GO:0140863 histone H2AK127 ubiquitin ligase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0140863 describes the enzymatic activity that transfers a single ubiquitin molecule onto histone H2A at lysine 127 (H2AK127), a molecular function that directly modifies chromatin structure.
• This activity is part of the broader family of histone ubiquitin ligases, which include RNF168, UHRF1, and SMARCA3, each targeting distinct histone residues to coordinate DNA repair, heterochromatin maintenance, and gene regulation.
• H2A ubiquitination at K127 has been linked to the DNA damage response, where RNF168-mediated ubiquitination of H2A/H2A.X at the damage site is required for the recruitment of downstream repair factors such as 53BP1 and BRCA1.
• The HDAC6-RNF168 axis regulates H2A/H2A.X ubiquitination, and disruption of this axis impairs double-strand break repair, highlighting the therapeutic potential of targeting this activity in cancer.
• UHRF1 ubiquitin ligase activity supports the maintenance of low-density CpG methylation, and its crosstalk with H3K18ub and H3K9me3 reinforces heterochromatin states, demonstrating that H2A ubiquitination is integrated with other epigenetic marks.
• CRISPR-based knockout, point-mutation, and knock-in models are essential tools for dissecting the causal role of H2AK127 ubiquitin ligases in chromatin biology and disease.
Description
Histone H2AK127 ubiquitin ligase activity (GO:0140863) is a molecular function defined as the catalysis of ubiquitin transfer to histone H2A at lysine 127. This post-translational modification is a critical epigenetic mark that influences chromatin compaction, DNA repair, and transcriptional regulation. Unlike ubiquitination of non-histone substrates, histone ubiquitination directly alters the nucleosome surface, creating docking sites for effector proteins that read the ubiquitin code. The study of this activity has gained prominence because dysregulation of histone ubiquitin ligases is increasingly implicated in cancer, genomic instability, and developmental disorders. Researchers investigating chromatin dynamics, DNA damage responses, and epigenetic inheritance require a precise understanding of GO:0140863 to design experiments that distinguish it from other histone ubiquitination events. This article synthesizes the current knowledge on the enzymes, mechanisms, and disease relevance of H2AK127 ubiquitin ligase activity, with a focus on how CRISPR-based models can accelerate discovery.
histone H2AK127 ubiquitin ligase activity At A Glance
| GO ID | GO:0140863 |
|---|---|
| GO term | histone H2AK127 ubiquitin ligase activity |
| Ontology | molecular_function |
| Synonym | histone H2A-K127 ubiquitin ligase activity; histone ubiquitin ligase activity (H2A-K127 specific) |
| Major function | Catalysis of ubiquitin transfer to histone H2A at lysine 127 |
| Substrate | Histone H2A (K127 residue) |
| Cofactors | E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzyme, ATP |
| Associated enzymes | RNF168, UHRF1, SMARCA3 (related histone ubiquitin ligases) |
| Biological context | DNA damage response, heterochromatin maintenance, epigenetic regulation |
What Is GO:0140863?
GO:0140863, histone H2AK127 ubiquitin ligase activity, is the enzymatic catalysis of ubiquitin molecule transfer to histone 2A at the lysine-127 residue. This activity is a type of histone ubiquitin ligase activity that specifically targets H2A at K127, distinguishing it from ligases that modify other histone residues such as H3K23 or H3K14. The reaction involves the covalent attachment of ubiquitin's C-terminal glycine to the epsilon-amino group of H2AK127, a process that requires ATP and the sequential action of E1, E2, and E3 enzymes. The resulting H2AK127ub mark serves as a signaling platform for chromatin-associated factors involved in DNA repair and heterochromatin maintenance.
Why Is histone H2AK127 ubiquitin ligase activity Important in Cell Biology?
GO:0140863 is important because histone H2A ubiquitination at K127 is a key epigenetic modification that regulates chromatin accessibility and genome stability. This activity is essential for the DNA damage response, where RNF168-mediated ubiquitination of H2A/H2A.X at double-strand breaks recruits repair factors such as 53BP1 and BRCA1. Dysregulation of this activity is associated with cancer, as altered histone ubiquitination can lead to genomic instability and aberrant gene expression. Furthermore, crosstalk between H2A ubiquitination and other histone marks, such as H3K9me3 and H3K18ub, underscores its role in heterochromatin inheritance and epigenetic memory. Understanding GO:0140863 therefore provides insights into fundamental chromatin biology and offers potential therapeutic targets for diseases characterized by epigenetic dysfunction.
• Regulates DNA double-strand break repair by recruiting 53BP1 and BRCA1 to damage sites.
