{"gene":"POLR2G","run_date":"2026-06-10T06:43:35","timeline":{"discoveries":[{"year":1993,"finding":"RPB7 is essential for cell viability in S. cerevisiae; deletion of RPB7 is lethal, whereas deletion of RPB4 causes only conditional growth defects. RPB7 association with Pol II is dependent on RPB4, as polymerase purified from rpb4Δ cells also lacks RPB7.","method":"Gene deletion, yeast genetics, polymerase purification","journal":"Yeast","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean genetic knockouts with defined viability phenotypes, replicated in multiple follow-up studies","pmids":["8488730"],"is_preprint":false},{"year":1995,"finding":"Human hsRPB7 is a functional ortholog of yeast RPB7: it assembles into a complete 12-subunit yeast Pol II complex (shown by immunoprecipitation) and rescues the essential RPB7 deletion at moderate temperatures. However, hsRPB7 shows reduced affinity for yeast RPB4 compared to yeast RPB7, explaining failure to fully complement at temperature extremes.","method":"Complementation assay, immunoprecipitation of Pol II from yeast expressing hsRPB7, yeast two-hybrid","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — complementation, immunoprecipitation, and two-hybrid results are mutually consistent and replicated by later studies","pmids":["7579693"],"is_preprint":false},{"year":1998,"finding":"Oncogenic EWS-Fli1 specifically interacts with hsRPB7 (the human Pol II seventh subunit) via the EWS amino terminus; this interaction was identified by yeast two-hybrid and confirmed by in vitro immunoprecipitation. hsRPB7 copurifies with EWS-Fli1 but not with wild-type Fli1 from nuclear extracts. Overexpression of hsRPB7 specifically enhances gene activation by EWS-chimeric transcription factors.","method":"Yeast two-hybrid, in vitro immunoprecipitation, nuclear extract copurification, overexpression transactivation assay","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 / Strong — yeast two-hybrid confirmed by reciprocal co-IP and nuclear extract copurification plus functional overexpression assay in single study","pmids":["9704926"],"is_preprint":false},{"year":1998,"finding":"Human hsRPB4 strongly and specifically interacts with hsRPB7 in yeast and mammalian cells and copurifies with intact Pol II. Structure-function mapping defined the interaction interface between hsRPB4 and hsRPB7. hsRPB4 does not interact with yeast RPB7, showing species-specific divergence while retaining partial complementation of rpb4− yeast phenotypes.","method":"Yeast two-hybrid, coimmunoprecipitation in mammalian cells, polymerase copurification, truncation/deletion mapping","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP in two cell systems plus copurification with Pol II and domain mapping","pmids":["9528765"],"is_preprint":false},{"year":1998,"finding":"Cryo-EM difference mapping showed that Rpb4/Rpb7 form part of the floor of the DNA-binding cleft of yeast Pol II. Surface plasmon resonance demonstrated that Rpb4/Rpb7 stabilize a minimal pre-initiation complex containing promoter DNA, TBP, TFIIB and polymerase, suggesting a role in coupling DNA entry into the active center cleft to cleft closure.","method":"Cryo-EM 3D reconstruction, difference mapping, surface plasmon resonance","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure with SPR functional validation in a single rigorous study","pmids":["9545247"],"is_preprint":false},{"year":1999,"finding":"Ligand-free RARα interacts with hsRPB7 through its DNA-binding domain (in the absence of retinoic acid); RA addition disrupts the interaction. Overexpressed hRARα (no RA) represses Pol II-dependent activators (AP-1, GR), and this repression is relieved by co-transfected hsRPB7, indicating that unliganded RARα sequesters hsRPB7 to repress transcription.","method":"Yeast two-hybrid, truncation analysis, transient transfection repression/rescue assays","journal":"Endocrine","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — two-hybrid plus functional rescue assay, single lab, no