GO:1904333 positive regulation of error-prone translesion synthesis: DNA Damage Tolerance Pathway, Genes, Functions and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:1904333 describes the biological process that increases the frequency, rate, or extent of error-prone translesion synthesis (TLS), a DNA damage tolerance mechanism.
• Error-prone TLS is carried out by specialized DNA polymerases such as Pol ι, Pol η, and Pol κ that bypass DNA lesions and can introduce mutations.
• Positive regulation of TLS is critical for cancer cell survival after chemotherapy or radiation, contributing to therapy resistance.
• Key regulators include RAD18, which monoubiquitinates PCNA to recruit TLS polymerases, and JNK/c-Jun signaling that upregulates Pol ι.
• Dysregulated TLS is implicated in colorectal, esophageal, bladder, and triple-negative breast cancers.
• CRISPR knockout, point mutation, knock-in, and overexpression models enable precise dissection of TLS regulatory networks.
Description
GO:1904333, positive regulation of error-prone translesion synthesis, is a biological process that activates or increases the frequency, rate, or extent of error-prone translesion synthesis (TLS). TLS is a DNA damage tolerance pathway in which specialized low-fidelity DNA polymerases replicate past DNA lesions that block the high-fidelity replicative polymerases, thereby allowing cell survival at the cost of potential mutations. This process is tightly regulated to balance genome maintenance and mutagenesis, and its dysregulation is linked to cancer progression and therapy resistance. Researchers study GO:1904333 to understand how cells tolerate DNA damage, how mutations arise, and how to target these pathways in diseases such as cancer.
positive regulation of error-prone translesion synthesis At A Glance
| GO ID | GO:1904333 |
|---|---|
| GO term | positive regulation of error-prone translesion synthesis |
| Ontology | biological_process |
| Synonym | activation of error-prone translesion synthesis; positive regulation of mutagenic PRR; upregulation of error-prone postreplication DNA repair |
| Major function | Upregulates error-prone translesion synthesis, allowing DNA lesion bypass by specialized polymerases |
| Key regulators | RAD18, JNK/c-Jun, PCNA monoubiquitination |
| Associated polymerases | Pol ι, Pol η, Pol κ |
| Disease relevance | Cancer progression, chemoresistance, metastasis |
What Is GO:1904333?
According to the Gene Ontology, GO:1904333 (positive regulation of error-prone translesion synthesis) encompasses any process that activates or increases the frequency, rate, or extent of error-prone translesion synthesis. This includes molecular events that promote the recruitment, activation, or function of error-prone DNA polymerases at sites of DNA damage, leading to lesion bypass and potentially mutagenic outcomes.
Why Is positive regulation of error-prone translesion synthesis Important in Cell Biology?
Positive regulation of error-prone translesion synthesis is crucial because it determines whether cells survive DNA damage at the expense of mutations. This process is a double-edged sword: it protects against replication fork collapse and cell death, but also drives mutagenesis and cancer development. In cancer, upregulation of TLS often contributes to resistance to genotoxic therapies, making it a target for therapeutic intervention.
• Enables cell survival after DNA damage by bypassing lesions that block replication.
• Contributes to mutagenesis and genomic instability, hallmarks of cancer.
• Mediates resistance to chemotherapy and radiotherapy in multiple cancers.
• Regulates cancer stemness and progression in triple-negative breast cancer.
• Promotes invasion and metastasis in esophageal squamous cell carcinoma.
• Involved in hypermutagenesis in bladder cancer.
• Potential biomarker for predicting therapy response.
• Target for sensitizing tumors to DNA-damaging agents.
• Plays a role in immune microenvironment crosstalk via TGF-β signaling.
• Provides a mechanism for adaptation to replication stress.
What Happens During positive regulation of error-prone translesion synthesis?
DNA Damage Recognition and PCNA Monoubiquitination
In simple terms: When DNA is damaged, a ring protein called PCNA gets tagged with a small protein called ubiquitin, which acts like a landing pad for repair polymerases.
The initiation of error-prone translesion synthesis requires the recognition of stalled replication forks and the monoubiquitination of proliferating cell nuclear antigen (PCNA). RAD18, an E3 ubiquitin ligase, together with RAD6, catalyzes this modification, which is a key step for recruiting TLS polymerases to the damage site. This process is positively regulated by various signaling pathways, including JNK/c-Jun, which can upregulate RAD18 or downstream polymerases.
Recruitment of Error-Prone DNA Polymerases
In simple terms: Specialized DNA polymerases that can copy past damage are brought to the stalled fork.