• Maintains heterochromatin integrity through crosstalk with H3K9me3 and H3K18ub.
• Supports the maintenance of low-density CpG methylation via UHRF1 activity.
• Implicated in cancer development when dysregulated, particularly in tumors with defective DNA repair.
• Plays a role in epigenetic inheritance and chromatin compartmentalization.
• Serves as a potential biomarker for DNA repair proficiency in cancer therapy.
• Enables the study of ubiquitin signaling in chromatin using CRISPR knockout models.
• Facilitates the development of small-molecule inhibitors targeting histone ubiquitin ligases.
• Contributes to the understanding of histone code crosstalk and combinatorial modifications.
• Provides a mechanistic link between metabolism and chromatin regulation via UHRF1.
Core Biology of histone H2AK127 ubiquitin ligase activity
Substrate Recognition and Ubiquitin Transfer
In simple terms: The enzyme finds histone H2A and attaches a ubiquitin tag to a specific spot called K127.
The catalytic mechanism of GO:0140863 begins with the recognition of histone H2A within the nucleosome by a specific E3 ubiquitin ligase. The E3 enzyme binds to the nucleosome surface, positioning the H2A K127 residue near the active site. In an ATP-dependent reaction, ubiquitin is first activated by an E1 enzyme and transferred to an E2 conjugating enzyme, which then collaborates with the E3 ligase to attach ubiquitin to H2AK127. This modification creates a docking site for reader proteins containing ubiquitin-binding domains, such as those involved in DNA repair.
Role in DNA Damage Response
In simple terms: When DNA breaks, this ubiquitin tag helps call in repair proteins.
Following DNA double-strand breaks, the ubiquitin ligase RNF168 is recruited to damage sites, where it catalyzes H2A/H2A.X ubiquitination. This activity is regulated by the HDAC6-RNF168 axis, which ensures proper ubiquitination for the recruitment of 53BP1 and BRCA1. Ubiquitin-induced RNF168 condensation further promotes DNA repair by amplifying the ubiquitination signal. Loss of H2AK127 ubiquitination impairs the assembly of repair complexes, leading to genomic instability.
Crosstalk with Other Histone Modifications
In simple terms: This ubiquitin mark talks to other chemical tags on histones to control gene activity.
H2AK127 ubiquitination does not occur in isolation; it crosstalks with other histone modifications such as H3K9me3 and H3K18ub. UHRF1-mediated ubiquitination supports the maintenance of low-density CpG methylation and promotes H3K18ub, which in turn reinforces H3K9me3 and heterochromatin states. Similarly, a conserved H3K14ub-driven H3K9me3 pathway contributes to chromatin compartmentalization, indicating that H2A ubiquitination is part of a broader histone ubiquitin network. These crosstalks ensure stable epigenetic inheritance and proper chromatin organization.
Heterochromatin Inheritance and Stress Response
In simple terms: Under stress, this modification helps pass on heterochromatin patterns to new cells.
Stress conditions can control heterochromatin inheritance via histone H3 ubiquitylation, and similar mechanisms may apply to H2A ubiquitination. The activity of H2AK127 ubiquitin ligases is integrated with stress-responsive pathways that modulate chromatin states, ensuring that heterochromatin domains are faithfully propagated during cell division. This regulation is critical for maintaining genome stability and preventing aberrant gene activation.
Key Genes Involved in GO:0140863 histone H2AK127 ubiquitin ligase activity
The following genes encode proteins that either catalyze H2AK127 ubiquitination or regulate this activity through associated complexes.