reciprocal co-IP","pmids":["10484292"],"is_preprint":false},{"year":1999,"finding":"Rpb7 can associate with Pol II lacking Rpb4 (Pol IIΔ4) when overexpressed, demonstrated by two reciprocal coimmunoprecipitation experiments. This Rpb4-independent interaction supports transcription under mild stress, establishing that a major role of Rpb4 is to augment the affinity of Rpb7 for Pol II.","method":"Suppressor screen, reciprocal coimmunoprecipitation, transcription assays in rpb4Δ cells","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP with functional (transcription) readout and genetic suppressor data converging on same conclusion","pmids":["10082533"],"is_preprint":false},{"year":2000,"finding":"The purified Rpb4/Rpb7 complex binds single-stranded DNA and RNA via an OB-fold motif in Rpb7. A small deletion in the OB-fold abolished nucleic acid binding without disrupting the Rpb4/Rpb7 complex or its association with Pol II, yet destroyed transcription activity. A separate OB-fold deletion blocked transcription but not nucleic acid binding, indicating the OB-fold mediates both nucleic acid binding and protein-protein interactions required for initiation. Template competition showed Rpb4/Rpb7 is not needed for stable polymerase recruitment but acts at a post-recruitment step.","method":"Purified complex binding assays (ssDNA/RNA), deletion mutagenesis, reconstituted transcription assay, template competition","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution with mutagenesis and multiple orthogonal functional assays in one study","pmids":["11087726"],"is_preprint":false},{"year":2001,"finding":"Crystal structure of the archaeal RPB7/RPB4 homologs (Methanococcus jannaschii subunits E and F) was determined, revealing an elongated two-domain structure for subunit E with two potential RNA-binding motifs, and subunit F wrapping around subunit E at the domain interface. A model was proposed in which the RNA-binding face of RPB7 contacts the nascent RNA transcript.","method":"X-ray crystallography","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure of archaeal ortholog complex, foundational structural study widely replicated","pmids":["11741548"],"is_preprint":false},{"year":2003,"finding":"pVHL directly binds to hsRPB7 through its beta-domain; naturally occurring beta-domain mutations decrease this interaction. Introduction of wild-type pVHL into VHL-mutant renal carcinoma cells promotes ubiquitylation and proteasomal degradation of hsRPB7, reduces its nuclear accumulation, and suppresses hsRPB7-induced VEGF promoter transactivation and VEGF secretion.","method":"Yeast two-hybrid, co-immunoprecipitation, ubiquitylation assay, proteasome inhibitor experiments, luciferase reporter assay, ELISA","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct binding confirmed by co-IP, ubiquitylation demonstrated biochemically, functional consequence shown by reporter and VEGF secretion assays; multiple orthogonal methods","pmids":["12912922"],"is_preprint":false},{"year":2003,"finding":"In fission yeast, Rpb7 directly interacts in vitro with Seb1 (the S. pombe homolog of Nrd1, an RNA-binding protein for 3′-end formation of snRNA/snoRNA). Two-hybrid and in vitro binding identified Glu166/Asp167 near the Rpb7 C-terminus as critical for the interaction. S. cerevisiae Rpb7 also interacts with Nrd1, showing evolutionary conservation. This suggests Rpb7 anchors an RNA-processing factor to the Pol II apparatus.","method":"Two-hybrid screen, in vitro direct binding, site-directed mutagenesis","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro direct binding plus two-hybrid with mutagenesis, single lab","pmids":["12907709"],"is_preprint":false},{"year":2003,"finding":"A conditional mutation in the shared Rpb6 subunit (Q100R) causes selective loss of Rpb4 and Rpb7 from purified Pol II. Interaction experiments support a direct association between Rpb6 and Rpb4, identifying Rpb6 as one contact point through which the Rpb4/Rpb7 subcomplex associates with the Pol II core.","method":"Genetic