GO:0031390 Ctf18 RFC-like complex: Components, Assembly and Research Methods, Genes, Functions and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0031390 describes the Ctf18 RFC-like complex, a heptameric clamp loader related to replication factor C that loads PCNA onto DNA and is essential for chromosome cohesion.
• The complex contains the subunits Ctf18p, Rfc2p, Rfc3p, Rfc4p, Rfc5p, Dcc1p, and Ctf8p in Saccharomyces cerevisiae.
• Ctf18-RFC functions in telomere positioning and maintenance, and its inactivation causes telomere defects in yeast.
• Ctf18-RFC is distinct from the Elg1-RFC complex, which acts as a PCNA unloader and guardian of genome stability.
• Dysregulation of clamp loaders like Ctf18-RFC is linked to genome instability, a hallmark of cancer and premature aging.
• CRISPR-based knockout, point mutation, knock-in, and overexpression models enable precise dissection of Ctf18-RFC subunit functions in human cells.
Description
The Ctf18 RFC-like complex (GO:0031390) is a conserved heptameric clamp loader that shares homology with replication factor C (RFC) but performs specialized roles in chromosome cohesion and telomere maintenance. Unlike the canonical RFC, which loads proliferating cell nuclear antigen (PCNA) during DNA replication, the Ctf18-RFC complex is critical for sister chromatid cohesion and proper telomere positioning. In Saccharomyces cerevisiae, the complex comprises Ctf18p, Rfc2p, Rfc3p, Rfc4p, Rfc5p, Dcc1p, and Ctf8p, and its inactivation leads to chromosome mis-segregation and telomere clustering defects. This complex is part of a family of alternative RFC-like clamp loaders, including Elg1-RFC and Rad17-RFC, that maintain genome stability through specialized PCNA loading and unloading activities. Research into Ctf18-RFC has revealed its importance in DNA damage responses, checkpoint control, and the prevention of aneuploidy. Understanding its molecular mechanism is essential for deciphering how cells preserve genomic integrity and how its dysfunction contributes to diseases such as cancer.
Ctf18 RFC-like complex At A Glance
| GO ID | GO:0031390 |
|---|---|
| GO term | Ctf18 RFC-like complex |
| Ontology | cellular_component |
| Synonym | Ctf18-RFC, Ctf18-RLC, RFC (Ctf18) |
| Major function | Loads PCNA onto DNA and promotes chromosome cohesion and telomere maintenance |
| Subunits (S. cerevisiae) | Ctf18p, Rfc2p, Rfc3p, Rfc4p, Rfc5p, Dcc1p, Ctf8p |
| Related complex | Elg1-RFC (PCNA unloader) and Rad17-RFC (checkpoint clamp loader) |
| Disease relevance | Genome instability, cancer predisposition, and telomere-related disorders |
What Is GO:0031390?
The Ctf18 RFC-like complex is a heptameric protein complex that is structurally related to replication factor C (RFC) and functions as a clamp loader for proliferating cell nuclear antigen (PCNA). It plays a vital role in chromosome cohesion and telomere maintenance, and in Saccharomyces cerevisiae its subunits are Ctf18p, Rfc2p, Rfc3p, Rfc4p, Rfc5p, Dcc1p, and Ctf8p.
Why Is Ctf18 RFC-like complex Important in Cell Biology?
The Ctf18 RFC-like complex is essential for maintaining genome stability through its roles in sister chromatid cohesion and telomere maintenance. Its dysfunction leads to chromosome mis-segregation, aneuploidy, and increased sensitivity to DNA-damaging agents, which are hallmarks of cancer and premature aging. Studying this complex provides insights into how cells coordinate DNA replication with cohesion establishment and how alternative clamp loaders contribute to genome integrity.
• Ensures proper sister chromatid cohesion during S phase, preventing chromosome mis-segregation.
• Positions telomeres at the nuclear periphery, influencing telomere maintenance and silencing.