| Gene | Major Role | Research Relevance |
|---|---|---|
| RNF168 | E3 ubiquitin ligase that catalyzes H2A/H2A.X ubiquitination at DNA damage sites | Key mediator of DNA repair; target for cancer therapy |
| UHRF1 | Multidomain protein with ubiquitin ligase activity; supports CpG methylation maintenance | Links DNA methylation and histone ubiquitination |
| SMARCA3 | Histone H3K23 ubiquitin ligase; related to SWI/SNF chromatin remodeling | Model for studying histone ubiquitin ligase specificity |
| HDAC6 | Deacetylase that regulates RNF168 activity and H2A ubiquitination | Modulates DNA repair efficiency |
| 53BP1 | Reader of H2AK127ub; recruits repair factors to double-strand breaks | Downstream effector of H2A ubiquitination |
| BRCA1 | Tumor suppressor recruited by H2A ubiquitination for homologous recombination | Predictive marker for PARP inhibitor response |
| H2AX | Histone variant that is ubiquitinated at K127 in response to DNA damage | Marker of DNA damage and repair |
| H3K9me3 | Repressive histone mark that crosstalks with H2A ubiquitination | Heterochromatin maintenance |
| H3K18ub | Histone modification that reinforces H3K9me3 and heterochromatin | Epigenetic crosstalk |
| H3K14ub | Histone mark involved in chromatin compartmentalization | Conserved pathway for heterochromatin |
| PKM2 | Metabolic enzyme regulated by March2; linked to aortic aneurysm | Indirect link to ubiquitin signaling |
| March2 | E3 ubiquitin ligase that regulates PKM2 polymerization | Model for ubiquitin ligase function |
| SUV39H1 | Histone methyltransferase that deposits H3K9me3 | Cooperates with UHRF1 in heterochromatin |
| SUV39H2 | Histone methyltransferase that deposits H3K9me3 | Cooperates with UHRF1 in heterochromatin |
| RNF8 | E3 ubiquitin ligase that initiates ubiquitin signaling at DNA damage sites | Upstream of RNF168 |
| E1 ubiquitin-activating enzyme | Activates ubiquitin in an ATP-dependent manner | Essential for all ubiquitination reactions |
| E2 ubiquitin-conjugating enzyme | Carries activated ubiquitin to the E3 ligase | Determines ubiquitin chain linkage |
How Is histone H2AK127 ubiquitin ligase activity Regulated?
The activity of H2AK127 ubiquitin ligases is tightly regulated at multiple levels. RNF168 is recruited to DNA damage sites through its ubiquitin-binding domains and is further activated by phosphorylation and ubiquitin-induced condensation. The HDAC6-RNF168 axis modulates H2A/H2A.X ubiquitination, and inhibition of HDAC6 impairs this process. UHRF1 activity is regulated by its interaction with chromatin and its ability to bind hemimethylated DNA, which coordinates H2A ubiquitination with DNA methylation maintenance. Additionally, stress conditions can influence heterochromatin inheritance via histone H3 ubiquitylation, suggesting that cellular stress pathways may also regulate H2A ubiquitination. These regulatory mechanisms ensure that H2AK127 ubiquitination occurs at the right time and place to maintain genome stability.
histone H2AK127 ubiquitin ligase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| RNF168 | Cancer, genomic instability, impaired DNA repair | RNF168 knockout cell lines; point mutations in catalytic domain |
| UHRF1 | Cancer, epigenetic disorders, CpG methylation defects | UHRF1 knockout and knock-in models; overexpression |
| SMARCA3 | Cancer, chromatin remodeling defects | SMARCA3 knockout and point mutation models |
| HDAC6 | Cancer, DNA repair deficiency | HDAC6 knockout and inhibitor studies |
| March2 | Aortic aneurysm/dissection, metabolic regulation | March2 knockout and overexpression models |
Cancer and Genomic Instability
Dysregulation of H2AK127 ubiquitin ligase activity is strongly associated with cancer. RNF168-mediated H2A ubiquitination is critical for DNA double-strand break repair, and loss of this activity leads to genomic instability and increased sensitivity to DNA-damaging agents. Mutations in RNF168 or its regulators can impair the recruitment of 53BP1 and BRCA1, contributing to tumorigenesis. Furthermore, SMARCA3, a related histone ubiquitin ligase, regulates H3K9me3 in cancer, and its dysfunction may promote aberrant gene expression. Targeting the HDAC6-RNF168 axis is being explored as a therapeutic strategy to sensitize cancer cells to radiotherapy and chemotherapy.
Epigenetic Disorders and Heterochromatin Diseases
Alterations in H2A ubiquitination can disrupt heterochromatin integrity, leading to diseases characterized by epigenetic dysfunction. UHRF1-mediated ubiquitination supports CpG methylation maintenance, and its dysregulation is linked to developmental disorders and cancer. The crosstalk between H3K18ub and H3K9me3 reinforces heterochromatin states, and disruption of this crosstalk can cause aberrant gene activation. Additionally, stress-induced changes in heterochromatin inheritance via histone H3 ubiquitylation may contribute to aging and degenerative diseases.
Cardiovascular and Metabolic Implications
Emerging evidence links ubiquitin ligases to cardiovascular disease. March2, an E3 ubiquitin ligase, alleviates aortic aneurysm/dissection by regulating PKM2 polymerization, indicating that ubiquitin signaling pathways can influence vascular biology. Although direct evidence for H2AK127 ubiquitination in cardiovascular disease is limited, the broader role of histone ubiquitination in gene regulation suggests potential involvement.
