Abstract
Tumor suppressor PTEN regulates cellular activities and controls genome stability through multiple mechanisms. In this study, we report that PTEN is necessary for the protection of DNA replication forks against replication stress. We show that deletion of PTEN leads to replication fork collapse and chromosomal instability upon fork stalling following nucleotide depletion induced by hydroxyurea. PTEN is physically associated with replication protein A 1 (RPA1) via the RPA1 C-terminal domain. STORM and iPOND reveal that PTEN is localized at replication sites and promotes RPA1 accumulation on replication forks. PTEN recruits the deubiquitinase OTUB1 to mediate RPA1 deubiquitination. RPA1 deletion confers a phenotype like that observed in PTEN knockout cells with stalling of replication forks. Expression of PTEN and RPA1 shows strong correlation in colorectal cancer. Heterozygous disruption of RPA1 promotes tumorigenesis in mice. These results demonstrate that PTEN is essential for DNA replication fork protection. We propose that RPA1 is a target of PTEN function in fork protection and that PTEN maintains genome stability through regulation of DNA replication.
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Acknowledgements
We thank L Liang for solving the in silico docking analysis and protein purification, C Song for generating the BRCA2+/− HCT116 cell line, X Du, C Zhang and Y Liu for data analysis and manuscript preparation, Y Li for OTUB1 antibody generation. We thank Dr Chyuan-Sheng Lin for Actin-Flpe and E2A-Cre transgenic mice. We thank J Liang, X Zhao, Q Yin, X Chen, S He, J Zhou, and W Zhao for technical assistance and critical discussions. This study was supported by grants to YY including the National Natural Scientific Foundation of China (NSFC 31420103905, 81430056, 81372491 and 81321003), the China National Major Scientific Program (2010CB912202), the Shu Fan Education and Research Foundation, and Lam Chung Nin Foundation for Systems Biomedicine.
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( Supplementary information is linked to the online version of the paper on the Cell Research website.)
Supplementary information
Supplementary information, Figure S1 (download PDF )
PTEN regulates DNA replication mainly through stalled replication forks protection (PDF 385 kb)
Supplementary information, Figure S2 (download PDF )
PTEN binds to RPA1 mainly through its PTP domain (PDF 338 kb)
Supplementary information, Figure S3 (download PDF )
RPA1 is required for stalled replication forks protection (PDF 264 kb)
Supplementary information, Figure S4 (download PDF )
RPA1 is stabilized by PTEN/OTUB1 and acts as a downstream target (PDF 414 kb)
Supplementary information, Figure S5 (download PDF )
Validation of Rpa1+/− mice (PDF 310 kb)
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Wang, G., Li, Y., Wang, P. et al. PTEN regulates RPA1 and protects DNA replication forks. Cell Res 25, 1189–1204 (2015). https://doi.org/10.1038/cr.2015.115
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DOI: https://doi.org/10.1038/cr.2015.115
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