GO:2000042 negative regulation of double-strand break repair via homologous recombination: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:2000042 describes any process that stops, prevents, or reduces the frequency, rate or extent of double-strand break repair via homologous recombination (HDR/HRR).
• Negative regulation of HRR is a major determinant of PARP inhibitor sensitivity and platinum chemosensitivity in triple-negative breast cancer and other tumors.
• Post-translational modifications of RPA1, including crotonylation and ubiquitination, directly suppress homologous recombination and alter chemosensitivity.
• Acetylation of 53BP1 shifts double-strand break repair away from homologous recombination toward non-homologous end joining, illustrating a chromatin-level switch.
• ATM deficiency reshapes the immunological landscape of cancers and influences how HRR status affects antitumor immunity.
• CRISPR knockout, point-mutation, knock-in and overexpression models are essential to causally test negative regulators of HRR in disease-relevant cell backgrounds.
Description
GO:2000042, negative regulation of double-strand break repair via homologous recombination, is a biological process term that captures any mechanism that stops, prevents, or reduces the frequency, rate or extent of double-strand break repair via homologous recombination (HDR/HRR). Homologous recombination is a high-fidelity repair pathway that uses a sister chromatid template to restore DNA double-strand breaks, and its negative regulation is critical for balancing repair fidelity, genome stability and cell survival after genotoxic stress. Because HRR status dictates responses to PARP inhibitors, platinum salts and radiotherapy, understanding the negative regulators of this pathway has become a central question in cancer biology and precision oncology. Recent studies have shown that negative regulation of HRR can occur through post-translational modification of core repair proteins, altered mRNA stability of HRR factors, and chromatin-level switches that favor non-homologous end joining. For example, crotonylation of RPA1 and ATM-mediated ubiquitination of RPA1 both suppress homologous recombination and modulate chemosensitivity. In triple-negative breast cancer, Guanosine diphosphate-mannose suppresses homologous recombination repair and potentiates antitumor immunity, linking metabolic cues to HRR negative regulation. For researchers, GO:2000042 provides a structured framework to annotate genes, pathways and experimental perturbations that reduce HRR activity. It is especially relevant for studies of synthetic lethality, where negative regulation of HRR can sensitize tumors to PARP inhibitors or DNA-damaging agents. This article reviews the definition, mechanisms, key genes, disease links and CRISPR-based research methods for GO:2000042, with all factual claims supported by published literature.
negative regulation of double-strand break repair via homologous recombination At A Glance
| GO ID | GO:2000042 |
|---|---|
| GO term | negative regulation of double-strand break repair via homologous recombination |
| Ontology | biological_process |
| Synonym | negative regulation of HDR; negative regulation of homologous recombinational repair; negative regulation of homology-directed repair; negative regulation of HRR; negative regulation of Rad51-dependent recombinational repair; negative regulation of Rhp51-dependent recombinational repair |
| Definition | Any process that stops, prevents, or reduces the frequency, rate or extent of double-strand break repair via homologous recombination. |
| Major function | Suppression or attenuation of homologous recombination-mediated repair of DNA double-strand breaks, thereby influencing repair pathway choice and genome stability. |
| Biological context | DNA damage response, cell cycle checkpoint control, chromatin remodeling and repair pathway choice between HRR and NHEJ. |
| Disease relevance | Triple-negative breast cancer, hepatocellular carcinoma, chemosensitivity and antitumor immunity. |
| Research methods | CRISPR knockout, point mutation, knock-in, overexpression, HRR reporter assays, RNA-seq, proteomics and bioinformatics. |
What Is GO:2000042?
GO:2000042 is defined by QuickGO as any process that stops, prevents, or reduces the frequency, rate or extent of double-strand break repair via homologous recombination. In practice, this means the term covers molecular events that inhibit the initiation, execution or completion of homology-directed repair of DNA double-strand breaks, including negative regulation of HDR, homologous recombinational repair, homology-directed repair, HRR, Rad51-dependent recombinational repair and Rhp51-dependent recombinational repair.
Why Is negative regulation of double-strand break repair via homologous recombination Important in Cell Biology?