screen, polymerase purification, interaction experiments (pulldown/co-IP)","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — purification of mutant polymerase plus direct interaction experiments, single lab","pmids":["12697831"],"is_preprint":false},{"year":2005,"finding":"UV cross-linking of uridine analogs in the first 6 nt of nascent RNA demonstrates that the emerging transcript contacts Rpb7 in Pol II elongation complexes carrying 26–32 nt transcripts; this interaction is greatly reduced at 41–43 nt and absent at 125 nt, establishing groove 1 (pointing toward Rpb4/Rpb7) as the nascent RNA exit path.","method":"UV cross-linking with uridine analogs on reconstituted elongation complexes","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstituted complexes with defined transcript lengths and site-specific cross-linking; directly identifies RNA-Rpb7 contact","pmids":["16327806"],"is_preprint":false},{"year":2005,"finding":"Crystal structure of the human Rpb4/Rpb7 heterodimer was determined at 2.7 Å. Site-directed mutagenesis of conserved surface residues in Rpb7 (and archaeal subunit E), combined with EMSA RNA-binding assays, identified an elongated surface spanning the OB-fold B4–B5 loop and extending to the N-terminal domain as the RNA-binding interface.","method":"X-ray crystallography, site-directed mutagenesis, EMSA","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure at 2.7 Å with mutagenesis-validated RNA-binding surface, multiple orthogonal methods","pmids":["16282592"],"is_preprint":false},{"year":2005,"finding":"Fission yeast Rpb7 has a specific role in transcription of centromeric pre-siRNA precursors required for RNAi-directed heterochromatin formation. The rpb7-G150D point mutation specifically impairs centromeric pre-siRNA transcription (and thus RNAi silencing) without equivalent effects seen with other Pol II subunit mutants.","method":"Point mutation analysis, centromeric transcription assays, RNAi/heterochromatin reporter assays","journal":"Genes & development","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — defined point mutation with specific transcriptional and silencing phenotypes, single lab","pmids":["16204182"],"is_preprint":false},{"year":2005,"finding":"In fission yeast, Rpb7 interacts with both GAPDH and actin. GAPDH was identified as an Rpb7-binding protein by two-hybrid screen and confirmed by affinity purification from S. pombe extract using an Rpb4/Rpb7-coupled column. Actin was identified as a Pol II-associated protein and shown to interact with Rpb7.","method":"Two-hybrid screen, affinity purification (Rpb4/Rpb7-coupled column pull-down)","journal":"FEBS letters","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, two-hybrid plus single affinity-pull-down without reciprocal co-IP","pmids":["15620689"],"is_preprint":false},{"year":2005,"finding":"Mutations in the N-terminal RNP-like domain of S. cerevisiae Rpb7 impair stress responses (high-temperature growth, sporulation) and increase Rpb7's dependence on Rpb4 for Pol II interaction. RNA polymerase pulldown assays revealed two crucial interaction points between the Rpb4/Rpb7 subcomplex and the core: the N-terminal RNP-like domain of Rpb7 and the partially ordered N-terminus of Rpb4 (contacting Rpb2).","method":"Mutagenesis, RNA polymerase pulldown assay, complementation analysis","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo pulldown with defined mutants identifying two interaction points, single lab","pmids":["18056993"],"is_preprint":false},{"year":2007,"finding":"In Pyrococcus furiosus, the archaeal RPB7 ortholog E' (subunit E') stimulates Pol II core enzyme activity at low temperatures and promotes open complex formation (bubble formation at multiple positions in the transcription bubble), as shown by permanganate footprinting of reconstituted transcription complexes. Subunit F did not significantly contribute to catalytic activity.","method":"Reconstitution of archaeal RNA polymerase, permanganate footprinting, in vitro transcription at varying temperatures","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — fully reconstituted system with permanganate footprinting and kinetic analyses; rigorous in vitro mechanistic dissection","pmids":["17311916"],"is_preprint":false},{"year":2008,"finding":"ChIP experiments in live human cells show that Rpb7 remains associated with Pol II during early elongation (not solely at initiation as previously believed), and under stress conditions Rpb7 association extends to late elongation stages, suggesting a role in stabilizing transcribing polymerase throughout the transcription cycle.","method":"TAP-ChIP (chromatin immunoprecipitation) with quantitative PCR across promoter-proximal and downstream transcribed regions in human cells","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP in living human cells with multiple gene loci and stress conditions, single lab","pmids":["17848138"],"is_preprint":false},{"year":2008,"finding":"Pull-down assays confirmed direct in vitro interaction between the first 57 residues of the EWS N-terminal activation domain (EAD) and hsRPB7, forming a stable complex.","method":"In vitro pull-down assay with recombinant proteins","journal":"Molecular biology reports","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, single in vitro pull-down method only","pmids":["18607770"],"is_preprint":false},{"year":2013,"finding":"Quantitative proteomics demonstrated that Rpb4/Rpb7 dissociate from Pol II upon interaction with transcriptional elongation-associated proteins recruited to the hyperphosphorylated CTD (Ser2-P form). Pol II isolated through Rpb7 is depleted in Ser2-CTD phosphorylation, indicating Rpb4/7 are dispensable during specific elongation stages.","method":"Quantitative proteomics (mass spectrometry), immunoprecipitation via Rpb7-tagged Pol II","journal":"Molecular & cellular proteomics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — quantitative MS with orthogonal co-IP approach, single lab","pmids":["23418395"],"is_preprint":false},{"year":2023,"finding":"Rpb7 functions as a repressor of transcription-coupled nucleotide excision repair (TCR) in S. cerevisiae. The Rpb7 region interacting with the KOW3 domain of Spt5 represses TCR through a common mechanism with Spt4/Spt5. The Rpb7 regions contacting Rpb4 and/or core RNAPII repress TCR largely independently of Spt4/Spt5, as shown by synergistic enhancement of TCR derepression when combined with spt4Δ.","method":"Genetic epistasis (double mutants), TCR assays at specific gene loci, UV sensitivity assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with defined domain mutations and multiple gene loci, single lab","pmids":["37380080"],"is_preprint":false},{"year":2025,"finding":"RPB7 stabilizes RPB1 (the largest Pol II subunit): depletion of RPB7 destabilizes RPB1 via the E3 ubiquitin ligase Cullin 3. Stabilization depends on the loop regions of RPB7, CDK9, the CTD and linker region of RPB1. RPB7 also interacts with the phosphatase CTDP1, which is required for RPB1 stability. RPB7 is essential for Pol II reinitiation, engages RNA-processing factors, and localizes to the RNA exit channel; its absence compromises RNA processing.","method":"RPB7 depletion experiments, co-immunoprecipitation (RPB7-CTDP1 interaction), ubiquitylation assay (Cullin 3), reinitiation assays, RNA processing assays, localization data","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (depletion, co-IP, ubiquitylation, functional assays) in single study, not yet independently replicated","pmids":["40038320"],"is_preprint":false},{"year":2005,"finding":"Mapping of the Rpb4–Rpb7 interaction interface by two-hybrid analysis with truncation mutants showed that both the amino- and carboxy-terminal domains of Rpb7 are required for interaction with Rpb4, and that N-terminal deletions of Rpb4 (beyond 49 residues) progressively abolish interaction with Rpb7.","method":"Yeast two-hybrid with truncation mutants, computational structural analysis","journal":"Biochemical and biophysical research