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DNA replication is initiated at sites containing predefined DNA sequences known as origins of replication. DNA is unwound at these sites by the minichromosome maintenance (MCM) helicase and other factors such as Cdc45 and the associated GINS complex.The unwound single strands are protected by replication protein A (RPA) until DNA polymerase starts synthesizing DNA at the 5’ end of the strand in the same direction as the replication fork. To prevent the replication fork from falling apart, a...

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Quantifying Replication Stress in Ovarian Cancer Cells Using Single-Stranded DNA Immunofluorescence

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Brc1-dependent recovery from replication stress.

Kirstin L Bass¹, Johanne M Murray, Matthew J O'Connell

¹Department of Oncological Sciences, Mount Sinai School of Medicine, New York, NY 10029, USA.

Journal of Cell Science

|February 28, 2012

Summary

This summary is machine-generated.

BRCT-containing protein 1 (Brc1) is crucial for recovery from replication stress in fission yeast. Cells lacking Brc1 recruit Rad52 to stalled replication forks, but this does not trigger DNA repair or recombination pathways.

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Assessment of Global DNA Double-Strand End Resection using BrdU-DNA Labeling coupled with Cell Cycle Discrimination Imaging

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Published on: April 28, 2021

Area of Science:

Molecular and Cell Biology
DNA Replication and Repair
Genetics and Genomics

Background:

BRCT-containing protein 1 (Brc1) in Schizosaccharomyces pombe is vital for surviving chronic replicative stress.
The precise role of Brc1 in either DNA repair or replication fidelity remains unclear.

Purpose of the Study:

To elucidate the function of Brc1 in response to replication stress.
To investigate the nature of Rad52 foci formation in brc1Δ cells under replication stress.

Main Methods:

Analysis of replication pausing recovery in brc1Δ cells.
Investigation of Rad52 foci formation and composition (RPA, Rad51, γH2A).
Assessment of recombination and post-replication repair.
Study of origin recognition complex (ORC) function in Brc1-deficient cells under replication stress.

Main Results:

brc1Δ cells exhibit delayed recovery from replication pausing without activating DNA damage checkpoints.
Rad52 is required for brc1Δ cell viability and forms RPA-positive foci under replication stress, but lacks Rad51 and γH2A.
These Rad52 foci are not associated with increased recombination or post-replication repair and are dependent on ORC function.

Conclusions:

Brc1-deficient cells employ an ORC-dependent mechanism to rescue replication stress, involving dormant origin firing and Rad52 recruitment.
The observed Rad52 foci represent structures that stabilize stalled forks but do not signal DNA damage requiring recombination or checkpoint activation.
This study clarifies Brc1's role in managing replication stress, distinct from canonical DNA damage response pathways.