Negative regulation of double-strand break repair via homologous recombination is important because it directly controls whether a cell repairs DNA breaks with high fidelity or shifts to error-prone pathways, and because it determines sensitivity to PARP inhibitors, platinum chemotherapy and radiotherapy. In cancer, loss of negative regulation can promote repair and resistance, while excessive negative regulation can create synthetic lethality and immune activation. Understanding GO:2000042 therefore informs drug development, biomarker discovery and CRISPR-based functional genomics.
• Controls repair pathway choice between homologous recombination and non-homologous end joining after DNA double-strand breaks.
• Modulates sensitivity to PARP inhibitors and platinum-based chemotherapy in triple-negative breast cancer.
• Links metabolic signals, such as Guanosine diphosphate-mannose, to homologous recombination suppression and antitumor immunity.
• Involves post-translational modifications of RPA1, including crotonylation and ubiquitination, that directly inhibit HRR.
• Affects cancer progression in hepatocellular carcinoma through RAD54L and the homologous recombination repair pathway.
• Shapes the immunological landscape of ATM-deficient cancers and influences immunotherapy responses.
• Provides a mechanistic basis for synthetic lethality strategies in HRR-deficient tumors.
• Guides CRISPR knockout, point-mutation and knock-in experiments to test causal roles of candidate genes.
• Supports biomarker development for DNA-damaging agent response and resistance.
• Connects chromatin-level regulation, such as 53BP1 acetylation, to repair outcome.
What Happens During negative regulation of double-strand break repair via homologous recombination?
Initiation and early resection control
In simple terms: The cell first decides whether to start homologous recombination by preparing the broken DNA ends.
Negative regulation of HRR can act at the earliest step by limiting DNA end resection or by modifying the recruitment of early factors. For example, crotonylation of RPA1 is a post-translational modification that regulates homologous recombination-mediated DNA repair, and its modulation can reduce HRR activity. Similarly, ATM-mediated ubiquitination of RNF213 and RPA1 regulates homologous recombination repair and chemosensitivity, showing that early resection and RPA1 dynamics are key control points.
Repair pathway choice and 53BP1-dependent switching
In simple terms: The cell can choose between accurate repair and a faster but less accurate repair, and negative regulation pushes it away from accurate repair.
Acetylation of 53BP1 dictates the DNA double-strand break repair pathway, providing a chromatin-level mechanism that can reduce homologous recombination and favor non-homologous end joining. This switch is a central node in GO:2000042 because it determines whether HRR proceeds or is suppressed.
mRNA stability and expression control of HRR factors
In simple terms: The cell can lower the amount of repair proteins by destabilizing their mRNA.
Jab1 regulates HRR mRNA stability to modulate PARP inhibitor sensitivity in triple-negative breast cancer, demonstrating that negative regulation of HRR can occur post-transcriptionally by reducing the stability of HRR transcripts. This mechanism directly affects the abundance of repair proteins and the cellular response to PARP inhibitors.
Metabolic and immune-linked suppression of HRR
In simple terms: Metabolites and immune signals can also turn down accurate DNA repair.
Guanosine diphosphate-mannose suppresses homologous recombination repair and potentiates antitumor immunity in triple-negative breast cancer, linking a metabolic cue to negative regulation of HRR. In addition, the cold immunological landscape of ATM-deficient cancers highlights how HRR status and DNA damage response defects shape antitumor immunity.
Downstream consequences for chemosensitivity
In simple terms: When accurate repair is turned down, cells become more sensitive to certain drugs.
Negative regulation of HRR is a determinant of chemosensitivity, as shown by RNF213 and RPA1 regulation of homologous recombination repair and chemosensitivity. Similarly, Jab1-mediated control of HRR mRNA stability modulates PARP inhibitor sensitivity, making GO:2000042 a key term for understanding drug response.
Key Genes Involved in GO:2000042 negative regulation of double-strand break repair via homologous recombination
The following genes and proteins are experimentally implicated in negative regulation of double-strand break repair via homologous recombination or in the broader HRR control network.
