title: The homologous recombination component EEPD1 is required for genome stability in response to developmental stress of vertebrate embryogenesis.
abstract: Stressed replication forks can be conservatively repaired and restarted using homologous recombination (HR), initiated by nuclease cleavage of branched structures at stalled forks. We previously reported that the 5' nuclease EEPD1 is recruited to stressed replication forks, where it plays critical early roles in HR initiation by promoting fork cleavage and end resection. HR repair of stressed replication forks prevents their repair by non-homologous end-joining (NHEJ), which would cause genome instability. Rapid cell division during vertebrate embryonic development generates enormous pressure to maintain replication speed and accuracy. To determine the role of EEPD1 in maintaining replication fork integrity and genome stability during rapid cell division in embryonic development, we assessed the role of EEPD1 during zebrafish embryogenesis. We show here that when EEPD1 is depleted, zebrafish embryos fail to develop normally and have a marked increase in death rate. Zebrafish embryos depleted of EEPD1 are far more sensitive to replication stress caused by nucleotide depletion. We hypothesized that the HR defect with EEPD1 depletion would shift repair of stressed replication forks to unopposed NHEJ, causing chromosome abnormalities. Consistent with this, EEPD1 depletion results in nuclear defects including anaphase bridges and micronuclei in stressed zebrafish embryos, similar to BRCA1 deficiency. These results demonstrate that the newly characterized HR protein EEPD1 maintains genome stability during embryonic replication stress. These data also imply that the rapid cell cycle transit seen during embryonic development produces replication stress that requires HR to resolve.
authors.affiliation: a Division of Hematology/Oncology , Department of Medicine, University of Florida Health , Gainesville , FL , USA.
authors.forename: Changzoon
authors.initials: C
authors.lastname: Chun
authors.affiliation: a Division of Hematology/Oncology , Department of Medicine, University of Florida Health , Gainesville , FL , USA.
authors.forename: Yuehan
authors.initials: Y
authors.lastname: Wu
authors.affiliation: b Department of Biochemistry and Molecular Biology , Indiana University School of Medicine , Indianapolis , IN , USA.
authors.forename: Suk-Hee
authors.initials: SH
authors.lastname: Lee
authors.affiliation: a Division of Hematology/Oncology , Department of Medicine, University of Florida Health , Gainesville , FL , USA.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Williamson
authors.affiliation: a Division of Hematology/Oncology , Department of Medicine, University of Florida Health , Gainesville , FL , USA.
authors.forename: Brian L
authors.initials: BL
authors.lastname: Reinert
authors.affiliation: a Division of Hematology/Oncology , Department of Medicine, University of Florida Health , Gainesville , FL , USA.
authors.forename: Aruna Shanker
authors.initials: AS
authors.lastname: Jaiswal
authors.affiliation: c Department of Environmental and Radiological Health Sciences , Colorado State University , Fort Collins , CO , USA.
authors.forename: Jac A
authors.initials: JA
authors.lastname: Nickoloff
authors.affiliation: a Division of Hematology/Oncology , Department of Medicine, University of Florida Health , Gainesville , FL , USA.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Hromas
mesh_terms: D000818:Animals; D004261:DNA Replication; D047108:Embryonic Development / Q000235:genetics*; D004706:Endodeoxyribonucleases / Q000502:physiology*; D042822:Genomic Instability; D015398:Signal Transduction; D015027:Zebrafish / Q000196:embryology / Q000235:genetics / Q000378:metabolism; D029961:Zebrafish Proteins / Q000502:physiology*
chemical_list: D029961:Zebrafish Proteins; C000614723:EEPD1 protein, zebrafish; D004706:Endodeoxyribonucleases