title: Drosophila RecQ4 is directly involved in both DNA replication and the response to UV damage in S2 cells.
abstract: The RecQ4 protein shows homology to both the S.cerevisiae DNA replication protein Sld2 and the DNA repair related RecQ helicases. Experimental data also suggest replication and repair functions for RecQ4, but the precise details of its involvement remain to be clarified.Here we show that depletion of DmRecQ4 by dsRNA interference in S2 cells causes defects consistent with a replication function for the protein. The cells show reduced proliferation associated with an S phase block, reduced BrdU incorporation, and an increase in cells with a subG1 DNA content. At the molecular level we observe reduced chromatin association of DNA polymerase-alpha and PCNA. We also observe increased chromatin association of phosphorylated H2AvD--consistent with the presence of DNA damage and increased apoptosis.Analysis of DmRecQ4 repair function suggests a direct role in NER, as the protein shows rapid but transient nuclear localisation after UV treatment. Re-localisation is not observed after etoposide or H₂O₂ treatment, indicating that the involvement of DmRecQ4 in repair is likely to be pathway specific.Deletion analysis of DmRecQ4 suggests that the SLD2 domain was essential, but not sufficient, for replication function. In addition a DmRecQ4 N-terminal deletion could efficiently re-localise on UV treatment, suggesting that the determinants for this response are contained in the C terminus of the protein. Finally several deletions show differential rescue of dsRNA generated replication and proliferation phenotypes. These will be useful for a molecular analysis of the specific role of DmRecQ4 in different cellular pathways.
authors.affiliation: Department Basic Medical Sciences, St. Georges University London, Cranmer Terrace, London, United Kingdom.
authors.forename: Gilles
authors.initials: G
authors.lastname: Crevel
authors.forename: Nicole
authors.initials: N
authors.lastname: Vo
authors.forename: Isabelle
authors.initials: I
authors.lastname: Crevel
authors.forename: Sana
authors.initials: S
authors.lastname: Hamid
authors.forename: Lily
authors.initials: L
authors.lastname: Hoa
authors.forename: Seiji
authors.initials: S
authors.lastname: Miyata
authors.forename: Sue
authors.initials: S
authors.lastname: Cotterill
mesh_terms: D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism / Q000528:radiation effects; D049109:Cell Proliferation / Q000528:radiation effects; D004249:DNA Damage / Q000528:radiation effects; D004261:DNA Replication / Q000528:radiation effects; D004330:Drosophila / Q000235:genetics* / Q000378:metabolism* / Q000528:radiation effects; D004789:Enzyme Activation; D005047:Etoposide / Q000494:pharmacology; D015870:Gene Expression; D006861:Hydrogen Peroxide / Q000494:pharmacology; D009154:Mutation; D054730:Protein Interaction Domains and Motifs; D021381:Protein Transport / Q000187:drug effects / Q000528:radiation effects; D012330:RNA, Double-Stranded / Q000235:genetics / Q000378:metabolism; D053484:RecQ Helicases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016196:S Phase / Q000528:radiation effects; D014466:Ultraviolet Rays / Q000009:adverse effects*
chemical_list: D012330:RNA, Double-Stranded; D005047:Etoposide; D006861:Hydrogen Peroxide; D053484:RecQ Helicases