Related Experiment Video
Updated: Dec 11, 2025
Capturing Common Fragile Site Breaks by Native γH2A.X ChIP
Published on: January 24, 2025
Degradation of 5hmC-marked stalled replication forks by APE1 causes genomic instability
Suhas S Kharat1, Xia Ding1, Divya Swaminathan1
1Mouse Cancer Genetics Program, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA.
Abstract:
Synthetic lethality between poly(ADP-ribose) polymerase (PARP) inhibition and BRCA deficiency is exploited to treat breast and ovarian tumors. However, resistance to PARP inhibitors (PARPis) is common. To identify potential resistance mechanisms, we performed a genome-wide RNAi screen in BRCA2-deficient mouse embryonic stem cells and validation in KB2P1.21 mouse mammary tumor cells. We found that resistance to multiple PARPi emerged with reduced expression of TET2 (ten-eleven translocation), which promotes DNA demethylation by oxidizing 5-methylcytosine (5mC) to 5-hydroxymethycytosine (5hmC) and other products. TET2 knockdown in BRCA2-deficient cells protected stalled replication forks (RFs). Increasing 5hmC abundance induced the degradation of stalled RFs in KB2P1.21 and human cancer cells by recruiting the base excision repair-associated apurinic/apyrimidinic endonuclease APE1, independent of the BRCA2 status. TET2 loss did not affect the recruitment of the repair protein RAD51 to sites of double-strand breaks (DSBs) or the abundance of proteins associated with RF integrity. The loss of TET2, of its product 5hmC, and of APE1 recruitment to stalled RFs promoted resistance to the chemotherapeutic cisplatin. Our findings reveal a previously unknown role for the epigenetic mark 5hmC in maintaining the integrity of stalled RFs and a potential resistance mechanism to PARPi and cisplatin.
Insights
Resistance to PARP inhibitors (PARPis) in cancer can emerge from reduced TET2 expression. TET2 loss protects stalled replication forks, potentially explaining PARPi resistance and impacting cisplatin efficacy.
Area of Science:
- Epigenetics
- DNA repair
- Cancer biology
Background:
- Synthetic lethality exploiting PARP inhibition and BRCA deficiency is a key cancer therapy.
- Acquired resistance to PARP inhibitors (PARPis) limits their clinical efficacy.
- Understanding resistance mechanisms is crucial for improving cancer treatment outcomes.
Purpose of the Study:
- To identify novel mechanisms of resistance to PARP inhibitors (PARPis).
- To investigate the role of TET2 and its epigenetic products in DNA replication fork integrity and drug resistance.
Main Methods:
- Genome-wide RNAi screening in BRCA2-deficient mouse embryonic stem cells.
- Validation in mouse mammary tumor cells and human cancer cells.
- Analysis of TET2 expression, 5-hydroxymethycytosine (5hmC) levels, and protein recruitment to stalled replication forks.
Main Results:
- Reduced TET2 expression was identified as a resistance mechanism to multiple PARPis.
- TET2 loss protected stalled replication forks (RFs) in BRCA2-deficient cells.
- Increased 5hmC abundance promoted stalled RF degradation via APE1 recruitment, conferring resistance to PARPis and cisplatin.
Conclusions:
- TET2 and its epigenetic product 5hmC play a critical role in maintaining stalled replication fork integrity.
- Loss of TET2 and 5hmC, along with impaired APE1 recruitment, represents a novel resistance mechanism to PARPis and cisplatin.
- These findings offer new insights into epigenetic regulation of DNA repair and drug resistance in cancer therapy.
More Related Videos
Related Concept Videos
02:37Restarting Stalled Replication Forks
02:37Restarting Stalled Replication Forks
02:37DNA Damage can Stall the Cell Cycle
02:37DNA Damage Can Stall the Cell Cycle
01:02The DNA Replication Fork
02:31Homologous Recombination