11. DNA DSB Repair

• gamma H2AX is a marker of DSBs, as these disappear it indicates that the DSBs are being repaired
• NHEJ doesnt require a homologous template and can occur in any phase of the cell cycle - small deletions can occur
• NHEJ:

  1. Ku70/Ku80 and DNA PK bind
  2. PAXX stabilises the 2 ends
  3. Artemis (nuclease) breaks down one side of each strand
  4. ligation

  possible loss of 1-4 nucleotides
• alternative end joining normally only occurs when there is an issue with NHEJ pathway
• AEJ:

  1. MRN complex and CtiP binds - CtiP facilitates DNA end resection
  2. PolQ extends DNA leading to flaps
  3. Fen1 trims any flaps
  4. DNA ends are ligated together using PARP1 and XRCC1 complex

  loss of over 4 nucleotides
• homologous recombination requires a homologous template so only occurs in late S and G2, has high accuracy
• HR:

  1. MRN complex and CtIP bind
  2. ssDNA overhangs generated, RPA binds there
  3. BRCA2 recruits Rad51 to replace RPA
  4. Rad51 invades sister chromatid forming a D-loop, uses intact DNA as a template
  5. DNA polymerase elongates this
  6. double holiday junction formed, dissolved by dissolution or resolution pathway

  dissolution = no DNA transfer between templates

  resolution = possible DNA transfer, uses nucleases to break DNA crosses