Affinage

RFC5

Replication factor C subunit 5 · UniProt P40937

Length
340 aa
Mass
38.5 kDa
Annotated
2026-06-10
11 papers in source corpus 10 papers cited in narrative 10 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 5/5 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

RFC5 encodes a small subunit of the replication factor C (RFC) clamp-loader complex, which is essential for processive DNA chain elongation by DNA polymerase delta/epsilon (PMID:7774928, PMID:8692942). Beyond its replication role, Rfc5 acts as a sensor and transducer of replication and DNA-damage checkpoint signals: the rfc5-1 mutation abolishes the S-phase checkpoint and impairs DNA-damage-induced Rad53 phosphorylation and RNR3 induction, defects rescued by overexpression of SPK1/Rad53 or TEL1, placing Rfc5 upstream of Rad53 in the Mec1/Tel1 signaling axis (PMID:8692942, PMID:9315648). Rfc5 physically interacts with the RFC-related checkpoint protein Rad24, an interaction dependent on Rad24's NTP-binding motif and required for checkpoint control across G1, S, and G2/M phases (PMID:9710632, PMID:10913172). RFC box motifs IV–VII of Rfc5 mediate inter-subunit contacts within the complex, and the replication and checkpoint functions are genetically separable (PMID:11129041). In human cancer cells, RFC5 supports homologous-recombination DNA repair and, by enhancing repair of chemotherapy-induced lesions, suppresses cGAS-STING innate-immune signaling to promote immune evasion (PMID:34042508, PMID:41192524); its expression is transcriptionally activated by FoxM1, contributing to chemoresistance (PMID:28185110).

Mechanistic history

Synthesis pass · year-by-year structured walk · 10 steps
  1. 1995 Medium

    Established the human gene product as a bona fide subunit of the multimeric RFC clamp loader required for processive DNA polymerase delta/epsilon activity, anchoring RFC5 to the replication machinery.

    Evidence PCR from somatic hybrid DNAs and FISH mapping of human RFC5 (p36.5)

    PMID:7774928

    Open questions at the time
    • No direct biochemical assay of Rfc5 clamp-loading activity
    • Subunit stoichiometry and contacts not resolved
  2. 1996 High

    Showed RFC has a function beyond replication itself — sensing replication state and transmitting an S-phase checkpoint signal — by separating the replication defect (PCNA-suppressible) from the checkpoint defect (Rad53-suppressible).

    Evidence Temperature-sensitive rfc5-1 mutant analysis and SPK1/POL30 overexpression suppression in S. cerevisiae, with chromosome segregation readout

    PMID:8692942

    Open questions at the time
    • Molecular nature of the replication-state signal sensed by RFC unknown
    • Direct biochemical link to Rad53 not shown
  3. 1997 High

    Placed Rfc5 in the Mec1/Tel1-Rad53 DNA-damage checkpoint axis, demonstrating it is needed to activate Rad53 and downstream transcriptional responses after damage during S phase.

    Evidence Rad53 phosphorylation shift, RNR3 induction, S-phase progression assays, and TEL1/RAD53 suppressor genetics in rfc5-1 mutants

    PMID:9315648

    Open questions at the time
    • Whether Rfc5 directly contacts Mec1/Tel1 not established
    • Mechanism of signal relay to Rad53 unresolved
  4. 1998 High

    Identified a physical partner for the checkpoint function — the RFC-related protein Rad24 — providing a molecular basis for how RFC subunits couple to the damage-response apparatus.

    Evidence Co-immunoprecipitation, co-sedimentation, and RAD24-overexpression suppression of rfc5-1 with Rad53 phosphorylation readout

    PMID:9710632

    Open questions at the time
    • Whether Rad24-Rfc5 forms a distinct alternative clamp-loader not defined here
    • Direct binding interface not mapped
  5. 2000 High

    Demonstrated the Rad24–RFC interaction operates across all cell cycle phases and depends on Rad24's NTP-binding motif, generalizing the checkpoint role beyond S phase.

