The Drosophila Werner exonuclease participates in an exonuclease-independent response to replication stress.
Members of the RecQ family of helicases are known for their roles in DNA repair, replication, and recombination. Mutations in the human RecQ helicases, WRN and BLM, cause Werner and Bloom syndromes, which are diseases characterized by genome instability and an increased risk of cancer. While WRN contains both a helicase and an exonuclease domain, the Drosophila melanogaster homolog, WRNexo, contains only the exonuclease domain. Therefore the Drosophila model system provides a unique opportunity to study the exonuclease functions of WRN separate from the helicase. We created a null allele of WRNexo via imprecise P-element excision. The null WRNexo mutants are not sensitive to double-strand break-inducing reagents, suggesting that the exonuclease does not play a key role in homologous recombination-mediated repair of DSBs. However, WRNexo mutant embryos have a reduced hatching frequency and larvae are sensitive to the replication fork-stalling reagent, hydroxyurea (HU), suggesting that WRNexo is important in responding to replication stress. The role of WRNexo in the HU-induced stress response is independent of Rad51. Interestingly, the hatching defect and HU sensitivity of WRNexo mutants do not occur in flies containing an exonuclease-dead copy of WRNexo, suggesting that the role of WRNexo in replication is independent of exonuclease activity. Additionally, WRNexo and Blm mutants exhibit similar sensitivity to HU and synthetic lethality in combination with mutations in structure-selective endonucleases. We propose that WRNexo and BLM interact to promote fork reversal following replication fork stalling and in their absence regressed forks are restarted through a Rad51-mediated process.
Department of Biology, Tufts University, Medford, Massachusetts 02155 Training in Education and Critical Research Skills (TEACRS) Program, Department of Physical Sciences and Mathematics, Mount St. Mary's College, Los Angeles, California elyse.bolterstein@tufts.edu mitch.mcvey@tufts.edu.
Elyse
E
Bolterstein
Department of Biology, Tufts University, Medford, Massachusetts 02155.
Rachel
R
Rivero
Department of Physical Sciences and Mathematics, Mount St. Mary's College, Los Angeles, California 90049.
Melissa
M
Marquez
Department of Biology, Tufts University, Medford, Massachusetts 02155 Training in Education and Critical Research Skills (TEACRS) Program, Department of Physical Sciences and Mathematics, Mount St. Mary's College, Los Angeles, California Program in Genetics, Tufts Sackler School of Graduate Biomedical Sciences, Boston, Massachusetts 02111 elyse.bolterstein@tufts.edu mitch.mcvey@tufts.edu.
Mitch
M
McVey
D000818:Animals; D053903:DNA Breaks, Double-Stranded; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism; D004260:DNA Repair; D004261:DNA Replication; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000201:enzymology* / Q000235:genetics; D005092:Exonucleases / Q000235:genetics / Q000378:metabolism*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D006918:Hydroxyurea; D008297:Male; D009154:Mutation; D051135:Rad51 Recombinase / Q000235:genetics / Q000378:metabolism; D053484:RecQ Helicases / Q000235:genetics / Q000378:metabolism*
D029721:Drosophila Proteins; D051135:Rad51 Recombinase; C495149:spn-A protein, Drosophila; D005092:Exonucleases; C542170:WRNexo protein, Drosophila; D004265:DNA Helicases; D053484:RecQ Helicases; C470639:blm protein, Drosophila; D006918:Hydroxyurea
DNA replication; Werner syndrome; double-strand break; replication restart