Monoubiquitinated PCNA recruits Y-family DNA polymerases such as Pol η, Pol ι, Pol κ, and REV1. These polymerases contain ubiquitin-binding domains that interact with ubiquitinated PCNA. Positive regulation of TLS involves increased expression or activation of these polymerases. For example, DNA polymerase iota (Pol ι) is overexpressed in bladder cancer and promotes hypermutagenesis, and its expression is regulated by JNK/c-Jun. Similarly, Pol η is a substrate for calpain, which affects its retention at UV-induced replication foci.
Lesion Bypass and Mutagenesis
In simple terms: The specialized polymerases copy DNA past the damage, often inserting wrong bases and causing mutations.
Once recruited, TLS polymerases catalyze the insertion of nucleotides opposite the lesion and extend the primer, allowing replication to continue. Because these polymerases lack proofreading activity, they can introduce mutations. Positive regulation of this process increases the frequency of lesion bypass and mutagenesis. In colorectal cancer cells, cetuximab treatment alters the network of translesion polymerases, suggesting that therapy can modulate TLS activity.
Regulation by Signaling Pathways
In simple terms: Cellular signals like stress kinases can boost the TLS process.
Positive regulation of error-prone TLS is controlled by various signaling cascades. The JNK/c-Jun pathway upregulates Pol ι, contributing to hypermutagenesis in bladder cancer. In esophageal squamous cell cancer, RAD18 promotes migration and invasion via the JNK-MMPs pathway. Additionally, a positive feedback loop involving RAD18-YAP-TGF-β between triple-negative breast cancer and macrophages regulates cancer stemness and progression. These examples illustrate how extracellular signals and microenvironmental interactions can enhance TLS.
Key Genes Involved in GO:1904333 positive regulation of error-prone translesion synthesis
The following genes and proteins are central to the positive regulation of error-prone translesion synthesis, based on published literature.
| Gene | Major Role | Research Relevance |
|---|---|---|
| RAD18 | E3 ubiquitin ligase that monoubiquitinates PCNA to recruit TLS polymerases | Promotes migration and invasion in esophageal cancer; regulates cancer stemness in TNBC |
| POLI | Error-prone DNA polymerase iota; bypasses lesions with low fidelity | Overexpressed in bladder and esophageal cancers; regulated by JNK/c-Jun |
| POLH | DNA polymerase eta; involved in UV lesion bypass | Substrate for calpain; retention at replication foci |
| PCNA | Sliding clamp; monoubiquitination serves as platform for TLS polymerase recruitment | Central node in TLS regulation; target for modulating TLS |
| REV1 | Y-family polymerase; scaffold for TLS polymerases | Part of TLS network; potential target in colorectal cancer |
| POLK | DNA polymerase kappa; error-prone lesion bypass | Analyzed in colorectal cancer following cetuximab treatment |
| JNK | Stress-activated kinase; upregulates Pol ι and RAD18 | Mediates signaling to TLS in cancer |
| c-Jun | Transcription factor downstream of JNK; regulates POLI expression | Contributes to hypermutagenesis in bladder cancer |
| YAP | Transcriptional co-activator; involved in RAD18-YAP-TGF-β feedback loop | Regulates cancer stemness in TNBC |
| TGF-β | Cytokine; part of feedback loop with RAD18 and YAP | Modulates tumor microenvironment in TNBC |
| MMPs | Matrix metalloproteinases; downstream of RAD18-JNK pathway | Promote invasion and metastasis in esophageal cancer |
| Calpain | Protease; cleaves Pol η | Regulates Pol η retention at UV-induced foci |
| RAD6 | E2 ubiquitin-conjugating enzyme; partners with RAD18 | Essential for PCNA monoubiquitination |
| Ubiquitin | Small protein modifier; monoubiquitinates PCNA | Key post-translational modification in TLS |
| EGFR | Receptor tyrosine kinase; cetuximab target | Treatment alters TLS polymerase network in colorectal cancer |
| Cetuximab | Monoclonal antibody against EGFR | Used to study TLS network changes in colorectal cancer |
How Is positive regulation of error-prone translesion synthesis Regulated?
Positive regulation of error-prone translesion synthesis is controlled at multiple levels. Post-translational modification of PCNA by RAD18-mediated monoubiquitination is a critical switch. Transcriptional upregulation of TLS polymerases, such as POLI by JNK/c-Jun, increases TLS capacity. Signaling pathways including JNK, YAP, and TGF-β form feedback loops that amplify TLS under stress conditions. Additionally, proteolytic processing by calpain can modulate Pol η availability. These regulatory mechanisms ensure that TLS is activated when needed but can also be hijacked in cancer.