    Evidence Co-IP with Rad24 K115R/K115E motif mutants and rfc5-1 rad24 double-mutant checkpoint assays in G1, S, G2/M

    PMID:10913172

    Open questions at the time
    • Structural basis of NTP-dependent interaction not resolved
    • Loaded sensor structure unknown
  6. 2000 Medium

    Genetically separated RFC5's replication and checkpoint functions and mapped inter-subunit contact determinants to conserved RFC box motifs IV–VII.

    Evidence RFC5 suppressor screen of rfc1-1, phenotypic readouts (MMS/HU sensitivity, telomere length, mutator), Rad53 phosphorylation, comparative structural inference

    PMID:11129041

    Open questions at the time
    • Structural inference is comparative, not direct
    • Single-lab characterization
  7. 2017 Medium

    Connected RFC5 to chemoresistance in human cancer by identifying transcriptional control through FoxM1.

    Evidence FoxM1 promoter-interaction assay, siRNA knockdown of FoxM1/RFC5, viability/apoptosis assays and thiostrepton inhibition in glioma cells

    PMID:28185110

    Open questions at the time
    • No ChIP confirmation of direct promoter binding
    • Single lab, no reconstitution
  8. 2021 Medium

    Implicated RFC5 in homologous-recombination repair in human cells, extending its DNA-metabolism role beyond replication.

    Evidence AEG-1 knockdown gene-expression array, γ-H2AX foci, colony formation, HR repair activity assay after ionizing radiation in glioma

    PMID:34042508

    Open questions at the time
    • AEG-1→RFC5 link is correlative from array
    • Direct role of RFC5 in HR step not mechanistically dissected
  9. 2025 Medium

    Linked RFC5-mediated DNA repair to tumor immune evasion, showing repair of chemotherapy lesions suppresses cytosolic DNA sensing and dampens anti-tumor immunity.

    Evidence RFC5 manipulation in NPC cells, micronuclei quantification, cGAS-STING assays, in vivo cytokine and T-cell exhaustion marker analysis

    PMID:41192524

    Open questions at the time
    • No direct RFC5 biochemical assay
    • Single study, single lab
    • Causal chain from repair to STING suppression inferred
  10. 2024 Low

    Raised the possibility that RFC5 function is modulated post-transcriptionally through SRSF10-dependent alternative splicing.

    Evidence SRSF10 knockdown/overexpression with gel electrophoresis of RFC5 splice variants and proliferation/cell-cycle assays in colorectal cancer cells

    PMID:39110418

    Open questions at the time
    • Splicing change detected only by gel electrophoresis
    • Functional consequence of the specific exon-2-excluded isoform not validated
    • Single lab

Open questions

Synthesis pass · forward-looking unresolved questions
  • How RFC5 mechanistically transduces a replication/damage signal to Rad53/Mec1 at the molecular level, and whether the human clamp loader retains a distinct checkpoint-sensing role, remains unresolved.
  • No structure of an RFC5-containing checkpoint sensor
  • Direct enzymatic activity of human RFC5 within HR not defined
  • Mechanism linking repair to cGAS-STING suppression not biochemically established

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0060089 molecular transducer activity 3 GO:0140657 ATP-dependent activity 2
Localization
GO:0005634 nucleus 2
Pathway
R-HSA-1640170 Cell Cycle 2 R-HSA-69306 DNA Replication 2 R-HSA-8953897 Cellular responses to stimuli 2 R-HSA-73894 DNA Repair 1
Partners
RAD24RAD53RFC2
Complex memberships
Replication factor C (RFC) complex

