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Updated: Jul 30, 2025

Quantifying Replication Stress in Ovarian Cancer Cells Using Single-Stranded DNA Immunofluorescence
Published on: February 10, 2023
p130RB2 positively contributes to ATR activation in response to replication stress via the RPA32-ETAA1 axis
Chiharu Uchida1, Hiroyuki Niida2, Satoshi Sakai2
1Advanced Research Facilities & Services, Preeminent Medical Photonics Education & Research Center, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka 431-3192, Japan.
p130RB2 protein is essential for activating Ataxia-telangiectasia mutated and Rad3-related (ATR) kinase during DNA replication stress. Its absence disrupts cell cycle progression and survival, highlighting its role in maintaining genome integrity.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- Ataxia-telangiectasia mutated and Rad3-related (ATR) kinase is vital for cell cycle control during DNA replication stress.
- ATR is activated via RPA32-ETAA1 and TopBP1 pathways, but the RPA32-ETAA1 mechanism is not fully understood.
Purpose of the Study:
- To elucidate the role of p130RB2 in the RPA32-ETAA1-mediated ATR activation pathway.
- To investigate the impact of p130RB2 on cell cycle progression and genome stability under replication stress.
Main Methods:
- Hydroxyurea-induced DNA replication stress model in cells.
- Co-immunoprecipitation assays to study protein interactions (p130RB2, ETAA1, TopBP1).
- Western blotting to assess ATR activation and target phosphorylation (RPA32, Chk1, ATR).
- Analysis of S phase re-progression, anaphase bridges, and cell survival.
Main Results:
- p130RB2 interacts with ETAA1 but not TopBP1.
- Depletion of p130RB2 impairs RPA32-ETAA1 interaction and ATR activation, including target phosphorylation.
- p130RB2 deficiency leads to aberrant S phase re-progression, increased anaphase bridges, and reduced cell survival.
- Restoration of p130RB2 rescues these phenotypes.
Conclusions:
- p130RB2 positively regulates the RPA32-ETAA1-ATR signaling axis.
- p130RB2 is crucial for proper cell cycle re-progression and maintaining genome integrity following replication stress.
- p130RB2 is a key component in the cellular response to DNA replication stress.
Related Concept Videos
DNA Damage can Stall the Cell Cycle
Restarting Stalled Replication Forks
MAPK Signaling Cascades
Regulation of the Unfolded Protein Response
Negative Regulator Molecules
The Unfolded Protein Response

