A phosphorylation–deubiquitination cascade regulates the BRCA2–RAD51 axis in homologous recombination

Luo, Kuntian; Li, Lei; Li, Yunhui; Wu, Chenming; Yin, Yujiao; Chen, Yuping; Deng, Min; Nowsheen, Somaira; Yuan, Jian; Lou, Zhenkun

doi:10.1101/gad.289439.116

Cited by 75 publications

(77 citation statements)

References 97 publications

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“…Hence we undertook GST pulldown assays and found the interaction between DDB2 and Rad51, suggesting that DDB2 stabilizes Rad51 by inhibiting its ubiquitination (Figure G). In previous studies, RING1, UCHL3, UAF1, FBH1 UBL1, UBC9, and BLM were reported to manipulate the ubiquitination of Rad51 by direct interaction. Therefore, using immunoprecipitation assays, we checked whether any of these proteins was involved in this process.…”

Section: Resultsmentioning

confidence: 94%

Depletion of DNA damage binding protein 2 sensitizes triple‐negative breast cancer cells to poly ADP‐ribose polymerase inhibition by destabilizing Rad51

Zhao

et al. 2019

Cancer Science

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Poly ADP‐ribose polymerase inhibitors (PARPi) have shown promising therapeutic efficacy in triple‐negative breast cancer (TNBC) patients. However, resistance ultimately develops, preventing a curative effect from being attained. Extensive investigations have indicated the diversity in the mechanisms underlying the PARPi sensitivity of breast cancer. In this study, we found that DNA damage binding protein 2 (DDB2), a DNA damage‐recognition factor, could protect TNBC cells from PARPi by regulating DNA double‐strand break repair through the homologous recombination pathway, whereas the depletion of DDB2 sensitizes TNBC cells to PARPi. Furthermore, we found that DDB2 was able to stabilize Rad51 by physical association and disrupting its ubiquitination pathway‐induced proteasomal degradation. These findings highlight an essential role of DDB2 in modulating homologous recombination pathway activity and suggest a promising therapeutic target for TNBC.

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Section: Resultsmentioning

confidence: 94%

Depletion of DNA damage binding protein 2 sensitizes triple‐negative breast cancer cells to poly ADP‐ribose polymerase inhibition by destabilizing Rad51

Zhao

et al. 2019

Cancer Science

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“…Additional possibilities may involve RADX post-translational modifications that either reduce interactions between RADX monomers and/or reduce the affinity of the RADX OB-folds for ssDNA. In direct analogy to RADX, both RAD51 and RPA are phosphorylated and SUMOylated throughout the cell cycle and in response to DNA damage (50)(51)(52). Another open question is the interplay between BRCA2/RAD51 and RADX.…”

Section: Discussionmentioning

confidence: 99%

RADX condenses single-stranded DNA to antagonize RAD51 loading

Zhang

Schaub

Finkelstein

2020

Preprint

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RADX is a mammalian single-stranded DNA-binding protein that stabilizes telomeres and stalled replication forks. Cellular biology studies have shown that the balance between RADX and Replication Protein A (RPA) activities is critical for DNA replication integrity. RADX is also a negative regulator of RAD51-mediated homologous recombination at stalled forks. However, the mechanism of RADX acting on DNA and its interactions with RPA and RAD51 are enigmatic. Using singlemolecule imaging of the key proteins in vitro, we reveal that RADX condenses ssDNA filaments, even when the ssDNA is coated with RPA at physiological protein ratios. RADX compacts RPA-coated ssDNA filaments via higher-order assemblies that can capture ssDNA in trans. Furthermore, RADX blocks RPA displacement by RAD51 and prevents RAD51 loading on ssDNA.Our results indicate that RADX is an ssDNA condensation protein that inhibits RAD51 filament formation and may antagonize other ssDNA-binding proteins on RPA-coated ssDNA.

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“…These inhibitors have been shown to be able to suppress HR activity and resensitize cisplatin-resistant cancer cells to cisplatin [15]. UCHL3 is another DUB found to deubiquitinate Rad51 to promote its interaction with BRCA2 for loading to the processed DSB sites [16]. It will be interesting to see if potent and specific inhibitors of UCHL3 can be developed to sensitize the HR-proficient cells to PARP inhibitors or other DNA-damaging agents.…”

Section: Ubiquitinationmentioning

confidence: 99%

Drugging Homologous Recombination: Back to the Future

2018

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If a person is lucky to live long enough, he/she will also have a significantly increased likelihood of developing a cancer. Cancer is caused by accumulation of gene mutations over time. Genomic instability is one of the two key enabling characteristics of cancer cells [1], first enabling initial tumorigenesis and later drug resistance. Genomic stability is critical to maintain cellular homeostasis and cellular identity. However, cellular DNA is constantly under the attack from both endogenous and exogenous threats, creating lesions. These lesions, particularly the DNA double-strand breaks (DSBs) are lethal to the cells. They must be repaired for the cells to survive and proliferate. There are two major pathways to repair the DSBs in mammalian cells. One is error-free homologous recombination repair (HR) and the other one is error-prone nonhomologous end-joining repair [2]. During HR, a homologous sister chromatin is used as a template to repair the breaks. In nonhomologous end-joining, on the other hand, the broken DNA ends are simply ligated together without a need for a homologous template, often leading to some deletions of the original DNA sequences. It is the HR process that maintains the integrity of our cellular genome. Reversing resistanceWhile genomic instability is a hallmark of cancer cells, the genome cannot be too unstable. After passing a certain intolerable threshold, the cancer cells will die from excessive DSBs or genomic changes. This creates a unique opportunity to develop novel cancer therapies by targeting the HR pathway. The viability of targeting HR repair for anti-cancer therapeutics is supported by the recent US FDA approval of three different PARP inhibitors (olaparib, rucaparib and niraparib) for high-grade serous ovarian cancer (HGSOC). These PARP inhibitors are mostly effective in patients with preexisting somatic or germline BRCA mutations through a process called synthetic lethality [3]. BRCA1 and BRCA2 are two distinct breast and ovarian cancer-associated tumor suppressor genes. They are essential for instigating the HR repair pathway in collaboration with the nuclease MRN (Mre11-Rad50-NBS1) complex [2]. Mutations in BRCA genes result in deficient HR repair activity, a process which cancer cells are particularly reliant on due to high level of replication stress [4]. In the case of HGSOC, carriers of BRCA, which account for only approximately 30% of the patients are more sensitive to existing drugs including platinum salts and PARP inhibitors present significantly better prognosis than noncarriers. Even these BRCA carriers eventually develop resistance to the PARP inhibitors or platinum drugs. Although the mechanisms underlying the resistance are incompletely understood, restoration of HR function is a major cause [5,6]. Therefore, pharmacological strategies to drug the HR pathway can potentially expand the utility of existing PARP inhibitors and platinum drugs by priming the non-BRCA carriers and reversing the resistance mechanisms seen in the clinic with PARP inhibitors and pla...

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A phosphorylation–deubiquitination cascade regulates the BRCA2–RAD51 axis in homologous recombination

Cited by 75 publications

References 97 publications

Depletion of DNA damage binding protein 2 sensitizes triple‐negative breast cancer cells to poly ADP‐ribose polymerase inhibition by destabilizing Rad51

Depletion of DNA damage binding protein 2 sensitizes triple‐negative breast cancer cells to poly ADP‐ribose polymerase inhibition by destabilizing Rad51

RADX condenses single-stranded DNA to antagonize RAD51 loading

Drugging Homologous Recombination: Back to the Future

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