Accumulation of FFA-1, the Xenopus Homolog of Werner Helicase, and DNA Polymerase δ on Chromatin in Response to Replication Fork Arrest

• Published in: Journal of biochemistry (Tokyo) Vol. 140; no. 1; pp. 95 - 103
• Main Authors: Sasakawa, Noriko, Fukui, Tomoyuki, Waga, Shou
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01-07-2006
• Subjects: Adenosine Triphosphatases - metabolism; Animals; Aphidicolin - pharmacology; Cell Cycle Proteins - pharmacology; Chromatin - drug effects; Chromatin - metabolism; Deoxycytosine Nucleotides - pharmacology; DNA Damage; DNA Helicases - metabolism; DNA Polymerase II - metabolism; DNA Polymerase III - metabolism; DNA polymerase δ; DNA polymerases δ and ε; DNA Replication - drug effects; DNA-Binding Proteins - metabolism; double-strand break; DSB; Female; FFA-1; focus-forming activity 1; Geminin; Male; MCM; minichromosome maintenance protein; Oocytes - drug effects; Oocytes - metabolism; ORC; origin recognition complex; PCNA; Polδ and ε; pre-RC; pre-replicative complex; proliferating cell nuclear antigen; Proliferating Cell Nuclear Antigen - metabolism; RecQ Helicases; replication fork arrest; replication protein A; Replication Protein A - metabolism; RPA; Spermatozoa - metabolism; Werner helicase; Xenopus; Xenopus egg extract; Xenopus Proteins - metabolism
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/jb/mvj130

Werner syndrome is a genetic disorder characterized by premature aging and cancer-prone symptoms, and is caused by mutation of the WRN gene. WRN is a member of the RecQ helicase family and is thought to function in processes implicated in DNA replication and repair to maintain genome stability; however, its precise function is still unclear. We found that replication fork arrest markedly enhances chromatin binding of focus-forming activity 1 (FFA-1), a Xenopus WRN homolog, in Xenopus egg extracts. In addition to FFA-1, DNA polymerase δ (Polδ) and replication protein A, but not DNA polymerase ε and proliferating cell nuclear antigen, accumulated increasingly on replication-arrested chromatin. Elevated accumulation of these proteins was dependent on formation of pre-replicative complexes (pre-RCs). Double-strand break (DSB) formation also enhanced chromatin binding of FFA-1, but not Polδ, independently of pre-RC formation. In contrast to FFA-1, chromatin binding of Xenopus Bloom syndrome helicase (xBLM) only slightly increased after replication arrest or DSB formation. Thus, WRN-specific, distinct processes can be reproduced in the in vitro system in egg extracts, and this system is useful for biochemical analysis of WRN functions during DNA metabolism.