Cell Reports
Volume 28, Issue 13, 24 September 2019, Pages 3497-3509.e4
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Functional Analysis of the Replication Fork Proteome Identifies BET Proteins as PCNA Regulators

https://doi.org/10.1016/j.celrep.2019.08.051Get rights and content
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Highlights

  • iPOND proteomics identifies 593 proteins enriched on nascent DNA

  • 101 proteins are found to be depleted from nascent DNA compared to bulk chromatin

  • Loss-of-function and bioinformatics analyses provide functional validation

  • The BET proteins BRD2, BRD3, and BRD4 bind ATAD5 and regulate PCNA unloading

Summary

Identifying proteins that function at replication forks is essential to understanding DNA replication, chromatin assembly, and replication-coupled DNA repair mechanisms. Combining quantitative mass spectrometry in multiple cell types with stringent statistical cutoffs, we generated a high-confidence catalog of 593 proteins that are enriched at replication forks and nascent chromatin. Loss-of-function genetic analyses indicate that 85% yield phenotypes that are consistent with activities in DNA and chromatin replication or already have described functions in these processes. We illustrate the value of this resource by identifying activities of the BET family proteins BRD2, BRD3, and BRD4 in controlling DNA replication. These proteins use their extra-terminal domains to bind and inhibit the ATAD5 complex and thereby control the amount of PCNA on chromatin.

Keywords

DNA replication
iPOND
ATAD5
BET domain
PCNA
chromatin
replisome
proteomics

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