title: The Werner and Bloom syndrome proteins help resolve replication blockage by converting (regressed) holliday junctions to functional replication forks.
abstract: Cells cope with blockage of replication fork progression in a manner that allows DNA synthesis to be completed and genomic instability minimized. Models for resolution of blocked replication involve fork regression to form Holliday junction structures. The human RecQ helicases WRN and BLM (deficient in Werner and Bloom syndromes, respectively) are critical for maintaining genomic stability and thought to function in accurate resolution of replication blockage. Consistent with this notion, WRN and BLM localize to sites of blocked replication after certain DNA-damaging treatments and exhibit enhanced activity on replication and recombination intermediates. Here we examine the actions of WRN and BLM on a special Holliday junction substrate reflective of a regressed replication fork. Our results demonstrate that, in reactions requiring ATP hydrolysis, both WRN and BLM convert this Holliday junction substrate primarily to a four-stranded replication fork structure, suggesting they target the Holliday junction to initiate branch migration. In agreement, the Holliday junction binding protein RuvA inhibits the WRN- and BLM-mediated conversion reactions. Importantly, this conversion product is suitable for replication with its leading daughter strand readily extended by DNA polymerases. Furthermore, binding to and conversion of this Holliday junction are optimal at low MgCl(2) concentrations, suggesting that WRN and BLM preferentially act on the square planar (open) conformation of Holliday junctions. Our findings suggest that, subsequent to fork regression events, WRN and/or BLM could re-establish functional replication forks to help overcome fork blockage. Such a function is highly consistent with phenotypes associated with WRN- and BLM-deficient cells.
authors: Graduate Center for Toxicology and Markey Cancer Center, University of Kentucky College of Medicine, Lexington, Kentucky 40536, United States.
authors: Amrita
authors: A
authors: Machwe
authors: Rajashree
authors: R
authors: Karale
authors: Xioahua
authors: X
authors: Xu
authors: Yilun
authors: Y
authors: Liu
authors: David K
authors: DK
authors: Orren
mesh_terms: D004247:DNA / Q000737:chemistry*; D004261:DNA Replication; D024202:Electrophoretic Mobility Shift Assay; D005090:Exodeoxyribonucleases / Q000737:chemistry / Q000502:physiology*; D006801:Humans; D053484:RecQ Helicases / Q000737:chemistry / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D000071657:Werner Syndrome Helicase
chemical_list: D011994:Recombinant Proteins; D004247:DNA; D005090:Exodeoxyribonucleases; C097151:Bloom syndrome protein; D053484:RecQ Helicases; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase