Affinage

POLR3F

DNA-directed RNA polymerase III subunit RPC6 · UniProt Q9H1D9

Round 2 corrected
Length
316 aa
Mass
35.7 kDa
Annotated
2026-04-28
69 papers in source corpus 9 papers cited in narrative 9 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

POLR3F (RPC6/hRPC39/C34) is a subunit of the RNA polymerase III initiation-specific heterotrimer (with RPC32 and RPC62) that is essential for promoter-dependent transcription initiation but dispensable for elongation and termination. The heterotrimer dissociates from the Pol III core under partial denaturing conditions, and reconstitution of recombinant hRPC32–hRPC39–hRPC62 restores specific initiation; POLR3F directly contacts both TFIIIB components (TBP, BRF1) and TFIIIC (hTFIIIC90), thereby bridging the polymerase to its promoter-recognition machinery (PMID:9171375, PMID:9312031, PMID:10523658). Cryo-EM structures of human Pol III reveal that an iron–sulfur cluster, absent in yeast, tethers this heterotrimer to the polymerase core, and the repressor MAF1 competes with BRF1 for binding to the WH1 domain of the C34 subunit in a phosphorylation-regulated manner (PMID:33558764, PMID:32641350). Beyond canonical gene transcription, the intact Pol III holoenzyme containing POLR3F functions as a cytosolic DNA sensor that transcribes AT-rich dsDNA into 5′-triphosphate RNA to trigger RIG-I–dependent innate immune signaling (PMID:19631370).

Mechanistic history

Synthesis pass · year-by-year structured walk · 5 steps
  1. 1997 High

    Establishing that C34/hRPC39 resides in an initiation-specific subcomplex and is required for promoter-dependent transcription but not for catalytic elongation resolved a long-standing question about how Pol III distinguishes specific promoters from random templates.

    Evidence Purification of mutant yeast Pol III and reconstitution of recombinant human hRPC32–hRPC39–hRPC62 subcomplex with functional transcription assays

    PMID:9171375 PMID:9312031

    Open questions at the time
    • Structural basis for how the heterotrimer engages the Pol III core was unknown
    • It was unclear whether hRPC39 contacts TFIIIC directly or only through TFIIIB
  2. 1999 High

    Mapping the C34–TFIIIB70(BRF1) and C34/hRPC39–hTFIIIC90 interfaces defined POLR3F as a dual bridge connecting the polymerase simultaneously to both basal transcription factors, explaining how Pol III is recruited to pre-initiation complexes.

    Evidence Mutagenesis of TFIIIB70 with co-lethality analysis in yeast; co-immunoprecipitation of hTFIIIC90 with hRPC39 and hRPC62 in human extracts

    PMID:10329159 PMID:10523658

    Open questions at the time
    • No atomic-resolution map of the C34–BRF1 or C34–TFIIIC90 contact surfaces
    • Whether the TFIIIC interaction is maintained throughout initiation or only during recruitment was unresolved
  3. 2009 High

    Discovery that the Pol III holoenzyme (containing POLR3F) acts as a cytosolic DNA sensor expanded its known biology beyond nuclear gene transcription into innate immunity, showing it transcribes AT-rich DNA into 5′-ppp RNA to activate RIG-I signaling.

    Evidence Biochemical purification of the responsible polymerase, siRNA knockdown of Pol III subunits, IFN-β reporter assays, and Legionella infection model

    PMID:19631370

    Open questions at the time
    • The specific contribution of POLR3F versus other subunits to cytosolic DNA sensing was not dissected
    • No structure of the Pol III–cytosolic DNA complex is available
  4. 2020 Medium

    Demonstration that MAF1 and BRF1 compete for the WH1 domain of C34 (POLR3F ortholog), with MAF1 affinity modulated by phosphorylation, revealed the molecular switch controlling Pol III repression and derepression.

    Evidence NMR structural analysis of citrus MAF1, co-immunoprecipitation, phosphorylation assays, and competition binding assays with plant orthologs

    PMID:32641350

    Open questions at the time
    • Demonstrated in a plant ortholog system; direct validation of this competitive mechanism on human POLR3F is lacking
    • Identity and hierarchy of kinases regulating MAF1 phosphorylation at the C34 interface in mammals remain unresolved
  5. 2021 High

    High-resolution cryo-EM of human Pol III revealed that an iron–sulfur cluster, absent in yeast, anchors the POLR3F-containing heterotrimer to the polymerase core, providing the first structural explanation for how this subcomplex is physically integrated into the human enzyme.

    Evidence Cryo-EM of human Pol III at 2.8–3.3 Å resolution in unbound and transcribing states

    PMID:33558764

    Open questions at the time
    • No structure captures the heterotrimer engaged with both TFIIIB and TFIIIC simultaneously
    • Functional role of the iron–sulfur cluster beyond structural tethering has not been tested

Open questions

Synthesis pass · forward-looking unresolved questions
  • It remains unknown how the POLR3F-containing heterotrimer coordinates promoter opening (open complex formation) at atomic resolution, whether the iron–sulfur cluster serves a redox-sensing or regulatory role, and how the MAF1–C34 competition is regulated in human cells under different nutrient and stress conditions.
  • No structural snapshot of the open complex with the heterotrimer engaged on melted DNA
  • No functional dissection of the Fe–S cluster beyond structural tethering
  • MAF1–C34 competitive binding not validated biochemically with human proteins

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0003677 DNA binding 3 GO:0140098 catalytic activity, acting on RNA 3 GO:0005198 structural molecule activity 2
Localization
GO:0005654 nucleoplasm 2 GO:0005829 cytosol 1
Pathway
R-HSA-74160 Gene expression (Transcription) 4 R-HSA-168256 Immune System 1
Partners
BRF1GTF3C4MAF1PKP2POLR3BPOLR3CPOLR3ETBP
Complex memberships
Pol III initiation-specific heterotrimer (RPC32–RPC39–RPC62)RNA polymerase III

Evidence

Reading pass · 9 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
1997 The yeast C34 subunit (ortholog of human POLR3F/hRPC39) is part of an RNA Pol III-specific subunit that directly interacts with TFIIIB70 (BRF1). Mutations in RPC34 that impair growth also impaired C34 interaction with TFIIIB70. Mutant pol III had normal catalytic activity on non-specific templates but was defective in promoter-dependent transcription initiation, demonstrating that C34 is required for Pol III recruitment to pre-initiation complexes and for open complex formation. Site-directed mutagenesis of RPC34, purification of mutant Pol III, in vitro transcription assays on poly[d(A-T)] and specific gene templates, interaction assays with TFIIIB70 The EMBO journal High 9312031
1997 Human POLR3F (hRPC39) is part of a specific subcomplex with hRPC32 and hRPC62 that dissociates from the human Pol III core under partial denaturing conditions. The core Pol III lacking this subcomplex can perform elongation and termination on tailed templates but cannot support promoter-dependent transcription initiation; addition of the recombinant hRPC32-hRPC39-hRPC62 subcomplex restores specific initiation. hRPC39 physically interacts with both hTBP and hTFIIIB90, indicating it mediates Pol III recruitment to the TFIIIB-DNA complex. Immunopurification of human Pol III, sucrose gradient sedimentation, reconstitution of recombinant subcomplex, in vitro transcription assays, physical interaction assays (co-immunoprecipitation, pulldown) Genes & development High 9171375
1999 Yeast TFIIIB70 C-terminal conserved regions II and III are required for interaction with the C34 subunit (yeast POLR3F ortholog). Conditional mutations in region II that impaired C34 interaction were co-lethal with rpc34 mutations, establishing a functional genetic interaction. These mutations also impaired assembly of TFIIIB·TFIIIC·DNA complexes and in vitro transcription of the SUP4 tRNA gene, placing C34-TFIIIB70 interaction as essential for pre-initiation complex assembly. Extensive mutagenesis of TFIIIB70, co-lethality/epistasis analysis with rpc34 mutations, in vitro transcription assays, interaction assays Journal of molecular biology High 10329159
1999 The hTFIIIC90 subunit of human TFIIIC physically interacts with hRPC39 (POLR3F) and hRPC62 subunits of the Pol III initiation-specific subcomplex, as shown by immunodepletion and immunoprecipitation. This interaction is proposed to facilitate TFIIIB and RNA Pol III recruitment to the pre-initiation complex. Immunodepletion, co-immunoprecipitation, in vitro interaction assays Molecular and cellular biology Medium 10523658
2001 Plakophilin 2, a dual-location desmosomal/nuclear protein, is present in the RNA Pol III holoenzyme (but not the core complex) and co-immunoselects with other Pol III subunits and TFIIIB. Plakophilin 2 binds specifically to RPC155 (the largest Pol III subunit) in vitro, placing it in a nuclear particle that contains the full Pol III holoenzyme including POLR3F-containing subcomplex. Co-immunoselection, in vitro binding assays, colocalization by immunofluorescence Proceedings of the National Academy of Sciences of the United States of America Medium 11416169
2019 Endogenous microDNA (small extrachromosomal circular DNAs) associate with RNA polymerase subunits including POLR3F, as identified by affinity purification, suggesting POLR3F participates in microDNA-mediated transcription of small regulatory RNAs independent of canonical promoters. Affinity purification of microDNA-associated proteins, identification by mass spectrometry Nucleic acids research Low 30828735
2020 In citrus (plant ortholog system), MAF1 binds predominantly to the WH1 domain of the Pol III C34 subunit (CsC34, ortholog of POLR3F), and the MAF1 phosphoregulatory region (loop-3 and α-helix-2) contributes to this interaction. Phosphorylation of MAF1's phosphoregulatory region decreases its affinity for CsC34, leading to Pol III derepression. The C-terminal region of BRF1 (TFIIIB component) competes with MAF1 for CsC34 interaction, revealing a competitive mechanism regulating Pol III activity. NMR structural analysis of CsMAF1, co-immunoprecipitation, phosphorylation assays with citrus AGC1 kinase, competition binding assays The Plant cell Medium 32641350
2021 Cryo-EM structures of human RNA Pol III at 2.8–3.3 Å resolution in unbound and transcribing states reveal that an iron-sulfur cluster tethers the heterotrimer subcomplex (which includes POLR3F/RPC39) to the Pol III core, an element absent in yeast Pol III. The cancer-associated RPC7α isoform binds the polymerase clamp and may interfere with Pol III inhibition by tumor suppressor MAF1. Disease-related mutations are mapped onto the structure. Cryo-electron microscopy at 2.8–3.3 Å resolution Nature structural & molecular biology High 33558764
2009 RNA polymerase III (which contains POLR3F as part of its initiation subcomplex) acts as a cytosolic DNA sensor: it transcribes AT-rich dsDNA (poly(dA-dT)) into 5'-triphosphate RNA, which then activates RIG-I and MAVS to induce IFN-β. Inhibition of RNA Pol III prevents IFN-β induction by cytosolic DNA and by intracellular bacteria (Legionella pneumophila). Biochemical purification of the responsible enzyme, siRNA knockdown of Pol III, in vitro transcription assays, reporter gene assays for IFN-β induction Cell High 19631370

Source papers

Stage 0 corpus · 69 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2005 A human protein-protein interaction network: a resource for annotating the proteome. Cell 1704 16169070
2002 Generation and initial analysis of more than 15,000 full-length human and mouse cDNA sequences. Proceedings of the National Academy of Sciences of the United States of America 1479 12477932
2008 Identification of host proteins required for HIV infection through a functional genomic screen. Science (New York, N.Y.) 1165 18187620
2015 The BioPlex Network: A Systematic Exploration of the Human Interactome. Cell 1118 26186194
2017 Architecture of the human interactome defines protein communities and disease networks. Nature 1085 28514442
2015 A human interactome in three quantitative dimensions organized by stoichiometries and abundances. Cell 1015 26496610
2009 RNA polymerase III detects cytosolic DNA and induces type I interferons through the RIG-I pathway. Cell 977 19631370
2020 A reference map of the human binary protein interactome. Nature 849 32296183
2009 A genome-wide RNAi screen identifies multiple synthetic lethal interactions with the Ras oncogene. Cell 843 19490893
2003 Complete sequencing and characterization of 21,243 full-length human cDNAs. Nature genetics 754 14702039
2021 Dual proteome-scale networks reveal cell-specific remodeling of the human interactome. Cell 705 33961781
2012 A census of human soluble protein complexes. Cell 689 22939629
2011 Phylogenetic-based propagation of functional annotations within the Gene Ontology consortium. Briefings in bioinformatics 656 21873635
1994 Oligo-capping: a simple method to replace the cap structure of eukaryotic mRNAs with oligoribonucleotides. Gene 492 8125298
2004 The status, quality, and expansion of the NIH full-length cDNA project: the Mammalian Gene Collection (MGC). Genome research 438 15489334
2022 OpenCell: Endogenous tagging for the cartography of human cellular organization. Science (New York, N.Y.) 432 35271311
2005 Diversification of transcriptional modulation: large-scale identification and characterization of putative alternative promoters of human genes. Genome research 409 16344560
2015 Panorama of ancient metazoan macromolecular complexes. Nature 407 26344197
1996 Generation and analysis of 280,000 human expressed sequence tags. Genome research 376 8889549
2011 Toward an understanding of the protein interaction network of the human liver. Molecular systems biology 207 21988832
2005 Different from the HIV fusion inhibitor C34, the anti-HIV drug Fuzeon (T-20) inhibits HIV-1 entry by targeting multiple sites in gp41 and gp120. The Journal of biological chemistry 203 15640162
2001 The DNA sequence and comparative analysis of human chromosome 20. Nature 168 11780052
2020 Synthetic Lethal and Resistance Interactions with BET Bromodomain Inhibitors in Triple-Negative Breast Cancer. Molecular cell 159 32416067
2009 Ubiquitin-mediated proteolysis of HuR by heat shock. The EMBO journal 142 19322201
2019 Small extrachromosomal circular DNAs, microDNA, produce short regulatory RNAs that suppress gene expression independent of canonical promoters. Nucleic acids research 136 30828735
1997 Three human RNA polymerase III-specific subunits form a subcomplex with a selective function in specific transcription initiation. Genes & development 132 9171375
2018 Histone Interaction Landscapes Visualized by Crosslinking Mass Spectrometry in Intact Cell Nuclei. Molecular & cellular proteomics : MCP 101 30021884
