genome instability

• • dna endonucleases - cleaves chain
  • dna ligase - join ends
  • dna glycolyases - removes damaged bases
  • dna polymerase - add add nucleotides
• dna damage

  structural abnormality
• classes of dna damage

  • loss of base - apurinic/apyrimidinic site
  • small adduct - oxygen/methyl group
  • bulky adduct
  • cross links - intra or inter strand
  • mismatched
  • single / double strand breaks
• result of small adduct

  • 8oxoG
  • O6methylG
  • cause mismatch during DNA replication
• bulky adducts

  • e.g benzopyrene
  • distorts DNA helix, blocks DNA replication, stalls transcription
• 3' end of DNA = OH
• 5' group of DNA = P
• ssb

  • often accompanied by loss of base + 5' +/ 3' termini damage
  • block DNA replication, stall transcription
• cross link / dbsb

  loss of genetic material
• what can damage dna?

  • ros -> small adducts / break strands
  • dna replicative stress
  • endogenous agents
  • exogenous
  • UV light -> melanoma
  • smoking -> lung cancer
  • infections
• pyrimidines

  C T U
• purines

  A G

  larger
• uv -> carcinoma

  • pyrimidine absorbs UV light -> reactive
  • pyrimidine dimerise e.g C-C
  • repaired by NER pathway
• smoking -> lung cancer

  • causes dna damage, oxidative damage, bulky
  • p53 mutation
  • exposure to polycyclic aromatic hydrocarbons
  • increased mutations at sites prone to adduct formation
• infections -> cancer

  • helicobacter pylori -> stomach cancer
  • hepatitis B + C -> liver cancer
  • chronic inflammation -> ROS
• how cell protects itself from dna damage

  • nucleus
  • how it is packaged
  • pumps pumping out harmful agents
  • enzymes breaking down harmful agents to safe
  • proteins conjugate harmful chemicals
  • detoxification
  • some proteins still functions during detoxification
• response to dna repair

  histone modification -> dna access

  damage detected

  atm/atr activation of many downstream molecules p53, chk 1/2

  repair/apoptosis
• dna damage responses

  apoptosis

  dna repair

  cell cycle checkpoints - arrest
• dna replication error

  strand slippage - insertion/deletion

  dna polymerase adds incorrect nucleotide
• mismatch repair

  MutSalpha recognises + binds to mispaired base

  MutL cuts mismatch

  endonuclease degrades faulty strand

  PCNA directs action

  DNA polymerase

  DNA ligase
• insertion/deletion mismatch repair

  mutSbeta recognises + binds

  mutL cuts mismatch

  endonucleases degrade faulty strand

  PCNA directs action

  DNA polymerase + ligase
• general formula for dna repair pathways

  recognition + excision of damaged

  incision of strand

  repair synthesis - dna polymerase

  dna ligase
• direct reversal repair

  MGMT repairs O6methylG

  by mopping up methyl group

  prevents mispairing with T
• BER

  repairs loss of a base, small adduct, single strand break

  AP site is an intermediate

  DNA glycosylases - excision (e.g OGG1)

  AP endonucleases - cut DNA backbone (APE1)

  DNA polymerase + ligase

  long vs short patch BER
• NER = removes damaged nucleotides

  bulky adducts - GG NER

  pyrimidine dimers - TC NER limits transcription
• dna repair pathway

  NER = bulky adduct / pyrimidine dimers

  BER = ssb / loss of base

  HRR/NHEJ = dsbr / crosslinks

  MMR - mismatched bases

  MGMT - guanine alkylation (small adduct)
• BER repair pathway

  ssb

  loss of base

  small adduct
• HRR/NHEJ reapir pathway

  dsbr

  crosslink
• dsbr

  nhej nonhomologous end joining

  hr homologous recombination
• nhej

  during cell cyle

  recognition protein Ku bind to double strand ends

  DNA-PKcs - pull strands together

  error prone

  often addition/deletion

  ligation
• hr

  at critical stages e.g S + G2

  resection

  Rad51 + BRCA2 take ends

  invades sister chromotids until matching sequence

  error proof
• mutations

  • permanent changes to dna sequence
  • transmitted to daughter cells
  • results from errors in DNA replication or damage to DNA
• mutation consequences on protein

  • prevent it being made
  • overexpression
  • change its function
• why does tumour (inter) heterogeneity occur?

  different mutations within the same cell type

  -> different phenotype
• why does tumour (intra) heterogeneity occur?

  different mutations as cancer cells are actively dividing + selected for
• how is genome instability an enabling characteristic?

  • dna damage repair pathway deregulated in cancer
  • cells with selective advantage survive
• forms of mutations

  • single nucleotide
  • aneuploidy
  • insertions/deletions
  • translocation - gain/loss of entire chromosomes
  • enabled by mitotic checkpoints
  • rearrangement in NH chromosomes
  • microsatellite instability MSI - shorten length, defect in mismatch repair
• microsatellite

  • short repeats (1-6 base pairs)
  • genetic fingerprint
• NH chromosomes

  belong to different chromosome pairs
• importance of genome instability

  • confers selective advantage
  • increases proliferation, decreases apoptosis
  • leads to increasingly aggressive genome instability
  • causes cancer
  • mutations to genes e.g TSGs
  • caretaker genes
  • more than mutations needed
  • resistance against treatments