• Acts as a specialized PCNA loader that is distinct from the canonical RFC.
• Its inactivation sensitizes cells to DNA replication stress and damage.
• Mutations or dysregulation are linked to genome instability and cancer.
• Provides a model for studying alternative clamp loaders and their specialized functions.
• Interacts genetically with Elg1-RFC and Rad17-RFC pathways.
• Serves as a target for understanding cohesion-related developmental disorders.
Ctf18 RFC-like complex
PCNA Loading and Chromosome Cohesion
In simple terms: The complex loads a ring-shaped protein onto DNA to hold sister chromatids together.
The Ctf18 RFC-like complex loads PCNA onto DNA during S phase, which is required for establishing sister chromatid cohesion. This loading activity is distinct from that of the canonical RFC and is thought to couple DNA replication with cohesion establishment. In Saccharomyces cerevisiae, loss of Ctf18-RFC leads to defective cohesion and chromosome mis-segregation.
Telomere Positioning and Maintenance
In simple terms: The complex helps anchor telomeres to the nuclear envelope and maintain their structure.
Ctf18-RFC positions yeast telomeres at the nuclear periphery but does not specify their replication timing. In Schizosaccharomyces pombe, Ctf18-RFC is involved in telomere maintenance, and its deletion causes telomere shortening and dysfunction. These roles are independent of its cohesion function and highlight the complex's multifunctional nature.
Distinction from Elg1-RFC and Rad17-RFC
In simple terms: Different clamp loaders do different jobs: Ctf18 loads PCNA for cohesion, Elg1 unloads PCNA, and Rad17 loads a checkpoint clamp.
The Ctf18-RFC complex is one of several alternative RFC-like complexes. Elg1-RFC acts as a PCNA unloader and guardian of genome stability, while Rad17-RFC loads the 9-1-1 checkpoint clamp. In fission yeast, inactivation of Ctf18-RFC and Elg1-RFC has contrasting effects in the absence of fully functional RFC. These distinctions are critical for understanding how cells coordinate different genome maintenance pathways.
Molecular Mechanism and Regulation
In simple terms: The complex uses ATP to open the PCNA ring and place it on DNA, and its activity is regulated by cell cycle and checkpoint signals.
Like canonical RFC, Ctf18-RFC uses ATP binding and hydrolysis to open the PCNA ring and load it onto primer-template junctions. The complex interacts with DNA polymerase processivity factors and is regulated by cell cycle kinases and checkpoint pathways. In Xenopus egg extracts, the Atad5 RFC-like complex is the major PCNA unloader, indicating that loading and unloading are tightly balanced. The Ctf18-RFC complex may also be regulated by its subunit composition and post-translational modifications.
Key Genes Involved in GO:0031390 Ctf18 RFC-like complex
The following genes encode subunits of the Ctf18 RFC-like complex and related clamp loaders, based on studies in Saccharomyces cerevisiae, Schizosaccharomyces pombe, and other model organisms.
