Rad52/Rad59-dependent recombination as a means to rectify faulty Okazaki fragment processing.
The correct removal of 5'-flap structures by Rad27 and Dna2 during Okazaki fragment maturation is crucial for the stable maintenance of genetic materials and cell viability. In this study, we identified RAD52, a key recombination protein, as a multicopy suppressor of dna2-K1080E, a lethal helicase-negative mutant allele of DNA2 in yeasts. In contrast, the overexpression of Rad51, which works conjointly with Rad52 in canonical homologous recombination, failed to suppress the growth defect of the dna2-K1080E mutation, indicating that Rad52 plays a unique and distinct role in Okazaki fragment metabolism. We found that the recombination-defective Rad52-QDDD/AAAA mutant did not rescue dna2-K1080E, suggesting that Rad52-mediated recombination is important for suppression. The Rad52-mediated enzymatic stimulation of Dna2 or Rad27 is not a direct cause of suppression observed in vivo, as both Rad52 and Rad52-QDDD/AAAA proteins stimulated the endonuclease activities of both Dna2 and Rad27 to a similar extent. The recombination mediator activity of Rad52 was dispensable for the suppression, whereas both the DNA annealing activity and its ability to interact with Rad59 were essential. In addition, we found that several cohesion establishment factors, including Rsc2 and Elg1, were required for the Rad52-dependent suppression of dna2-K1080E. Our findings suggest a novel Rad52/Rad59-dependent, but Rad51-independent recombination pathway that could ultimately lead to the removal of faulty flaps in conjunction with cohesion establishment factors.
From the Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea and.
Miju
M
Lee
From the Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea and.
Chul-Hwan
CH
Lee
From the Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea and.
Annie Albert
AA
Demin
From the Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea and.
Palinda Ruvan
PR
Munashingha
From the Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea and.
Tamir
T
Amangyeld
From the Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea and.
Buki
B
Kwon
the Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, Utah 84112.
Tim
T
Formosa
From the Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea and yeonsooseo@kaist.ac.kr.
Yeon-Soo
YS
Seo
D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism; D004260:DNA Repair; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D059765:Homologous Recombination; D015151:Immunoblotting; D009154:Mutation; D051721:Rad52 DNA Repair and Recombination Protein / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; C022262:Okazaki fragments; C497197:RAD52 protein, S cerevisiae; C400213:RAD59 protein, S cerevisiae; D051721:Rad52 DNA Repair and Recombination Protein; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA; D004265:DNA Helicases; C410222:DNA2 protein, S cerevisiae
DNA Helicase; DNA Recombination; DNA Repair; DNA Replication; Dna2; Genome Integrity; Homologous Recombination; Lagging Strand Synthesis; Rad52; Rad59