Evidence

Reading pass · 10 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
1996 Rfc5 is a small subunit of the replication factor C (RFC) complex in S. cerevisiae and is required for the S-phase checkpoint that couples DNA replication to mitotic entry. The rfc5-1 temperature-sensitive mutation causes cells to enter mitosis with unevenly separated or fragmented chromosomes, and overexpression of SPK1/Rad53 suppresses this defect, placing Rfc5 upstream of Rad53 in the checkpoint pathway. Overexpression of PCNA (POL30) suppresses the replication defect but not the checkpoint defect, indicating RFC has a direct role in sensing replication state and transmitting the checkpoint signal. Genetic epistasis (suppressor screens, temperature-sensitive mutant analysis, overexpression of SPK1/POL30), cell biology (chromosome segregation analysis) Proceedings of the National Academy of Sciences of the United States of America High 8692942
1997 Rfc5 is required for the DNA damage checkpoint: in rfc5-1 mutants, Rad53 phosphorylation in response to DNA damage is reduced during S phase, RNR3 transcription induction is impaired, and S-phase progression is not slowed in response to DNA damage. Overexpression of TEL1 suppresses rfc5-1 defects and restores Rad53 phosphorylation and RNR3 induction, placing Rfc5 in the Mec1/Tel1-Rad53 signaling axis upstream of Rad53 activation. Genetic epistasis (suppressor analysis with TEL1, RAD53 overexpression), phosphorylation assays (Rad53 phosphorylation shift), transcription induction assay (RNR3) Molecular and cellular biology High 9315648
1998 Rad24, a protein structurally related to RFC subunits, physically interacts with RFC subunits Rfc2 and Rfc5 and co-sediments with Rfc5. RAD24 overexpression suppresses rfc5-1 sensitivity to DNA-damaging agents and restores Rad53 phosphorylation, demonstrating a physical and functional interaction between Rad24 and Rfc5 in checkpoint pathways. Co-immunoprecipitation, co-sedimentation, genetic suppressor analysis, Rad53 phosphorylation assay Molecular and cellular biology High 9710632
2000 RFC5 functions in G1-, S-, and G2/M-phase DNA damage checkpoints in cooperation with Rad24. In rfc5-1 rad24-K115R double mutants, G1 and G2/M checkpoint defects appear that are absent in either single mutant. Co-immunoprecipitation showed that the Rad24(K115R) protein (NTP-binding motif mutant) fails to interact with RFC proteins in rfc5-1 mutants, establishing that the Rad24–RFC interaction is essential for checkpoint control across all cell cycle phases. Co-immunoprecipitation, site-directed mutagenesis of NTP-binding motif (K115E/K115R), double-mutant genetic epistasis, cell cycle checkpoint assays Molecular and cellular biology High 10913172
2000 RFC5 suppressor mutations that rescue rfc1-1 cold-sensitive growth map to conserved RFC box motifs IV–VII of Rfc5p, regions predicted to mediate inter-subunit contacts. These RFC5 suppressors do not interfere with Rad53 phosphorylation, unlike the checkpoint-defective rfc5-1 mutation, separating replication and checkpoint functions and suggesting RFC box motifs IV–VII coordinate neighboring-subunit interactions within the RFC complex. Genetic suppressor screen (isolation and characterization of RFC5 suppressor alleles), phenotypic analysis (MMS/HU sensitivity, telomere length, mutator phenotype), Rad53 phosphorylation assay, structural comparison Molecular & general genetics : MGG Medium 11129041
1995 Human RFC5 (p36.5 subunit) is a component of the multimeric human replication factor C complex, which is essential for processive DNA chain elongation by DNA polymerase delta or epsilon. The RFC5 gene was mapped to human chromosome band 12q24.2–q24.3. PCR amplification from somatic hybrid DNAs, fluorescence in situ hybridization (FISH) Genomics Medium 7774928
2017 FoxM1 transcriptionally activates RFC5 expression by directly interacting with the RFC5 promoter. Knockdown of FoxM1 or RFC5 re-sensitizes glioma cells to temozolomide, indicating the FoxM1–RFC5 transcriptional axis mediates TMZ resistance. Promoter interaction assay (FoxM1 binding to RFC5 promoter), siRNA knockdown of FoxM1/RFC5, cell viability/apoptosis assays, pharmacological inhibition (thiostrepton) Cell biology and toxicology Medium 28185110
2021 AEG-1 positively regulates RFC5 expression in glioma cells (identified by gene expression array). AEG-1 knockdown reduces RFC5 levels and impairs homologous recombination DNA repair activity induced by ionizing radiation, demonstrating that RFC5 is required for HR repair in glioma cells. Affymetrix gene expression array (AEG-1 KD → RFC5 downregulation), γ-H2AX foci assay, colony formation assay, flow cytometry, HR repair activity assay DNA and cell biology Medium 34042508
2025 High RFC5 expression in nasopharyngeal carcinoma cells enhances cisplatin-induced DNA damage repair, reduces micronucleus formation, suppresses cGAS-STING pathway activation, limits inflammatory mediator production, and promotes T cell exhaustion (elevated PD-1, LAG-3, CTLA-4; reduced IFN-γ and TNF-α secretion by CD8+ T cells) in vivo, linking RFC5-mediated DNA repair to immune evasion. RFC5 expression manipulation in NPC cells, micronuclei quantification, cGAS-STING pathway activity assays, in vivo tumor immune microenvironment analysis (cytokine measurement, T cell marker expression) Cellular signalling Medium 41192524
2024 SRSF10 promotes colorectal cancer progression by generating an aberrantly spliced exclusion isoform of RFC5 (AS1 exclusion of exon 2). SRSF10 knockdown alters RFC5 pre-mRNA splicing (detected by agarose gel electrophoresis of transcripts), affecting DNA replication and cell cycle progression in CRC cells. SRSF10 knockdown/overexpression, agarose gel electrophoresis of RFC5 splice variants, CCK8, transwell, flow cytometry Technology in cancer research & treatment Low 39110418