| Gene | Major Role | Research Relevance |
|---|---|---|
| CTF18 | ATPase subunit of Ctf18-RFC; essential for cohesion and telomere positioning | Knockout causes chromosome mis-segregation and telomere clustering defects |
| RFC2 | Small subunit of RFC and Ctf18-RFC; involved in clamp loading | Required for PCNA loading and DNA replication |
| RFC3 | Small subunit of RFC and Ctf18-RFC | Mutations affect clamp loading and genome stability |
| RFC4 | Small subunit of RFC and Ctf18-RFC | Essential for PCNA loading and cohesion |
| RFC5 | Small subunit of RFC and Ctf18-RFC | Interacts with Ctf18p and Dcc1p for complex assembly |
| DCC1 | Accessory subunit of Ctf18-RFC; required for cohesion | Deletion leads to sister chromatid cohesion defects |
| CTF8 | Accessory subunit of Ctf18-RFC; required for cohesion | Deletion causes chromosome instability |
| ELG1 | Subunit of Elg1-RFC, a PCNA unloader | Plays a role in sister chromatid cohesion and genome stability |
| RAD17 | Subunit of Rad17-RFC, loads 9-1-1 checkpoint clamp | Involved in DNA damage checkpoint and telomere maintenance |
| PCNA | Processivity factor loaded by RFC and Ctf18-RFC | Essential for DNA replication and repair |
| ATAD5 | Subunit of Atad5 RFC-like complex, major PCNA unloader | Regulates PCNA unloading in Xenopus and human cells |
| RFC1 | Large subunit of canonical RFC | Required for PCNA loading during replication |
| CTF4 | Accessory factor for cohesion establishment | Interacts with Ctf18-RFC to promote cohesion |
| POL30 | Yeast PCNA homolog | Loaded by Ctf18-RFC onto DNA |
| SMC1 | Cohesin subunit | Functional partner of Ctf18-RFC in cohesion |
| SMC3 | Cohesin subunit | Functional partner of Ctf18-RFC in cohesion |
| ECO1 | Acetyltransferase for cohesin | Works with Ctf18-RFC to establish cohesion |
How Is Ctf18 RFC-like complex Regulated?
The Ctf18 RFC-like complex is regulated at multiple levels. Its activity is cell cycle-dependent, peaking during S phase when cohesion is established. Post-translational modifications of subunits, such as phosphorylation by cyclin-dependent kinases and checkpoint kinases, modulate its interactions and localization. In fission yeast, the complex genetically interacts with the DNA damage checkpoint and with Elg1-RFC, suggesting coordinated regulation of PCNA loading and unloading. The abundance of accessory subunits like Dcc1 and Ctf8 may also control complex assembly and function.
Ctf18 RFC-like complex and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| CTF18 | Chromosome instability, cancer predisposition | Knockout human cell lines (e.g., HCT116) |
| RFC2 | Genome instability, replication stress | Point mutation knock-in in yeast or human cells |
| DCC1 | Cohesinopathy-like phenotypes | Overexpression and knockout in zebrafish or mouse models |
| ELG1 | Cancer, genome instability | Knockout mouse models and CRISPR screens |
| ATAD5 | PCNA unloading defects, cancer | Knockout in Xenopus egg extracts and human cells |
Cancer and Genome Instability
Dysregulation of clamp loaders, including Ctf18-RFC, leads to genome instability, a hallmark of cancer. Loss of Ctf18-RFC function causes chromosome mis-segregation and aneuploidy, which can promote tumorigenesis. In human cells, mutations in RFC subunits are associated with increased cancer susceptibility.
Telomere-Related Disorders
Ctf18-RFC is required for telomere maintenance in yeast, and its dysfunction causes telomere shortening and dysfunction. Telomere attrition is linked to premature aging syndromes and cancer predisposition in humans. Thus, Ctf18-RFC may contribute to telomere-related pathologies.
Cohesinopathies and Developmental Disorders
Sister chromatid cohesion defects caused by Ctf18-RFC inactivation resemble cohesinopathies such as Roberts syndrome. These developmental disorders are characterized by limb abnormalities, craniofacial defects, and intellectual disability. Studying Ctf18-RFC provides insights into the molecular basis of cohesion-related diseases.
