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Mutation and DNA Repair

Mutation vs DNA damage

  • DNA damage does not necessarily equal a mutation.
  • a mutation occurs when DNA damage is preserved through a cell cycle (i.e., fixed into the DNA sequence).

Sources of mutation (mutagens)

Internal

Replication errors

  • adding incorrect nucleotides during DNA replication

Reactive oxygen species (ROS)

Spontaneous nucleotide modifications

Tautomeric shift

  • hydrogen atom shifts between forms (keto/enol), altering H-bonding and base pairing

Depurination

  • loss of the glycosidic bond → base loss (AP site)

Deamination

  • loss of an amine group from a base
  • example: cytosine → uracil

External

Radiation

Ionizing radiation

  • breaks chemical bonds and ejects electrons
  • examples: X-rays, gamma rays

Non-ionizing radiation

  • excites electrons
  • UV forms thymine dimers (intra-strand dimerization)

Alkylating agents

  • add alkyl groups to DNA bases
  • example: mustard gas

Polycyclic aromatic hydrocarbons (PAHs)

  • fused carbon/hydrogen ring structures
  • form DNA adducts and can disrupt glycosidic bonds

Base analogs

  • molecules that resemble bases/nucleotides and can be misincorporated

RNA viruses

  • HIV, HCV, HTLV
  • can insert viral nucleic acid into host DNA

Bacteria and chronic inflammation

  • chronic inflammation can increase damage risk and mutagenic processes

Types of mutations

DNA-level mutations

Base-pair substitutions (point mutations)

  • transition: purine ↔ purine (A ↔ G) or pyrimidine ↔ pyrimidine (T ↔ C)
  • transversion: purine ↔ pyrimidine (or vice versa)

Insertions and deletions

  • can cause frameshift mutations (ribosomal reading frame shift)

Splice-site mutations

  • mutations at splice signals recognized by the spliceosome

  • can cause:

  • intron retention, or

  • exon skipping / truncation

Protein-level outcomes

Silent mutation

  • no amino-acid change

Non-silent mutations

  • missense: amino-acid substitution
  • nonsense: introduces a stop codon (often inactive protein)

Chromosomal mutations

Translocations (often from double-strand breaks)

  • commonly associated with ionizing radiation

Reciprocal translocation

  • segments exchanged between two different chromosomes
  • forms derivative chromosomes

  • typical findings:

  • length discrepancy

  • altered banding pattern
  • 46 chromosomes present

Robertsonian translocation

  • short arms lost, long arms fuse
  • commonly involves chromosomes 13, 14, 15, 21, 22

  • typical findings:

  • length discrepancy

  • altered banding pattern
  • chromosome count can appear normal depending on notation (your note: 46 present)

Ploidy and aneuploidy

Ploidy

  • number of complete sets of chromosomes

  • monoploid: 1 complete set

  • diploid: 2 complete sets

  • polyploidy: more than 2 complete sets (often from mitotic failure)

  • triploidy: 3 complete sets

  • tetraploidy: 4 complete sets

Aneuploidy

  • not a multiple of 23

  • commonly due to nondisjunction

  • monosomy: 1 copy

  • trisomy: 3 copies

Dispermy

  • can cause triploidy (as noted)

DNA repair pathways (redundant systems)

Direct reversal

  • removes damage without excising nucleotides
  • “suicide enzymes”: act once, not catalytic, expensive
  • can only repair specific/small damage types
  • example: methyltransferase (dealkylation)

Excision-based mechanisms (general steps)

  1. recognize damage
  2. excise/remove
  3. repair synthesis
  4. rejoin/seal

Base excision repair (BER)

  • for small lesions (oxidation, deamination, alkylation)

Steps:

  1. DNA glycosylase recognizes lesion (multiple types)
  2. cleaves glycosidic bond → AP site
  3. AP endonuclease + phosphodiesterase remove sugar-phosphate backbone
  4. DNA polymerase \(\beta\) fills
  5. DNA ligase seals

Nucleotide excision repair (NER) — global genomic

  • for bulky lesions (DNA adducts, pyrimidine dimers)

Steps:

  1. XPC + XPE detect helix distortion (e.g., CPD)
  2. XPB + XPD helicase unwind → lesion bubble
  3. XPA verifies damage
  4. XPF + XPG excise ~30 nt segment
  5. DNA polymerase \(\delta\) or \(\epsilon\) fills
  6. DNA ligase I seals

Mismatch repair (MMR)

  • fixes “no damage” base mismatches (non–Watson-Crick pairing)

Steps:

  1. MSH detects mismatch kink
  2. recruits MLH
  3. nick introduced on the incorrect strand
  4. EXO1 removes ~50–200 nt
  5. DNA polymerase \(\delta\) or \(\epsilon\) fills
  6. DNA ligase I seals

Non-homologous end joining (NHEJ)

  • active in late mitosis, G1, early S
  • no homologous template

Steps:

  1. Ku heterodimer binds both DNA ends
  2. ends must be blunt (or processed to become blunt)
  3. ligase IV joins ends

Notes:

  • end processing (“blunting”) can delete nucleotides

Homologous recombination (HR)

  • active in late S, G2, early mitosis
  • requires sister chromatids

Steps:

  1. MRN complex trims from the \(5'\) end to create an overhang

  2. MRN interacts with CtIP

  3. Rad51 mediates strand invasion with sister chromatid

  4. requires BRCA1

  5. DNA polymerase \(\delta\) / \(\epsilon\) extends
  6. DNA ligase I seals

Transcription-coupled repair (TCR) — a type of NER

  • RNA polymerase stalls at damage during transcription

Steps:

  1. stalled RNA Pol acts as the signal
  2. Cockayne syndrome B (CSB) binds RNA Pol
  3. CSA binds with ubiquitin ligase activity → ubiquitinates RNA Pol
  4. RNA Pol is degraded

  5. NER steps proceed:

  6. XPB/XPD unwind

  7. XPA verifies
  8. XPF/XPG excise ~30 nt
  9. DNA polymerase \(\delta\)/\(\epsilon\) fills
  10. DNA ligase I seals

Translesion synthesis (TLS)

  • prevents replicative polymerases from stalling indefinitely

Steps:

  1. when DNA pol \(\delta\)/\(\epsilon\) stalls, PCNA is monoubiquitinated
  2. replicative polymerase is displaced

  3. DNA pol \(\eta\) (error-prone) inserts nucleotides across lesion

  4. reduced nucleotide discrimination (truncated finger region)

  5. after bypass, PCNA loses monoubiquitination
  6. DNA pol \(\eta\) replaced by DNA pol \(\delta\)/\(\epsilon\)

Clinical implication

  • malfunction in repair pathways increases mutation accumulation and cancer risk