2001 Nuclear particles containing RNA polymerase III complexes associated with the junctional plaque protein plakophilin 2. Proceedings of the National Academy of Sciences of the United States of America 98 11416169
1997 Dual role of the C34 subunit of RNA polymerase III in transcription initiation. The EMBO journal 94 9312031
2013 A Y2H-seq approach defines the human protein methyltransferase interactome. Nature methods 93 23455924
2006 Maternal transfer of complement components C3-1, C3-3, C3-4, C4, C5, C7, Bf, and Df to offspring in rainbow trout (Oncorhynchus mykiss). Immunogenetics 93 16550351
1999 The TFIIIC90 subunit of TFIIIC interacts with multiple components of the RNA polymerase III machinery and contains a histone-specific acetyltransferase activity. Molecular and cellular biology 86 10523658
2021 Cryo-EM structures of human RNA polymerase III in its unbound and transcribing states. Nature structural & molecular biology 80 33558764
2003 HIV-1 resistance to the gp41-dependent fusion inhibitor C-34. Antiviral research 62 12895697
2008 Albumin-conjugated C34 peptide HIV-1 fusion inhibitor: equipotent to C34 and T-20 in vitro with sustained activity in SCID-hu Thy/Liv mice. The Journal of biological chemistry 59 18809675
2013 Conjugation of cholesterol to HIV-1 fusion inhibitor C34 increases peptide-membrane interactions potentiating its action. PloS one 52 23565220
1999 Mutagenesis of yeast TFIIIB70 reveals C-terminal residues critical for interaction with TBP and C34. Journal of molecular biology 44 10329159
2021 TLR4-IN-C34 Inhibits Lipopolysaccharide-Stimulated Inflammatory Responses via Downregulating TLR4/MyD88/NF-κB/NLRP3 Signaling Pathway and Reducing ROS Generation in BV2 Cells. Inflammation 33 34727285
2017 Alismanin A, a Triterpenoid with a C34 Skeleton from Alisma orientale as a Natural Agonist of Human Pregnane X Receptor. Organic letters 33 29016144
2005 Differential inhibition of HIV-1 and SIV envelope-mediated cell fusion by C34 peptides derived from the C-terminal heptad repeat of gp41 from diverse strains of HIV-1, HIV-2, and SIV. Journal of medicinal chemistry 28 15828842
2001 Variant toxin B and a functional toxin A produced by Clostridium difficile C34. FEMS microbiology letters 27 11430410
2019 Heterologous Expression of a Cryptic Gene Cluster from Streptomyces leeuwenhoekii C34T Yields a Novel Lasso Peptide, Leepeptin. Applied and environmental microbiology 22 31562169
1993 Anticodon bases C34 and C35 are major, positive, identity elements in Saccharomyces cerevisiae tRNA(Trp). Nucleic acids research 15 8255761
2019 Pharmacological inhibition of TLR4/NF-κB with TLR4-IN-C34 attenuated microcystin-leucine arginine toxicity in bovine Sertoli cells. Journal of applied toxicology : JAT 14 30671980
2016 Glycosyl Phosphatidylinositol-Anchored C34 Peptide Derived From Human Immunodeficiency Virus Type 1 Gp41 Is a Potent Entry Inhibitor. Journal of neuroimmune pharmacology : the official journal of the Society on NeuroImmune Pharmacology 14 27155865
2005 Development of anti-HIV agents targeting dynamic supramolecular mechanism: entry and fusion inhibitors based on CXCR4/CCR5 antagonists and gp41-C34-remodeling peptides. Current HIV research 14 16250877
2020 Bungsteroid A: One Unusual C34 Pentacyclic Steroid Analogue from Zanthoxylum bungeanum Maxim. The Journal of organic chemistry 12 32702985
2019 Dimeric C34 Derivatives Linked through Disulfide Bridges as New HIV-1 Fusion Inhibitors. Chembiochem : a European journal of chemical biology 12 31012222
