title: Identification of proteins at active, stalled, and collapsed replication forks using isolation of proteins on nascent DNA (iPOND) coupled with mass spectrometry.
abstract: Both DNA and chromatin need to be duplicated during each cell division cycle. Replication happens in the context of defects in the DNA template and other forms of replication stress that present challenges to both genetic and epigenetic inheritance. The replication machinery is highly regulated by replication stress responses to accomplish this goal. To identify important replication and stress response proteins, we combined isolation of proteins on nascent DNA (iPOND) with quantitative mass spectrometry. We identified 290 proteins enriched on newly replicated DNA at active, stalled, and collapsed replication forks. Approximately 16% of these proteins are known replication or DNA damage response proteins. Genetic analysis indicates that several of the newly identified proteins are needed to facilitate DNA replication, especially under stressed conditions. Our data provide a useful resource for investigators studying DNA replication and the replication stress response and validate the use of iPOND combined with mass spectrometry as a discovery tool.
authors.affiliation: From the Department of Biochemistry.
authors.forename: Bianca M
authors.initials: BM
authors.lastname: Sirbu
authors.forename: W Hayes
authors.initials: WH
authors.lastname: McDonald
authors.forename: Huzefa
authors.initials: H
authors.lastname: Dungrawala
authors.forename: Akosua
authors.initials: A
authors.lastname: Badu-Nkansah
authors.forename: Gina M
authors.initials: GM
authors.lastname: Kavanaugh
authors.forename: Yaoyi
authors.initials: Y
authors.lastname: Chen
authors.forename: David L
authors.initials: DL
authors.lastname: Tabb
authors.forename: David
authors.initials: D
authors.lastname: Cortez
mesh_terms: D004247:DNA / Q000096:biosynthesis / Q000378:metabolism*; D004249:DNA Damage; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*
chemical_list: D004268:DNA-Binding Proteins; D004247:DNA
keywords: ATR; Chromatin; DNA Binding Protein; DNA Damage Response; DNA Repair; DNA Replication; SNF2H; SNF2L; iPOND
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