DNA End Resection: Nucleases Team Up with the Right Partners to Initiate Homologous Recombination

Abstract

The repair of DNA double-strand breaks by homologous recombination commences by nucleolytic degradation of the 5'-terminated strand of the DNA break. This leads to the formation of 3'-tailed DNA, which serves as a substrate for the strand exchange protein Rad51. The nucleoprotein filament then invades homologous DNA to drive template-directed repair. In this review, I discuss mainly the mechanisms of DNA end resection in Saccharomyces cerevisiae, which includes short-range resection by Mre11-Rad50-Xrs2 and Sae2, as well as processive long-range resection by Sgs1-Dna2 or Exo1 pathways. Resection mechanisms are highly conserved between yeast and humans, and analogous machineries are found in prokaryotes as well.

Keywords: DNA end resection; DNA endonuclease; DNA helicase; DNA repair; nuclease; protein phosphorylation; recombination.

FIGURE 1.

DNA end resection is required for all recombination processes. The resection of the 5′-terminated DNA strand is required for all recombination pathways, including the SSA, synthesis-dependent strand annealing, and canonical double-strand break repair pathways. DNA end resection prevents mutagenic NHEJ. Microhomology-mediated end-joining was omitted from the scheme and text for simplicity.

FIGURE 2.

DNA end resection of free and blocked DNA ends. a, resection of free (clean) DNA ends. The MRX complex is rapidly recruited to DNA ends upon break formation. The nuclease activity of Mre11 is not required for resection, but the MRX complex has a role to recruit components of the processive pathways that include either Sgs1-Dna2 or Exo1. In some cases, the structural role of the MRX complex can be bypassed. DNA is subsequently resected by either Sgs1-Dna2 or Exo1 in a processive manner. Only a monomer of MRX is depicted for clarity reasons. b, resection of blocked (dirty) DNA ends. The MRX complex is rapidly recruited to DNA ends, which is followed by Sae2. The nuclease activity of Mre11 is required, and it cleaves endonucleolytically the 5′-terminated DNA strand away from the end in a reaction stimulated by phosphorylated (P) Sae2. Furthermore, MRX also likely recruits Sgs1-Dna2 or Exo1 to the endonuclease cut site. The endonuclease cut site provides an entry point for the Sgs1-Dna2 or Exo1 nucleases, which carry out long-range resection. The exonuclease of Mre11 then might degrade DNA in a 3′ → 5′ direction back toward the DNA break.

FIGURE 3.

Mechanism of long-range DNA end resection by Sgs1-Dna2 or Exo1 pathways. a, DNA end resection by Sgs1-Dna2. Sgs1 translocates with a 3′ → 5′ polarity on one DNA strand and unwinds DNA. Unwound ssDNA is coated by RPA, which directs the nucleolytic activity of Dna2 toward the 5′-terminated DNA strand. Whether the 5′ → 3′ motor activity of Dna2 participates in DNA end resection to form a bidirectional helicase remains to be demonstrated. b, DNA end resection by Exo1. The Exo1 nuclease is specific for dsDNA and has a 5′ → 3′ polarity, which directly results in 3′ tailed DNA.