Source papers

Stage 0 corpus · 11 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
1996 Rfc5, a small subunit of replication factor C complex, couples DNA replication and mitosis in budding yeast. Proceedings of the National Academy of Sciences of the United States of America 87 8692942
1997 Rfc5, a replication factor C component, is required for regulation of Rad53 protein kinase in the yeast checkpoint pathway. Molecular and cellular biology 80 9315648
1998 Functional and physical interaction between Rad24 and Rfc5 in the yeast checkpoint pathways. Molecular and cellular biology 69 9710632
2000 Rfc5, in cooperation with rad24, controls DNA damage checkpoints throughout the cell cycle in Saccharomyces cerevisiae. Molecular and cellular biology 67 10913172
1995 Assignment of the 36.5-kDa (RFC5), 37-kDa (RFC4), 38-kDa (RFC3), and 40-kDa (RFC2) subunit genes of human replication factor C to chromosome bands 12q24.2-q24.3, 3q27, 13q12.3-q13, and 7q11.23. Genomics 40 7774928
2017 FoxM1-mediated RFC5 expression promotes temozolomide resistance. Cell biology and toxicology 37 28185110
2021 AEG-1 Knockdown Sensitizes Glioma Cells to Radiation Through Impairing Homologous Recombination Via Targeting RFC5. DNA and cell biology 11 34042508
2023 RFC5, regulated by circ_0038985/miR-3614-5p, functions as an oncogene in the progression of colorectal cancer. Molecular carcinogenesis 10 36988339
2024 High Expression of SRSF10 Promotes Colorectal Cancer Progression by Aberrant Alternative Splicing of RFC5. Technology in cancer research & treatment 6 39110418
2000 Allele-specific interactions between the yeast RFC1 and RFC5 genes suggest a basis for RFC subunit-subunit interactions. Molecular & general genetics : MGG 5 11129041
2025 RFC5 enhances DNA damage response and immune escape via suppressing the cGAS-STING pathway in nasopharyngeal carcinoma. Cellular signalling 2 41192524

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