From Ctf18 RFC-like complex-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does Ctf18-RFC load PCNA onto DNA during S phase? | In vitro PCNA loading assays with purified components |
| What is the role of Ctf18-RFC in sister chromatid cohesion? | Yeast knockout strains and live-cell imaging of cohesin |
| How does Ctf18-RFC affect telomere positioning? | Yeast telomere FISH and nuclear pore colocalization |
| What are the human disease implications of Ctf18-RFC mutations? | CRISPR knockout human cell lines and organoids |
| How is Ctf18-RFC regulated by the cell cycle? | Synchronized cell cultures and phospho-proteomics |
| Can Ctf18-RFC be targeted for cancer therapy? | CRISPR library screens and drug sensitivity assays |
How to Study the Ctf18 RFC-like complex Process
| Method | What It Measures | Typical Application |
|---|---|---|
| CRISPR knockout | Loss-of-function phenotypes | Cohesion and telomere defects in yeast and human cells |
| Co-immunoprecipitation | Protein-protein interactions | Subunit assembly and partner identification |
| Live-cell imaging | Complex localization and chromosome dynamics | Telomere positioning and cohesion |
| In vitro PCNA loading | ATP-dependent clamp loading | Mechanistic studies of RFC-like complexes |
| RNA-seq | Transcriptional changes upon knockout | Pathway analysis in Ctf18-RFC mutants |
| Proteomics | Post-translational modifications and interactome | Regulation of Ctf18-RFC |
| Yeast genetics | Synthetic lethality and genetic interactions | Pathway crosstalk with Elg1-RFC and Rad17-RFC |
| FISH | Telomere length and localization | Telomere maintenance defects |
Genetic Knockout and Knockdown
CRISPR-Cas9 knockout of CTF18, DCC1, or CTF8 in yeast and human cells is used to assess cohesion, telomere maintenance, and genome stability. RNAi knockdown provides a complementary approach for acute depletion.
Protein-Protein Interaction Studies
Co-immunoprecipitation, yeast two-hybrid, and mass spectrometry can identify interactions between Ctf18-RFC subunits and partners like PCNA and cohesin. These methods help define the complex's assembly and regulation.
Live-Cell Imaging and FISH
Fluorescence microscopy of tagged subunits and telomere FISH are used to monitor complex localization and telomere positioning. Time-lapse imaging of cohesin and chromosomes reveals cohesion defects.
Biochemical Assays for PCNA Loading
In vitro PCNA loading assays with purified RFC-like complexes measure ATP-dependent clamp opening and loading onto DNA. These assays can distinguish Ctf18-RFC from canonical RFC and Elg1-RFC.
How CRISPR Can Be Used to Study GO:0031390 Ctf18 RFC-like complex
Knockout
CRISPR-Cas9 knockout of CTF18, DCC1, or CTF8 in human cell lines (e.g., HCT116, HeLa) can recapitulate cohesion and telomere defects observed in yeast. These models are useful for studying genome instability and aneuploidy.
Point Mutation
Point mutations in the ATPase domain of CTF18 can be introduced via CRISPR to dissect its catalytic activity from structural roles. Such models help distinguish PCNA loading from other functions.
Knock-in
Knock-in of tagged CTF18 (e.g., GFP or HA) allows live-cell imaging and proteomic analysis of the complex. Knock-in of disease-associated mutations can model human pathologies.
Overexpression
Overexpression of Ctf18-RFC subunits or the entire complex can be achieved via CRISPR activation or lentiviral vectors to study dosage effects on cohesion and telomere maintenance. This is useful for identifying gain-of-function phenotypes.
How EDITGENE Supports Ctf18 RFC-like complex Research
Researchers studying Ctf18 RFC-like complex-related genes often need to determine whether a candidate gene is causally involved in cohesion, telomere maintenance, or genome stability. EDITGENE provides a comprehensive suite of CRISPR-based services to generate precisely engineered cell models for such investigations.
Contact EDITGENE today to design your custom CRISPR model for Ctf18 RFC-like complex research.
Frequently Asked Questions About Ctf18 RFC-like complex
What is the Ctf18 RFC-like complex?
The Ctf18 RFC-like complex is a heptameric clamp loader related to replication factor C that loads PCNA onto DNA and is essential for chromosome cohesion and telomere maintenance.
What genes are involved in the Ctf18 RFC-like complex?