2012 Preexposure prophylaxis with albumin-conjugated C34 peptide HIV-1 fusion inhibitor in SCID-hu Thy/Liv mice. Antimicrobial agents and chemotherapy 12 22252805
2007 C34, a membrane fusion inhibitor, blocks HIV infection of langerhans cells and viral transmission to T cells. The Journal of investigative dermatology 12 17255952
2019 Synthesis and Biological Activities of Aplyronine A Analogues toward the Development of Antitumor Protein-Protein Interaction Inducers between Actin and Tubulin: Conjugation of the C1-C9 Macrolactone Part and the C24-C34 Side Chain. ACS omega 11 31459949
2005 Clinical significance and neuropathology of primary MADD in C34-T and G468-T mutations of the AMPD1 gene. Clinical neuropathology 11 15803807
2018 Analysis of metabolic networks of Streptomyces leeuwenhoekii C34 by means of a genome scale model: Prediction of modifications that enhance the production of specialized metabolites. Biotechnology and bioengineering 10 29578590
2017 A first-in-human study of the novel HIV-fusion inhibitor C34-PEG4-Chol. Scientific reports 9 28842581
2015 Molecular weight-dependent degradation of D-lactate-containing polyesters by polyhydroxyalkanoate depolymerases from Variovorax sp. C34 and Alcaligenes faecalis T1. Applied microbiology and biotechnology 9 26109003
2015 The C34 Peptide Fusion Inhibitor Binds to the Six-Helix Bundle Core Domain of HIV-1 gp41 by Displacement of the C-Terminal Helical Repeat Region. Biochemistry 9 26506247
2011 The effect of SAMe and betaine on Hepa 1-6, C34 and E47 liver cell survival in vitro. Experimental and molecular pathology 9 22032937
2022 Pharmacological inhibition of toll-like receptor 4 with TLR4-IN-C34 modulates the intestinal flora homeostasis and the MyD88/NF-κB axis in ulcerative colitis. European journal of pharmacology 8 36152840
2022 TLR4-IN-C34 protects against acute kidney injury via modulating TLR4/MyD88/NF-κb axis, MAPK, and apoptosis. Iranian journal of basic medical sciences 7 36474570
2021 The C29-C34 parts of antitumor macrolide aplyronine A serve as versatile actin-affinity tags. Chemical communications (Cambridge, England) 7 34553712
2020 The MAF1 Phosphoregulatory Region Controls MAF1 Interaction with the RNA Polymerase III C34 Subunit and Transcriptional Repression in Plants. The Plant cell 7 32641350
2014 Molecular dynamics studies of the inhibitor C34 binding to the wild-type and mutant HIV-1 gp41: inhibitory and drug resistant mechanism. PloS one 6 25393106
2024 A New Chimeric Antibody against the HIV-1 Fusion Inhibitory Peptide MT-C34 with a High Affinity and Fc-Mediated Cellular Cytotoxicity. Biology 5 39336102
2025 Benzo[a]phenoxazine derivative C34 efficacy against fluconazole-resistant Candida spp. Microbial pathogenesis 3 40122407
2005 Hyperleptinemia and its relation with peripheral C34(+)CD7(+) stem cells in renal transplant recipients. Transplant immunology 3 16431293
2025 Neuroprotective Effects of Early TLR4 Blockade with Compound C34 in Temporal Lobe Epilepsy: Alleviation of Neuroinflammation and Apoptosis. Iranian journal of pharmaceutical research : IJPR 0 40718441
2025 TLR4-IN-C34 attenuates the progression of osteoarthritis through inhibiting inflammation, angiogenesis and pain. Cellular signalling 0 40854507
2025 Metabolic Engineering of Streptomyces leeuwenhoekii C34T to Increase Chaxamycin Production Based on the iVR1007 Genome-Scale Model. Biotechnology and bioengineering 0 40956005
2024 [Methods to Increase the Efficiency of Knock-in of a Construct Encoding the HIV-1 Fusion Inhibitor, MT-C34 Peptide, into the CXCR4 Locus in the CEM/R5 T Cell Line]. Molekuliarnaia biologiia 0 39709562