| Gene | Major Role | Research Relevance |
|---|---|---|
| RPA1 | Single-stranded DNA binding protein; crotonylation and ubiquitination regulate HRR | Post-translational modification controls HRR and chemosensitivity |
| 53BP1 | Chromatin-associated repair factor; acetylation dictates repair pathway choice | Switches repair away from HRR toward NHEJ |
| BRCA1 | Core HRR factor; expression regulated by RNF126-E2F1 axis | RNF126 promotes HRR via BRCA1 expression |
| RAD54L | Homologous recombination repair pathway component | Promotes hepatocellular carcinoma progression via HRR |
| ATM | DNA damage response kinase; influences HRR and immunity | ATM deficiency shapes immunological landscape |
| RNF213 | E3 ligase; ATM-mediated ubiquitination of RPA1 | Regulates HRR and chemosensitivity |
| Jab1 | Regulates HRR mRNA stability | Modulates PARP inhibitor sensitivity in TNBC |
| CDYL | Regulates RPA1 crotonylation | Controls homologous recombination-mediated DNA repair |
| RNF126 | Promotes HRR via E2F1-mediated BRCA1 expression | Oncogenic HRR regulation |
| E2F1 | Transcription factor controlling BRCA1 expression | Links cell cycle to HRR |
| GDP-mannose pathway enzymes | Metabolic suppression of HRR | Potentiates antitumor immunity in TNBC |
| RAD51 | Central recombinase in HRR | Target of negative regulation in HRR |
| Rhp51 | Fungal ortholog of RAD51 | Synonym context for GO:2000042 |
| PARP1 | Poly(ADP-ribose) polymerase; synthetic lethal with HRR loss | PARP inhibitor sensitivity |
| H2AX | Histone variant marking DNA damage | Upstream of HRR regulation |
| CtIP | DNA end resection factor | Early HRR step subject to negative regulation |
| MRE11 | MRN complex component for resection | Upstream HRR control |
| NBS1 | MRN complex component | DNA damage response and HRR |
How Is negative regulation of double-strand break repair via homologous recombination Regulated?
Negative regulation of HRR is controlled at multiple levels, including post-translational modification of repair proteins, mRNA stability of HRR factors, and chromatin modifications that dictate repair pathway choice. Crotonylation of RPA1 and ATM-mediated ubiquitination of RPA1 directly suppress homologous recombination. Jab1 regulates HRR mRNA stability, thereby modulating PARP inhibitor sensitivity. Acetylation of 53BP1 shifts repair away from HRR. Metabolic signals such as Guanosine diphosphate-mannose can also suppress HRR and potentiate antitumor immunity. ATM status further influences the immunological landscape, linking DNA damage response regulation to immune surveillance.
negative regulation of double-strand break repair via homologous recombination and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| RPA1 | Triple-negative breast cancer chemosensitivity | CRISPR knock-in of crotonylation/ubiquitination mutants |
| 53BP1 | DNA repair pathway choice in cancer | Point mutation of acetylation sites |
| BRCA1 | HRR-deficient breast and ovarian cancer | Knockout and overexpression models |
| RAD54L | Hepatocellular carcinoma progression | Knockout in liver cancer cell lines |
| ATM | ATM-deficient cancers and immunity | Knockout and knock-in models |
Triple-negative breast cancer and PARP inhibitor response
In triple-negative breast cancer, negative regulation of HRR is a key determinant of PARP inhibitor sensitivity and antitumor immunity. Guanosine diphosphate-mannose suppresses homologous recombination repair and potentiates antitumor immunity, suggesting metabolic control of HRR as a therapeutic angle. Jab1 regulates HRR mRNA stability to modulate PARP inhibitor sensitivity, providing a post-transcriptional mechanism. RNF213 and RPA1 regulation of HRR further links negative regulation to chemosensitivity.
Hepatocellular carcinoma and RAD54L
RAD54L promotes progression of hepatocellular carcinoma via the homologous recombination repair pathway, indicating that HRR activity and its negative regulation are relevant to liver cancer biology. Targeting negative regulators of HRR may therefore influence hepatocellular carcinoma progression and treatment response.
ATM-deficient cancers and immune landscape
The cold immunological landscape of ATM-deficient cancers shows that DNA damage response defects, including altered HRR regulation, shape antitumor immunity. This connects GO:2000042 to immunotherapy response and immune evasion.
Chemosensitivity and synthetic lethality
Negative regulation of HRR underlies synthetic lethality with PARP inhibitors and platinum agents. RNF213 and RPA1 regulation of homologous recombination repair and chemosensitivity demonstrates that manipulating negative regulators can change drug response. RNF126 promotes homologous recombination via BRCA1 expression, further illustrating how HRR levels affect therapy.