In Saccharomyces cerevisiae, the complex comprises CTF18, RFC2, RFC3, RFC4, RFC5, DCC1, and CTF8.
What is the function of GO:0031390?
GO:0031390 describes the Ctf18 RFC-like complex, which loads PCNA onto DNA and plays a vital role in chromosome cohesion and telomere positioning.
How does Ctf18-RFC differ from canonical RFC?
Ctf18-RFC contains the Ctf18, Dcc1, and Ctf8 subunits instead of the large Rfc1 subunit, and it specializes in cohesion and telomere maintenance rather than bulk DNA replication.
What diseases are associated with Ctf18-RFC dysfunction?
Dysfunction of Ctf18-RFC leads to genome instability, aneuploidy, and telomere defects, which are linked to cancer and cohesinopathies.
How can I study Ctf18-RFC using CRISPR?
CRISPR knockout, point mutation, knock-in, and overexpression models can be used to dissect subunit functions in cohesion, telomere maintenance, and PCNA loading.
What is the role of Ctf18-RFC in telomere maintenance?
Ctf18-RFC positions yeast telomeres at the nuclear periphery and is required for telomere length maintenance in fission yeast.
Is Ctf18-RFC a PCNA loader or unloader?
Ctf18-RFC acts as a PCNA loader, while Elg1-RFC and Atad5-RFC are PCNA unloaders.
What model organisms are used to study Ctf18-RFC?
Saccharomyces cerevisiae, Schizosaccharomyces pombe, and Xenopus egg extracts are commonly used to study Ctf18-RFC function.
What are the subunits of Ctf18-RFC?
The subunits are Ctf18p, Rfc2p, Rfc3p, Rfc4p, Rfc5p, Dcc1p, and Ctf8p in Saccharomyces cerevisiae.
Conclusion
The Ctf18 RFC-like complex (GO:0031390) is a specialized clamp loader that is critical for chromosome cohesion and telomere maintenance. Its unique subunit composition and functions distinguish it from canonical RFC and other RFC-like complexes. Dysregulation of Ctf18-RFC leads to genome instability and is implicated in cancer and developmental disorders. Continued research using CRISPR-based models will further elucidate its molecular mechanisms and therapeutic potential.
References
- 1. Kawasoe Y et al.. 2024. The Atad5 RFC-like complex is the major unloader of proliferating cell nuclear antigen in Xenopus egg extracts.. J Biol Chem 300(1):105588 PMID: 38141767
- 2. Hiraga S et al.. 2006. The Ctf18 RFC-like complex positions yeast telomeres but does not specify their replication time.. EMBO J 25(7):1505-14 PMID: 16525505
- 3. Khair L et al.. 2010. Roles of the checkpoint sensor clamp Rad9-Rad1-Hus1 (911)-complex and the clamp loaders Rad17-RFC and Ctf18-RFC in Schizosaccharomyces pombe telomere maintenance.. Cell Cycle 9(11):2237-48 PMID: 20505337
- 4. Kim J et al.. 2005. Contrasting effects of Elg1-RFC and Ctf18-RFC inactivation in the absence of fully functional RFC in fission yeast.. Nucleic Acids Res 33(13):4078-89 PMID: 16040599
- 5. Lee KY et al.. 2020. Eukaryotic clamp loaders and unloaders in the maintenance of genome stability.. Exp Mol Med 52(12):1948-1958 PMID: 33339954
- 6. Aroya SB et al.. 2005. The Elg1 replication factor C-like complex: a novel guardian of genome stability.. DNA Repair (Amst) 4(4):409-17 PMID: 15725622
- 7. Parnas O et al.. 2009. The ELG1 clamp loader plays a role in sister chromatid cohesion.. PLoS One 4(5):e5497 PMID: 19430531
- 8. Kanellis P et al.. 2003. Elg1 forms an alternative PCNA-interacting RFC complex required to maintain genome stability.. Curr Biol 13(18):1583-95 PMID: 13678589