communications","confidence":"Low","confidence_rationale":"Tier 3 / Weak — two-hybrid only, single lab, no in vitro validation","pmids":["15913559"],"is_preprint":false}],"current_model":"POLR2G (RPB7/hsRPB7) encodes the seventh subunit of RNA polymerase II that forms an obligate heterodimer with RPB4; this subcomplex protrudes from the Pol II core at the RNA exit groove (groove 1), contacts the nascent transcript through an OB-fold RNA-binding domain, stabilizes the RPB1 largest subunit via recruitment of the CTDP1 phosphatase (countering Cullin 3-mediated ubiquitylation), mediates a post-recruitment step in transcription initiation (open-complex formation and/or clamp closure), remains associated with elongating polymerase through early elongation and dissociates during late elongation, represses transcription-coupled nucleotide excision repair, couples RNA processing to transcription, and serves as a direct binding target/effector for oncoproteins (EWS-Fli1), tumor suppressors (pVHL, which ubiquitylates it for proteasomal degradation), and nuclear receptors (unliganded RARα), thereby linking core transcription machinery to signaling and post-transcriptional gene regulation."},"narrative":{"mechanistic_narrative":"POLR2G (RPB7/hsRPB7) encodes the essential seventh subunit of RNA polymerase II, which forms an obligate heterodimer with RPB4 that contacts the Pol II core and is required for cell viability [PMID:8488730, PMID:9528765]. The subcomplex sits in the floor of the DNA-binding cleft and stabilizes a minimal pre-initiation complex of promoter DNA, TBP, TFIIB and polymerase, acting at a post-recruitment step rather than in stable polymerase recruitment; RPB7's OB-fold mediates both nucleic-acid binding and the protein-protein contacts required for initiation, and the subunit promotes open-complex formation [PMID:9545247, PMID:11087726, PMID:17311916]. RPB7 lines the RNA exit path (groove 1): the nascent transcript cross-links to RPB7 during early elongation and the contact is lost as the transcript lengthens, consistent with RPB7 marking the RNA exit channel [PMID:16327806, PMID:16282592]. RPB7 stays associated with elongating polymerase through early elongation and dissociates upon recruitment of Ser2-phosphorylated CTD elongation factors during late elongation [PMID:17848138, PMID:23418395]. Through these contacts RPB7 couples transcription to RNA processing by anchoring 3'-end/processing factors such as Nrd1/Seb1 to the polymerase [PMID:12907709, PMID:40038320], and it stabilizes the largest subunit RPB1 by recruiting the CTDP1 phosphatase and countering Cullin-3-mediated ubiquitylation [PMID:40038320]. Beyond core transcription, RPB7 is a binding target for the oncogenic EWS-Fli1 fusion, whose EWS N-terminus contacts RPB7 to enhance EWS-chimera-driven transactivation [PMID:9704926], and for the tumor suppressor pVHL, which binds RPB7 through its beta-domain and targets it for ubiquitylation and proteasomal degradation, thereby limiting RPB7-driven VEGF promoter activation [PMID:12912922]. RPB7 also represses transcription-coupled nucleotide excision repair via domain contacts overlapping with the Spt4/Spt5 pathway [PMID:37380080].","teleology":[{"year":1993,"claim":"Established that RPB7 is an essential, RPB4-dependent component of Pol II, distinguishing it from the dispensable RPB4 and defining the architecture of the subcomplex's attachment to the core.","evidence":"Gene deletion and polymerase purification in S. cerevisiae","pmids":["8488730"],"confidence":"High","gaps":["Did not define the molecular contacts anchoring RPB7 to the core","Did not establish the biochemical function of RPB7"]},{"year":1995,"claim":"Showed functional conservation by demonstrating the human ortholog assembles into and rescues yeast Pol II, validating yeast as a model for human RPB7 mechanism.","evidence":"Complementation, immunoprecipitation and two-hybrid in yeast expressing hsRPB7","pmids":["7579693"],"confidence":"High","gaps":["Reduced affinity for yeast RPB4 left interface determinants undefined","No