-DDB (DDB1-DDB2/XPE) complex assists. In TC-NER, a stalled RNA Pol II plus CSB (ERCC6) and CSA (ERCC8) flag the lesion.
• 2. Opening and verification. The ten-subunit transcription/repair factor TFIIH is recruited. Its ATP-dependent helicases XPB (ERCC3) and XPD (ERCC2) unwind ~25-30 bp around the damage; XPD's translocation stalls at the lesion, verifying real damage.
• 3. Pre-incision complex. XPA confirms the chemistry and positions the nucleases, while RPA coats and protects the undamaged strand.
• 4. Dual incision. The endonuclease XPG (ERCC5) cuts 3' of the lesion; the XPF-ERCC1 complex cuts 5'. This releases a 24-32 nt oligonucleotide carrying the damage.
• 5. Gap filling. DNA polymerase δ/ε (with PCNA and RFC) resynthesizes the gap using the intact strand as template.
• 6. Ligation. DNA ligase I (or ligase III/XRCC1) seals the final nick.

Key Molecules and Characteristic Numbers

The pathway's precision comes from a specific cast of proteins encoded by genes named largely after the disease they cause when mutated (XP = xeroderma pigmentosum, CS = Cockayne syndrome, ERCC = excision repair cross-complementing).

• XPC-RAD23B: the initiator sensor of GG-NER; binds distortion, not the lesion itself.
• TFIIH: a ~500 kDa, ten-subunit complex shared with transcription; XPB is a 3'-5' and XPD a 5'-3' ATP-dependent helicase.
• XPG and XPF-ERCC1: structure-specific endonucleases making the 3' and 5' cuts, respectively.

Concrete numbers: the excised patch is ~24-32 nt in humans (the E. coli UvrABC system removes a shorter ~12-13 nt fragment). A 6-4 photoproduct is repaired within roughly 1-2 hours, whereas the less-distorting CPD is far slower, taking 24-48 hours to clear from the genome by GG-NER. The lesion opening consumes ATP through the TFIIH helicases, and each repair event costs the cell the resynthesis of ~30 nucleotides.

How NER Is Studied and Regulated

NER has one of the richest experimental histories in molecular biology, because UV damage is easy to induce and quantify.

• Unscheduled DNA synthesis (UDS): the classic assay measures repair synthesis outside S phase by pulse-labeling with tritiated thymidine or, today, EdU; XP cells show sharply reduced UDS.
• Host-cell reactivation and cell-free extract assays: UV-damaged reporter plasmids incubated with cell extracts let researchers reconstitute dual incision in vitro, which is how Aziz Sancar and colleagues purified the human system.
• Excision-repair sequencing (XR-seq): a modern method that captures and sequences the excised 24-32 nt oligonucleotides, mapping repair across the whole genome at single-nucleotide resolution.
• Complementation groups: fusing cells from different XP patients restores repair only if they carry defects in different genes, defining the seven groups XP-A through XP-G.

Regulation ties NER to chromatin (histone modifications and remodelers such as the ATP-dependent remodeler open access), to the cell cycle, and to the p53 tumor suppressor, which transcriptionally upregulates XPC and DDB2 after UV.

How NER Differs from Its Cousins

The four excision-type repair pathways are easy to confuse, but they solve distinct chemical problems:

• vs. base excision repair (BER): BER handles small, non-distorting lesions (oxidized guanine, uracil, alkylated bases) using lesion-specific glycosylases and typically replaces just 1 nucleotide. NER handles bulky, distorting lesions and replaces ~30. BER senses chemistry; NER senses shape.
• vs. mismatch repair (MMR): MMR corrects replication errors (mispaired bases, small loops) and must know which strand is new; NER acts on damaged bases regardless of replication and needs no strand-discrimination signal.
• vs. photolyase/direct reversal: many organisms (not placental mammals) carry photolyase, which uses blue-light energy to directly monomerize a CPD without cutting DNA. NER instead excises the whole region and is the mammalian fallback.

NER is also uniquely substrate-versatile: one machine clears UV dimers, cisplatin adducts, and cigarette-smoke adducts alike, because it reads distortion rather than any single chemical bond.

Significance, Disease, and Open Questions

NER's importance is written in human disease. Xeroderma pigmentosum (XP), caused by biallelic mutations in XPA-XPG (or the translesion polymerase POLH in the XP-V variant), produces extreme photosensitivity and a ~1,000-2,000-fold increase in skin cancer risk, often before age 10. Defects in the transcription-coupled branch cause Cockayne syndrome (developmental and neurological degeneration) and, with TFIIH mutations, trichothiodystrophy (brittle sulfur-deficient hair). Because cisplatin acts by forming NER-repairable adducts, tumors with high NER (e.g. ERCC1 overexpression) resist platinum chemotherapy, making NER a pharmacogenomic biomarker.

• Landmark history: Setlow and Carrier, and Boyce and Howard-Flanders, described dimer excision in bacteria in 1964; James Cleaver linked defective repair to XP in 1968. Aziz Sancar shared the 2015 Nobel Prize in Chemistry for mechanistic studies of DNA repair including NER.
• Open questions: how chromatin is unpacked and restored around each lesion, how TC-NER coordinates with removal of stalled Pol II, and why some NER mutations cause cancer while others cause premature aging remain active research areas.

NER compared with other single-strand DNA repair pathways

Pathway	Lesions handled	Recognition	Patch size / hallmark
Nucleotide excision repair (NER)	Bulky, helix-distorting (CPDs, 6-4PPs, cisplatin adducts)	Detects helix distortion, not specific base	~24-32 nt oligo excised (human); dual incision
Base excision repair (BER)	Small non-distorting damage (oxidized/deaminated/alkylated bases)	Lesion-specific DNA glycosylases	1 nt (short-patch) or 2-10 nt; abasic-site intermediate
Mismatch repair (MMR)	Replication mispairs, small insertion/deletion loops	MutSα/MutLα scan post-replication	Long patch (>100 nt); strand-discrimination
Global genome NER (GG-NER)	Damage anywhere in genome	XPC-RAD23B (+ UV-DDB for CPDs)	Slow for CPDs; whole-genome surveillance
Transcription-coupled NER (TC-NER)	Damage in transcribed strand of active genes	Stalled RNA Pol II + CSA/CSB	Fast; couples repair to transcription