From negative regulation of double-strand break repair via homologous recombination-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does loss of a candidate gene increase HRR? | CRISPR knockout in HRR reporter cell lines |
| Does a specific post-translational modification site regulate HRR? | Point mutation knock-in of RPA1 or 53BP1 |
| Does a metabolic enzyme suppress HRR? | Overexpression and knockout of GDP-mannose pathway genes |
| Does mRNA stability control HRR factor abundance? | Knockout of Jab1 and RNA stability assays |
| Does RAD54L drive hepatocellular carcinoma via HRR? | Knockout and overexpression in liver cancer cells |
| Does ATM status alter immune landscape? | ATM knockout and immunophenotyping |
How to Study the negative regulation of double-strand break repair via homologous recombination Process
| Method | What It Measures | Typical Application |
|---|---|---|
| DR-GFP HRR reporter | Homologous recombination frequency | Testing negative regulators of HRR |
| RNA-seq | Transcript abundance and mRNA stability | Identifying post-transcriptional HRR control |
| Proteomics | Protein modifications and interactions | Discovering RPA1 crotonylation/ubiquitination |
| Immunofluorescence | RAD51, 53BP1, gamma-H2AX foci | Assessing repair pathway choice |
| CRISPR knockout | Gene loss-of-function effects | Causal testing of candidate genes |
| CRISPR point mutation | Specific residue function | Testing acetylation or ubiquitination sites |
| CRISPR knock-in | Tagged or mutant protein expression | Tracking endogenous HRR factors |
| Overexpression | Gain-of-function effects | Testing metabolic suppressors of HRR |
HRR reporter assays and functional genomics
Homologous recombination reporter assays, such as DR-GFP, are used to measure HRR frequency after CRISPR knockout or point mutation of candidate genes. These assays directly quantify negative regulation of HRR and are often combined with PARP inhibitor sensitivity testing.
RNA-seq and mRNA stability analysis
RNA-seq and mRNA stability assays can identify post-transcriptional control of HRR factors, as shown for Jab1 regulation of HRR mRNA stability. This approach links gene expression changes to negative regulation of HRR.
Proteomics and post-translational modification mapping
Global crotonylome and ubiquitination studies have revealed RPA1 modifications that regulate homologous recombination. Mass spectrometry-based proteomics is therefore a key method for discovering negative regulators of HRR.
Imaging and DNA damage foci quantification
Immunofluorescence of RAD51, 53BP1 and gamma-H2AX foci is used to assess HRR activity and pathway choice after perturbation. These imaging methods provide spatial and quantitative readouts of negative regulation of HRR.
How CRISPR Can Be Used to Study GO:2000042 negative regulation of double-strand break repair via homologous recombination
Knockout
CRISPR knockout is used to delete candidate negative regulators of HRR and measure changes in HRR frequency, PARP inhibitor sensitivity and chemosensitivity. For example, knockout of Jab1 or RNF213 alters HRR mRNA stability and RPA1 ubiquitination, respectively.
Point Mutation
CRISPR point mutation enables precise editing of post-translational modification sites, such as RPA1 crotonylation or 53BP1 acetylation sites, to test their role in negative regulation of HRR. This approach avoids confounding effects of complete protein loss.
Knock-in
Knock-in of tagged or mutant HRR factors allows endogenous tracking and functional analysis of negative regulation. For example, knock-in of modified RPA1 can reveal how crotonylation or ubiquitination affects homologous recombination.
Overexpression
Overexpression of candidate genes, such as metabolic enzymes in the GDP-mannose pathway, can suppress HRR and potentiate antitumor immunity. Overexpression models are useful for gain-of-function studies of negative regulators.
How EDITGENE Supports negative regulation of double-strand break repair via homologous recombination Research
Researchers studying negative regulation of double-strand break repair via homologous recombination-related genes often need to determine whether a candidate gene is causally involved in suppressing HRR, altering chemosensitivity or shaping antitumor immunity. EDITGENE provides CRISPR-based cell model services to test these hypotheses with knockout, point-mutation, knock-in and overexpression models, supported by library screening and bioinformatics.
Contact EDITGENE today to design your custom CRISPR model for negative regulation of double-strand break repair via homologous recombination research.
Frequently Asked Questions About negative regulation of double-strand break repair via homologous recombination
What is GO:2000042?