positive regulation of error-prone translesion synthesis and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| RAD18 | Esophageal squamous cell cancer; triple-negative breast cancer | Knockout in cancer cell lines; xenograft models |
| POLI | Bladder cancer; esophageal cancer | Overexpression and knockout in bladder/esophageal cell lines |
| POLH | UV-induced skin damage; potential role in cancer | Point mutation to prevent calpain cleavage; UV irradiation models |
| PCNA | General cancer therapy resistance | Knock-in of ubiquitination-deficient PCNA mutant |
| REV1 | Colorectal cancer | Knockout in colorectal cancer cells; cetuximab treatment |
Cancer Progression and Metastasis
Dysregulated positive regulation of error-prone TLS contributes to cancer progression and metastasis. In esophageal squamous cell carcinoma, RAD18 promotes migration and invasion via the JNK-MMPs pathway, and Pol ι promotes invasion and metastasis. In triple-negative breast cancer, a RAD18-YAP-TGF-β feedback loop between cancer cells and macrophages regulates cancer stemness and progression. These findings highlight TLS as a driver of aggressive cancer phenotypes.
Therapy Resistance
Upregulation of error-prone TLS can confer resistance to DNA-damaging therapies. In colorectal cancer cells, cetuximab treatment alters the network of translesion polymerases, suggesting that TLS activity may influence treatment response. Targeting positive regulators of TLS could sensitize tumors to chemotherapy and radiotherapy.
Mutagenesis and Genomic Instability
Overexpression of error-prone polymerases such as Pol ι leads to hypermutagenesis in bladder cancer. This increased mutation load can drive tumor heterogeneity and evolution. Understanding the regulation of TLS is therefore crucial for comprehending how cancers acquire mutations that fuel progression and drug resistance.
From positive regulation of error-prone translesion synthesis-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does RAD18 promote metastasis in esophageal cancer? | RAD18 knockout in esophageal cancer cell lines; invasion assays |
| Does Pol ι overexpression increase mutagenesis? | POLI overexpression in bladder cancer cells; mutation frequency assays |
| How does calpain cleavage affect Pol η function? | Point mutation of calpain cleavage site in POLH; UV foci imaging |
| Does PCNA monoubiquitination drive therapy resistance? | Knock-in of PCNA K164R mutant in colorectal cancer cells; cetuximab treatment |
| What is the role of RAD18-YAP-TGF-β loop in TNBC? | Knockout of RAD18 or YAP in TNBC cells; co-culture with macrophages |
| Can targeting TLS sensitize tumors to radiation? | Overexpression of dominant-negative REV1; radiation survival assays |
How to Study the positive regulation of error-prone translesion synthesis Process
| Method | What It Measures | Typical Application |
|---|---|---|
| CRISPR knockout | Loss-of-function effects on TLS | Identify essential TLS regulators |
| Point mutation knock-in | Specific residue function | Block PCNA monoubiquitination |
| Overexpression | Gain-of-function effects | Induce hypermutagenesis |
| Proteomics | Protein interactions and modifications | Map TLS network changes |
| Imaging | Subcellular localization | Track Pol η foci after UV |
| Mutation frequency assay | Error-prone TLS activity | Quantify mutagenesis |
| Invasion/migration assay | Metastatic potential | Assess RAD18 or Pol ι effects |
| Co-culture systems | Tumor-microenvironment interactions | Study RAD18-YAP-TGF-β loop |
CRISPR-Cas9 Knockout Screens
Genome-wide CRISPR knockout screens can identify genes that positively regulate error-prone TLS. For example, knocking out RAD18 or POLI can reduce TLS activity and sensitize cells to DNA-damaging agents. These screens are powerful for discovering novel regulators and potential therapeutic targets.
Point Mutation and Knock-in Models
Introducing point mutations in key residues, such as PCNA K164R, can block monoubiquitination and prevent TLS polymerase recruitment. Knock-in of such mutations allows precise dissection of regulatory mechanisms. Similarly, mutating calpain cleavage sites in Pol η can reveal its regulation.
Overexpression and Reporter Assays
Overexpressing TLS polymerases like Pol ι can increase mutagenesis and promote cancer phenotypes. Reporter assays measuring lesion bypass or mutation frequency can quantify TLS activity. These approaches help establish causality between positive regulators and TLS outcomes.
Proteomics and Network Analysis
Mass spectrometry-based proteomics can identify interaction partners and post-translational modifications of TLS components. Network analysis of translesion polymerases in colorectal cancer cells following cetuximab treatment revealed dynamic changes in TLS-related proteins. Such methods provide systems-level insights into TLS regulation.
How CRISPR Can Be Used to Study GO:1904333 positive regulation of error-prone translesion synthesis
Knockout
CRISPR knockout of positive regulators such as RAD18 or POLI can abolish error-prone TLS, leading to reduced mutagenesis and increased sensitivity to DNA-damaging agents. Knockout models are essential for validating the role of specific genes in TLS and cancer phenotypes.
