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J Biol Chem. 2013 Nov 1;288(44):31458-67. doi: 10.1074/jbc.M113.511337. Epub 2013 Sep 18.

Identification of proteins at active, stalled, and collapsed replication forks using isolation of proteins on nascent DNA (iPOND) coupled with mass spectrometry.

Author information

1
From the Department of Biochemistry.

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.

KEYWORDS:

ATR; Chromatin; DNA Binding Protein; DNA Damage Response; DNA Repair; DNA Replication; SNF2H; SNF2L; iPOND

PMID:
24047897
PMCID:
PMC3814742
DOI:
10.1074/jbc.M113.511337
[Indexed for MEDLINE]
Free PMC Article

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