Recombinational DNA Repair in Eukaryotes

Abstract

Double‐strand breaks and other deoxyribonucleic acid (DNA) damage can be repaired in cells by a process called homologous recombinational repair (HRR) that involves recombination with an undamaged DNA molecule. HRR is mechanistically complex and requires many enzymatic steps. It is conserved throughout eukaryotes and is key to maintaining genome stability, resolving collapsed replication forks and avoiding cancer. In HRR, a broken strand invades a homologous molecule and primes new DNA synthesis using this undamaged molecule as a template. Subsequent steps involve reannealing to the original molecule, further DNA synthesis and ligation. An intermediate called a double Holliday junction may be formed that links the damaged and undamaged DNA duplexes together. This structure can be dissolved by helicase/topoisomerase or endonuclease reactions that may or may not lead to crossing‐over of outside regions away from the break. HRR is distinct from but mechanistically related to recombination in meiosis and is critical to gene editing.

Key Concepts

  • Homologous recombinational repair (HRR) is a key process that enables cells to survive DNA damage in bacteria, archaeans and eukaryotes.
  • HRR and Nonhomologous end joining are the two main mechanisms for repairing DNA double‐strand breaks.
  • Recombinational repair uses information from a second, undamaged homologous copy of DNA to repair the damaged copy.
  • HRR is a very complex process needing a large number of genes and proteins.
  • The highly conserved RAD51 protein coats single‐stranded DNA tails created by the cell on each side of a double‐strand break.
  • One or both coated tails invade an unbroken DNA duplex to pair with a homologous strand while displacing the other strand.
  • New DNA synthesis and mechanisms for processing of annealed partners restore the original molecule.
  • Heteroduplex DNA, where information on the two strands derives from different parental molecules, is formed during the repair process.
  • The outside regions of the two original duplexes may or may not be exchanged with each other by crossover when HRR happens.
  • In vertebrates, HRR is key to avoiding genomic instability and cancer.

Keywords: recombination; recombinational repair; DNA double‐strand breaks; cancer; histone modifications; Holliday junction; RAD51; heteroduplex DNA; D‐loop

Figure 1. Steps in synthesis‐dependent strand annealing: (a) A DNA break in both strands of one chromatid. (b) Resection from only the 5′ ends on each side of the break leaves 3′ tails on the other strands. (c) Strand invasion by a 3′ tail leads to formation of a D‐loop (brown) and primes new synthesis (dashed blue line). (d) Further newly synthesised DNA anneals to the other 3′ end. (e) Additional synthesis and ligations restore an intact molecule.
Figure 2. Generation of a double Holiday Junction (dHJ): (a) and (b) as in Figure . (c) The invading 3′ tail primes new synthesis and the displaced D‐loop anneals with the other 3′end. (d) This second end primes new synthesis using the D‐loop strand as a template. (e) Ligation of the newly synthesised regions (blue dashed lines) to the resected 5′ ends of the broken molecule generates a dHJ.
Figure 3. Three ways to resolve a double Holliday Junction (see text): (a) Endonucleases cut at the sites shown by arrows, followed by ligation to generate heteroduplex DNA (solid over dashed lines) with recombined outside regions (green chromatid joined to brown chromatid). (b) As panel (a) except that endonucleases cut differently (see arrows) to generate molecules with hDNA but no recombination of outside regions. (c) No endonuclease cuts occur. Instead, junctions move towards each other by branch migration (see text) and a topoisomerase enzyme passes strands through each other, leaving hDNA but no outside recombination.
Figure 4. Steps in single‐strand annealing: (a) Homologous sequences (shown in black) occur in the same molecule on each side of a break site. (b) The homologous sequences are exposed by resection of the 5′ ends on each side of the break site. (c) The homologous sequences anneal with each other to leave unpaired single stranded overhangs on each side. (d) The overhangs are removed and resynthesis occurs (dashed blue lines) to generate a shorter repaired molecule deleted for the sequence between the homologous regions. The other chromatid is not involved but is shown for comparison.
Figure 5. Steps in break‐induced replication: (a) A single‐ended break is shown in one molecule. (b) Resection of one strand leaves a 3′ tail. (c) Strand invasion by this tail displaces a D‐loop and primes new synthesis (dashed blue line). (d) Further synthesis pushed the D‐loop forward leaving a single‐stranded newly synthesised strand behind it. (e) This new strand becomes a template for a second new strand synthesised back to the broken end, followed by ligation to generate an intact duplex with two newly synthesised strands paired with each other.
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Game, John C, and Chernikova, Sophia B(Oct 2020) Recombinational DNA Repair in Eukaryotes. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0029220]