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The Caenorhabditis elegans homolog of Gen1/Yen1 resolvases links DNA damage signaling to DNA double-strand break repair.

Bailly AP, Freeman A, Hall J, Déclais AC, Alpi A, Lilley DM, Ahmed S, Gartner A - PLoS Genet. (2010)

Bottom Line: DNA double-strand breaks (DSBs) can be repaired by homologous recombination (HR), which can involve Holliday junction (HJ) intermediates that are ultimately resolved by nucleolytic enzymes.An N-terminal fragment of human GEN1 has recently been shown to act as a Holliday junction resolvase, but little is known about the role of GEN-1 in vivo.Furthermore, GEN-1 acts redundantly with the 9-1-1 complex to ensure genome stability.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Gene Regulation and Expression, University of Dundee, Dundee, United Kingdom.

ABSTRACT
DNA double-strand breaks (DSBs) can be repaired by homologous recombination (HR), which can involve Holliday junction (HJ) intermediates that are ultimately resolved by nucleolytic enzymes. An N-terminal fragment of human GEN1 has recently been shown to act as a Holliday junction resolvase, but little is known about the role of GEN-1 in vivo. Holliday junction resolution signifies the completion of DNA repair, a step that may be coupled to signaling proteins that regulate cell cycle progression in response to DNA damage. Using forward genetic approaches, we identified a Caenorhabditis elegans dual function DNA double-strand break repair and DNA damage signaling protein orthologous to the human GEN1 Holliday junction resolving enzyme. GEN-1 has biochemical activities related to the human enzyme and facilitates repair of DNA double-strand breaks, but is not essential for DNA double-strand break repair during meiotic recombination. Mutational analysis reveals that the DNA damage-signaling function of GEN-1 is separable from its role in DNA repair. GEN-1 promotes germ cell cycle arrest and apoptosis via a pathway that acts in parallel to the canonical DNA damage response pathway mediated by RPA loading, CHK1 activation, and CEP-1/p53-mediated apoptosis induction. Furthermore, GEN-1 acts redundantly with the 9-1-1 complex to ensure genome stability. Our study suggests that GEN-1 might act as a dual function Holliday junction resolvase that may coordinate DNA damage signaling with a late step in DNA double-strand break repair.

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L1 stage worms IR sensitivity assay with gen-1 and gen-1 atm-1 double mutants.Assays were performed as described in Figure 3. Error bars represent s.e.m.
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pgen-1001025-g007: L1 stage worms IR sensitivity assay with gen-1 and gen-1 atm-1 double mutants.Assays were performed as described in Figure 3. Error bars represent s.e.m.

Mentions: We next wished to determine genetic interactions between GEN-1, ATR and ATM PI3-like kinases, which are predicted to act upstream of the 9-1-1 complex in DNA damage signaling. Given that an atl-1(tm853) deletion leads to excessive genome instability in germ cells and concomitant sterility [61], we could not assess the possibility of enhanced IR sensitivity in gen-1 atl-1 double mutants. In contrast, C. elegans atm-1 plays a minor role in DNA damage signaling and atm-1(gk186) results in partial defect in IR-induced cell cycle arrest and apoptosis [62]. Consistent with this notion, we found that the atm-1(gk186) deletion is not hypersensitive to IR when subjected to the L4 IR survival assay, and that the IR sensitivity is not enhanced by the gen-1(yp30) mutant (Figure S10). In contrast, the atm-1(gk186) mutant is sensitive to IR in the L1 assay, and IR sensitivity is enhanced in combination with both gen-1(tm2940) and gen-1(yp30) (Figure 7).


The Caenorhabditis elegans homolog of Gen1/Yen1 resolvases links DNA damage signaling to DNA double-strand break repair.

Bailly AP, Freeman A, Hall J, Déclais AC, Alpi A, Lilley DM, Ahmed S, Gartner A - PLoS Genet. (2010)

L1 stage worms IR sensitivity assay with gen-1 and gen-1 atm-1 double mutants.Assays were performed as described in Figure 3. Error bars represent s.e.m.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2908289&req=5

pgen-1001025-g007: L1 stage worms IR sensitivity assay with gen-1 and gen-1 atm-1 double mutants.Assays were performed as described in Figure 3. Error bars represent s.e.m.
Mentions: We next wished to determine genetic interactions between GEN-1, ATR and ATM PI3-like kinases, which are predicted to act upstream of the 9-1-1 complex in DNA damage signaling. Given that an atl-1(tm853) deletion leads to excessive genome instability in germ cells and concomitant sterility [61], we could not assess the possibility of enhanced IR sensitivity in gen-1 atl-1 double mutants. In contrast, C. elegans atm-1 plays a minor role in DNA damage signaling and atm-1(gk186) results in partial defect in IR-induced cell cycle arrest and apoptosis [62]. Consistent with this notion, we found that the atm-1(gk186) deletion is not hypersensitive to IR when subjected to the L4 IR survival assay, and that the IR sensitivity is not enhanced by the gen-1(yp30) mutant (Figure S10). In contrast, the atm-1(gk186) mutant is sensitive to IR in the L1 assay, and IR sensitivity is enhanced in combination with both gen-1(tm2940) and gen-1(yp30) (Figure 7).

Bottom Line: DNA double-strand breaks (DSBs) can be repaired by homologous recombination (HR), which can involve Holliday junction (HJ) intermediates that are ultimately resolved by nucleolytic enzymes.An N-terminal fragment of human GEN1 has recently been shown to act as a Holliday junction resolvase, but little is known about the role of GEN-1 in vivo.Furthermore, GEN-1 acts redundantly with the 9-1-1 complex to ensure genome stability.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Gene Regulation and Expression, University of Dundee, Dundee, United Kingdom.

ABSTRACT
DNA double-strand breaks (DSBs) can be repaired by homologous recombination (HR), which can involve Holliday junction (HJ) intermediates that are ultimately resolved by nucleolytic enzymes. An N-terminal fragment of human GEN1 has recently been shown to act as a Holliday junction resolvase, but little is known about the role of GEN-1 in vivo. Holliday junction resolution signifies the completion of DNA repair, a step that may be coupled to signaling proteins that regulate cell cycle progression in response to DNA damage. Using forward genetic approaches, we identified a Caenorhabditis elegans dual function DNA double-strand break repair and DNA damage signaling protein orthologous to the human GEN1 Holliday junction resolving enzyme. GEN-1 has biochemical activities related to the human enzyme and facilitates repair of DNA double-strand breaks, but is not essential for DNA double-strand break repair during meiotic recombination. Mutational analysis reveals that the DNA damage-signaling function of GEN-1 is separable from its role in DNA repair. GEN-1 promotes germ cell cycle arrest and apoptosis via a pathway that acts in parallel to the canonical DNA damage response pathway mediated by RPA loading, CHK1 activation, and CEP-1/p53-mediated apoptosis induction. Furthermore, GEN-1 acts redundantly with the 9-1-1 complex to ensure genome stability. Our study suggests that GEN-1 might act as a dual function Holliday junction resolvase that may coordinate DNA damage signaling with a late step in DNA double-strand break repair.

Show MeSH
Related in: MedlinePlus