Point Mutation
Point mutations can be introduced to disrupt specific functions, such as the ubiquitination site on PCNA (K164R) or calpain cleavage sites on Pol η. These models allow precise interrogation of regulatory mechanisms without completely eliminating the protein.
Knock-in
Knock-in of tagged versions of TLS proteins (e.g., GFP-POLH) enables live-cell imaging and tracking of protein dynamics at replication foci. Knock-in of disease-associated mutations can also model cancer predisposition.
Overexpression
CRISPR activation (CRISPRa) or lentiviral overexpression can drive high levels of TLS polymerases like Pol ι, mimicking the overexpression observed in cancers and allowing study of hypermutagenesis and metastasis.
How EDITGENE Supports positive regulation of error-prone translesion synthesis Research
Researchers studying positive regulation of error-prone translesion synthesis-related genes often need to determine whether a candidate gene is causally involved in DNA damage tolerance, mutagenesis, or cancer progression. EDITGENE provides a comprehensive suite of CRISPR services to enable such investigations with precision and reproducibility.
Contact EDITGENE today to design your custom CRISPR model for positive regulation of error-prone translesion synthesis research.
Frequently Asked Questions About positive regulation of error-prone translesion synthesis
What is GO:1904333?
GO:1904333 is the Gene Ontology term for positive regulation of error-prone translesion synthesis, a biological process that increases the frequency, rate, or extent of error-prone TLS.
What genes are involved in positive regulation of error-prone translesion synthesis?
Key genes include RAD18, POLI, POLH, PCNA, REV1, POLK, JNK, c-Jun, YAP, and TGF-β.
How does RAD18 regulate error-prone translesion synthesis?
RAD18 monoubiquitinates PCNA, which recruits error-prone DNA polymerases to sites of DNA damage, thereby promoting TLS.
What is the role of Pol ι in cancer?
Pol ι is an error-prone polymerase that is overexpressed in bladder and esophageal cancers, where it promotes hypermutagenesis, invasion, and metastasis.
How is error-prone translesion synthesis linked to chemotherapy resistance?
Upregulation of TLS allows cancer cells to bypass DNA damage induced by chemotherapy, leading to therapy resistance.
What experimental models are used to study positive regulation of error-prone TLS?
CRISPR knockout, point mutation knock-in, overexpression, and proteomics are commonly used to study TLS regulation.
Which diseases are associated with dysregulated error-prone TLS?
Dysregulated TLS is associated with colorectal, esophageal, bladder, and triple-negative breast cancers.
How does JNK signaling regulate TLS?
JNK/c-Jun signaling upregulates Pol ι expression, contributing to hypermutagenesis in bladder cancer.
What is the RAD18-YAP-TGF-β feedback loop?
It is a signaling loop between triple-negative breast cancer cells and macrophages that regulates cancer stemness and progression via RAD18.
Can CRISPR screens identify new regulators of error-prone TLS?
Yes, genome-wide CRISPR screens can uncover novel positive regulators of TLS and potential therapeutic targets.
Conclusion
Positive regulation of error-prone translesion synthesis (GO:1904333) is a critical biological process that enables cells to survive DNA damage at the cost of mutations. Its dysregulation is intimately linked to cancer progression, metastasis, and therapy resistance. Understanding the molecular players and regulatory mechanisms of TLS is essential for developing new strategies to target this pathway in cancer and other diseases. EDITGENE offers comprehensive CRISPR solutions to accelerate research on this important process.
References
- 1. Das A et al.. 2024. Analysis of translesion polymerases in colorectal cancer cells following cetuximab treatment: A network perspective.. Cancer Med 13(1):e6945 PMID: 39102671
- 2. Zou S et al.. 2018. RAD18 promotes the migration and invasion of esophageal squamous cell cancer via the JNK-MMPs pathway.. Cancer Lett 417:65-74 PMID: 29306013
- 3. Yan X et al.. 2022. A positive feedback loop: RAD18-YAP-TGF-β between triple-negative breast cancer and macrophages regulates cancer stemness and progression.. Cell Death Discov 8(1):196 PMID: 35413945
- 4. Zou S et al.. 2016. DNA polymerase iota (Pol ι) promotes invasion and metastasis of esophageal squamous cell carcinoma.. Oncotarget 7(22):32274-85 PMID: 27057634
- 5. Nettersheim JA et al.. 2021. DNA polymerase η is a substrate for calpain: a possible mechanism for pol η retention in UV-induced replication foci.. J Cell Sci 134(13) PMID: 34080634
- 6. Yuan F et al.. 2013. Overexpressed DNA polymerase iota regulated by JNK/c-Jun contributes to hypermutagenesis in bladder cancer.. PLoS One 8(7):e69317 PMID: 23922701