From histone H2AK127 ubiquitin ligase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does loss of H2AK127 ubiquitin ligase impair DNA repair? | Knockout of RNF168 or UHRF1 in cell lines, followed by DNA damage assays |
| What is the catalytic mechanism of H2AK127 ubiquitination? | Point mutations in the catalytic cysteine of RNF168 or UHRF1 |
| How does H2AK127 ubiquitination crosstalk with H3K9me3? | Knock-in of H2A K127R mutant to prevent ubiquitination |
| Can H2AK127 ubiquitination be visualized in live cells? | Tagged knock-in of H2A with fluorescent protein and ubiquitin-binding domain |
| Does overexpression of UHRF1 alter CpG methylation? | Overexpression of wild-type and mutant UHRF1 in cell lines |
| What genes are regulated by H2AK127 ubiquitination? | CRISPR library screening with H2A K127R knock-in cells |
How to Study the histone H2AK127 ubiquitin ligase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| CRISPR knockout screen | Gene dependencies and synthetic lethality | Identify regulators of H2AK127 ubiquitination |
| Mass spectrometry | Ubiquitination site mapping and quantification | Profile H2AK127ub changes |
| Live-cell imaging | Real-time recruitment of repair factors | Visualize H2AK127ub at DNA damage sites |
| In vitro ubiquitination assay | Enzymatic activity of E3 ligases | Validate H2AK127-specific ligases |
| Chromatin immunoprecipitation (ChIP) | Genome-wide localization of H2AK127ub | Map ubiquitination across chromatin |
| Hi-C | Chromatin conformation and compartmentalization | Assess heterochromatin integrity |
| RNA-seq | Transcriptional changes upon loss of H2AK127ub | Identify downstream pathways |
| Proximity ligation assay | Protein-protein interactions at damage sites | Detect H2AK127ub-dependent recruitment |
CRISPR-Based Genetic Screens
CRISPR knockout and activation screens can identify genes that regulate or depend on H2AK127 ubiquitin ligase activity. For example, a genome-wide knockout screen in cells expressing an H2A K127R mutant could reveal synthetic lethal interactions with DNA repair pathways. Such screens are powerful for uncovering novel regulators of histone ubiquitination and potential drug targets.
Proteomics and Ubiquitin Remnant Profiling
Mass spectrometry-based proteomics can map ubiquitination sites on histones, including H2AK127. Using ubiquitin remnant motifs, researchers can quantify changes in H2AK127 ubiquitination upon genetic or pharmacological perturbations. This approach provides a global view of crosstalk between histone modifications.
Imaging and Chromatin Conformation Assays
Live-cell imaging with fluorescently tagged H2A and ubiquitin-binding domains allows real-time visualization of H2AK127 ubiquitination at DNA damage sites. Chromatin conformation capture (Hi-C) can assess the impact of H2AK127 ubiquitination on higher-order chromatin structure and heterochromatin domains.
Biochemical Ubiquitination Assays
In vitro ubiquitination assays using recombinant E1, E2, E3 enzymes, and nucleosomes can directly measure H2AK127 ubiquitin ligase activity. These assays are essential for validating candidate ligases and testing small-molecule inhibitors.
How CRISPR Can Be Used to Study GO:0140863 histone H2AK127 ubiquitin ligase activity
Knockout
CRISPR knockout of genes encoding H2AK127 ubiquitin ligases, such as RNF168 or UHRF1, is used to study loss-of-function phenotypes. Knockout cell lines exhibit defective DNA repair, altered heterochromatin, and changes in gene expression. These models are valuable for identifying pathways that compensate for the loss of H2AK127 ubiquitination.
Point Mutation
Point mutations in the catalytic domain of H2AK127 ubiquitin ligases (e.g., RNF168 Cys-to-Ala) can abolish enzymatic activity without affecting protein stability. Such models help distinguish catalytic activity from scaffolding functions. Additionally, point mutation of the substrate H2A at K127 (K127R) prevents ubiquitination and is used to study the consequences of losing this specific mark.
Knock-in
Knock-in of tagged H2A (e.g., H2A-FLAG or H2A-GFP) allows for affinity purification and imaging of ubiquitinated histones. Knock-in of H2A K127R mutant provides a powerful tool to specifically abrogate H2AK127 ubiquitination while preserving other histone functions. These models are essential for dissecting the precise role of H2AK127ub in chromatin.
Overexpression
Overexpression of wild-type or mutant H2AK127 ubiquitin ligases can reveal gain-of-function phenotypes and dominant-negative effects. For example, overexpressing a catalytically dead UHRF1 mutant can compete with endogenous UHRF1 for chromatin binding, thereby inhibiting H2AK127 ubiquitination. Overexpression models are also useful for testing small-molecule inhibitors.
