3. DNA damage

DNA damage is caused by…

1. replication errors

2. spontaneous

3. mutagen induced damage

DNA damage: replication errors

Proofreading activity of DNA polymerases detects and repairs mismatches
Fidelity of replication: errors approx.1 in every 10^9 -10^10 nt

DNA damage: spontaneous types

Alkylation, Depurination, Deamination

DNA repair

Complex processes employed to fix: endogenous damage & exogenous damage

Hundreds of thousands of mutagenic lesions repaired daily

exogenous damage

DNA damaged by mutagens

(endogenous damage)

Mismatched bases during replication

mutations

permanent changes

undetected/ unrepaired

errors ARE NOT THE SAME as mutations

endogenous mutagens

free radicals

exogenous mutagens

alkylating agents

radiation (ionizing, xrays, UV)

chemicals (pollutants, pesticides, etc)

carcinogens

mutagens that induce normal cells to become cancerous

types of DNA repair for single stranded damage

Direct damage reversal
– Excision repair
■ Base excision repair (BER)
■ Mismatch repair (MMR)
■ Nucleotide excision repair (NER)

ss damage: Direct damage reversal

Simplest mechanism

repair done without cuts in the DNA backbone
wasteful process

Mostly for removal of drug-induced (chemotherapy) alkylation of guanine (O6-methylguanine)
– Methyl groups removed by methytransferase
– Methylated enzyme becomes inactive → one enzyme is needed for each methyl group removed

Generic DNA excision repair pathway

Common steps:

1. Detection of damage

2. Excision of damaged DNA by endo-/exonucleases

3. Replacement of excised region by DNA polymerase

4. Nick sealing by DNA ligase

base excision repair pathway (BER)

Repairs small distortions in DNA caused by damaged bases

After exposure to alkylating or oxidizing agents

– Spontaneous
■ Depurination (>1000/day)
■ Deamination (C → U) (>100/day)

• Deamination of methylated C → T escapes repair
  
  Sugar-phosphate backbone remains intact!

mismatch repair (MMR)

Removes and repairs mispaired, but undamaged bases added during replication (escape DNA polymerase proofreading)

Corrects small insertions or deletions

Mismatched bases in newly synthesized DNA produce distortions and trigger MMR
Newly synthesized strand recognized by presence of nicks
Parental strand used as template

Defects in MMR

Defects in MMR → Lynch syndrome (AD)
Major phenotype (15% of all colon cancers)

nucleotide excision repair (NER)

Detects and corrects bulky damage that distort the DNA double helix → replication
block

Cigarette smoke – benzo(a)pyrene-guanine adducts
UV radiation - adjacent pyrimidine dimers (T-T)
Unresolved T-T dimers result in UV-induced skin cancers

defects in NER →

Defects in NER result in xeroderma pigmentosum (AR)
Homozygotes show UV-sensitivity with increased incidence of skin cancer = “children of the night”

types of DNA repair for double stranded damage

Recombination repair
■ Homologous recombination (HR)
■ Non-homologous end joining (NHEJ)

repair of double stranded breaks-

Ionizing radiation, ROS, and inadvertent action of nuclear enzymes induce double-strand breaks(DSBs) in the DNA, which are the main cause for genomic instability and large mutations

homologus recombination

information from the sister chromatid is used as template to replace the missing region
■ More complex, but more precise
■ Mostly during the S-phase of the cell cycle
■ Preferred mechanism in yeast (lots of repetitive sequences in animal DNA → gene amplification)

non homologous end joining

Primary method of DSB repair in mammalian cells
■ Joining of strand breaks with no sequence similarity = simplest and fastest mechanism to
heal DSBs
■ Functions throughout the cell cycle (mostly G1)
■ Generally results in deletions or small insertions
■ Sequence of repaired DNA is NEVER the same with the original