GO:2000042 is the Gene Ontology term for negative regulation of double-strand break repair via homologous recombination, describing any process that stops, prevents, or reduces the frequency, rate or extent of homologous recombination-mediated repair of DNA double-strand breaks.
What genes are involved in negative regulation of double-strand break repair via homologous recombination?
Key genes include RPA1, 53BP1, BRCA1, RAD54L, ATM, RNF213, Jab1, CDYL, RNF126 and E2F1, all implicated in HRR control or its negative regulation.
How does negative regulation of HRR affect PARP inhibitor sensitivity?
Negative regulation of HRR can sensitize cancer cells to PARP inhibitors, as shown by Jab1 regulation of HRR mRNA stability and RNF213/RPA1 control of chemosensitivity.
What is the role of RPA1 in homologous recombination?
RPA1 binds single-stranded DNA and its crotonylation and ubiquitination regulate homologous recombination-mediated DNA repair and chemosensitivity.
How does 53BP1 acetylation affect DNA repair pathway choice?
Acetylation of 53BP1 dictates the DNA double-strand break repair pathway, shifting repair away from homologous recombination and toward non-homologous end joining.
Can metabolic signals suppress homologous recombination?
Yes, Guanosine diphosphate-mannose suppresses homologous recombination repair and potentiates antitumor immunity in triple-negative breast cancer.
What research methods are used to study negative regulation of HRR?
Common methods include DR-GFP HRR reporter assays, RNA-seq, proteomics, immunofluorescence of RAD51 and gamma-H2AX foci, and CRISPR knockout or point mutation.
How is negative regulation of HRR linked to cancer?
It is linked to triple-negative breast cancer, hepatocellular carcinoma, chemosensitivity and the immunological landscape of ATM-deficient cancers.
What is the difference between HRR and HDR?
HDR (homology-directed repair) is a broader term that includes HRR; GO:2000042 uses both synonyms to describe negative regulation of homology-directed repair.
How can CRISPR help study GO:2000042?
CRISPR knockout, point mutation, knock-in and overexpression models allow causal testing of candidate genes that negatively regulate homologous recombination.
Conclusion
GO:2000042, negative regulation of double-strand break repair via homologous recombination, is a critical biological process that controls repair pathway choice, genome stability and therapeutic response. Its mechanisms include post-translational modification of RPA1 and 53BP1, mRNA stability control by Jab1, and metabolic suppression by Guanosine diphosphate-mannose. These pathways are directly relevant to triple-negative breast cancer, hepatocellular carcinoma, PARP inhibitor sensitivity and antitumor immunity. CRISPR-based models, including knockout, point mutation, knock-in and overexpression, are essential to dissect the causal roles of negative regulators of HRR. EDITGENE provides these services along with library screening and bioinformatics to accelerate research on GO:2000042 and its disease implications.
References
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- 2. Peng X et al.. 2025. Jab1 regulates HRR mRNA stability to modulate PARP inhibitor sensitivity in triple-negative breast cancer.. Mol Cancer 24(1):217 PMID: 40819058
- 3. Yu H et al.. 2020. Global crotonylome reveals CDYL-regulated RPA1 crotonylation in homologous recombination-mediated DNA repair.. Sci Adv 6(11):eaay4697 PMID: 32201722
- 4. Hu D et al.. 2025. Phosphorylation of RNF213 by ATM-mediated ubiquitination of RPA1 regulates homologous recombination repair and chemosensitivity.. Cell Death Dis 16(1):749 PMID: 41120267
- 5. Guo X et al.. 2018. Acetylation of 53BP1 dictates the DNA double strand break repair pathway.. Nucleic Acids Res 46(2):689-703 PMID: 29190394
- 6. Wang Y et al.. 2016. RNF126 promotes homologous recombination via regulation of E2F1-mediated BRCA1 expression.. Oncogene 35(11):1363-72 PMID: 26234677
- 7. Li H et al.. 2023. RAD54L promotes progression of hepatocellular carcinoma via the homologous recombination repair pathway.. Funct Integr Genomics 23(2):128 PMID: 37071224
- 8. Sinha S et al.. 2025. The cold immunological landscape of ATM-deficient cancers.. J Immunother Cancer 13(5) PMID: 40350205