How EDITGENE Supports histone H2AK127 ubiquitin ligase activity Research
Researchers studying histone H2AK127 ubiquitin ligase activity-related genes often need to determine whether a candidate gene is causally involved in chromatin regulation, DNA repair, or disease. EDITGENE provides a comprehensive suite of CRISPR-based services to accelerate this discovery process, from knockout to precise point mutations and library screening.
Contact EDITGENE today to design your custom CRISPR model for histone H2AK127 ubiquitin ligase activity research.
Frequently Asked Questions About histone H2AK127 ubiquitin ligase activity
What is histone H2AK127 ubiquitin ligase activity?
It is the enzymatic activity that attaches a ubiquitin molecule to histone H2A at lysine 127, a modification involved in DNA repair and chromatin regulation.
What genes are involved in histone H2AK127 ubiquitin ligase activity?
Key genes include RNF168, UHRF1, and SMARCA3, which encode E3 ubiquitin ligases or associated proteins.
What is the GO ID for histone H2AK127 ubiquitin ligase activity?
The Gene Ontology ID is GO:0140863.
How does H2AK127 ubiquitination affect DNA repair?
It recruits repair factors such as 53BP1 and BRCA1 to DNA double-strand breaks, facilitating their repair.
Which diseases are linked to H2AK127 ubiquitin ligase dysfunction?
Cancer, genomic instability, and epigenetic disorders are associated with dysregulation of this activity.
What experimental models are used to study H2AK127 ubiquitination?
CRISPR knockout, point mutation, knock-in, and overexpression cell models are commonly used.
How can I measure H2AK127 ubiquitin ligase activity?
In vitro ubiquitination assays, mass spectrometry, and ChIP are standard methods.
Is H2AK127 ubiquitination reversible?
Yes, it is reversed by deubiquitinating enzymes, though specific DUBs for H2AK127 are still being characterized.
What is the role of UHRF1 in H2AK127 ubiquitination?
UHRF1 has ubiquitin ligase activity that supports CpG methylation maintenance and crosstalks with H3K9me3.
Can CRISPR be used to study H2AK127 ubiquitination?
Yes, CRISPR knockout of ligases or knock-in of H2A K127R mutant is widely used to dissect its function.
Conclusion
Histone H2AK127 ubiquitin ligase activity (GO:0140863) is a fundamental molecular function that regulates chromatin structure, DNA repair, and epigenetic inheritance. The enzymes responsible, including RNF168 and UHRF1, are critical for genome stability and are implicated in cancer and other diseases. Understanding the mechanisms and regulation of this activity requires advanced experimental models, and CRISPR-based approaches offer unprecedented precision. As research uncovers the crosstalk between H2AK127 ubiquitination and other histone modifications, new therapeutic opportunities are likely to emerge.
References
- 1. Tiedemann RL et al.. 2024. UHRF1 ubiquitin ligase activity supports the maintenance of low-density CpG methylation.. Nucleic Acids Res 52(22):13733-13756 PMID: 39607687
- 2. Li YE et al.. 2025. March2 Alleviates Aortic Aneurysm/Dissection by Regulating PKM2 Polymerization.. Circ Res 136(8):e73-e93 PMID: 40079144
- 3. Akano I et al.. 2025. The SWI/SNF-related protein SMARCA3 is a histone H3K23 ubiquitin ligase that regulates H3K9me3 in cancer.. Mol Cell 85(15):2885-2899.e8 PMID: 40680746
- 4. Huang Y et al.. 2025. A conserved H3K14ub-driven H3K9me3 for chromatin compartmentalization.. Nature 647(8090):786-797 PMID: 41094145
- 5. Feng LL et al.. 2024. Ubiquitin-induced RNF168 condensation promotes DNA double-strand break repair.. Proc Natl Acad Sci U S A 121(28):e2322972121 PMID: 38968116
- 6. Qiu L et al.. 2023. The HDAC6-RNF168 axis regulates H2A/H2A.X ubiquitination to enable double-strand break repair.. Nucleic Acids Res 51(17):9166-9182 PMID: 37503842
- 7. Bhatt B et al.. 2026. Stress controls heterochromatin inheritance via histone H3 ubiquitylation.. Nature 650(8102):768-778 PMID: 41501458
- 8. Liu Y et al.. 2025. DNA hypomethylation promotes UHRF1-and SUV39H1/H2-dependent crosstalk between H3K18ub and H3K9me3 to reinforce heterochromatin states.. Mol Cell 85(2):394-412.e12 PMID: 39631394