human-cell functional assay"]},{"year":1998,"claim":"Mapped the obligate RPB4–RPB7 heterodimer interface in human cells and placed the subcomplex in the floor of the DNA-binding cleft, where it stabilizes the pre-initiation complex.","evidence":"Reciprocal co-IP and domain mapping in mammalian cells; cryo-EM difference mapping and SPR in yeast","pmids":["9528765","9545247"],"confidence":"High","gaps":["Mechanism linking cleft positioning to clamp closure inferred not proven","Atomic-resolution interface not yet resolved at this stage"]},{"year":1998,"claim":"Identified RPB7 as a direct target of the oncogenic EWS-Fli1 fusion, linking the core transcription subunit to oncogenic transactivation.","evidence":"Yeast two-hybrid, in vitro co-IP, nuclear-extract copurification and overexpression transactivation assay","pmids":["9704926"],"confidence":"High","gaps":["Endogenous relevance in Ewing sarcoma cells not established","Mechanism by which RPB7 enhances chimera activity unresolved"]},{"year":2000,"claim":"Defined RPB7's OB-fold as the dual nucleic-acid-binding and initiation-competent element and showed the subcomplex acts at a post-recruitment step, not in stable polymerase loading.","evidence":"Purified-complex binding assays, OB-fold deletion mutagenesis, reconstituted transcription and template competition","pmids":["11087726"],"confidence":"High","gaps":["Precise post-recruitment step (open complex vs clamp closure) not separated","In vivo confirmation of OB-fold requirement absent"]},{"year":2001,"claim":"Provided the first atomic view of the RPB7/RPB4 fold from the archaeal homolog, predicting an RNA-binding face that contacts the nascent transcript.","evidence":"X-ray crystallography of Methanococcus jannaschii subunits E and F","pmids":["11741548"],"confidence":"High","gaps":["RNA contact was modeled, not observed","Archaeal divergence from eukaryotic Pol II left untested"]},{"year":2003,"claim":"Connected RPB7 to tumor-suppressor regulation by showing pVHL binds and targets RPB7 for ubiquitin-proteasome degradation, controlling RPB7-driven VEGF activation.","evidence":"Two-hybrid, co-IP, ubiquitylation and proteasome-inhibitor assays, luciferase reporter and ELISA in renal carcinoma cells","pmids":["12912922"],"confidence":"High","gaps":["Whether degradation affects assembled Pol II or only free RPB7 unclear","Direct E3 ligase chemistry not reconstituted"]},{"year":2003,"claim":"Implicated RPB7 in coupling transcription to RNA processing by identifying a conserved interaction with the 3'-end formation factor Nrd1/Seb1, and identified RPB6 as a core attachment point for the subcomplex.","evidence":"Two-hybrid, in vitro direct binding and site-directed mutagenesis (Seb1/Nrd1); conditional Rpb6 mutant and interaction assays","pmids":["12907709","12697831"],"confidence":"Medium","gaps":["No reciprocal co-IP for the Nrd1 interaction in mammalian cells","Functional consequence for processing not directly measured here"]},{"year":2005,"claim":"Localized the nascent RNA exit path to RPB7 and resolved the human heterodimer crystal structure with a mutagenesis-validated RNA-binding surface.","evidence":"Site-specific UV cross-linking on reconstituted elongation complexes; 2.7 Å human Rpb4/Rpb7 crystal structure with mutagenesis and EMSA","pmids":["16327806","16282592"],"confidence":"High","gaps":["Functional role of the transient RNA contact in elongation not defined","Dynamics of contact loss at longer transcripts not mechanistically explained"]},{"year":2007,"claim":"Demonstrated a catalytic-stimulatory role by showing the archaeal RPB7 ortholog promotes open-complex (bubble) formation and stimulates core polymerase activity.","evidence":"Reconstituted archaeal RNA polymerase, permanganate footprinting and in vitro transcription","pmids":["17311916"],"confidence":"High","gaps":["Generalization to eukaryotic Pol II not directly tested","Mechanistic link between RNA binding and bubble formation unresolved"]},{"year":2008,"claim":"Revised the initiation-only view by showing RPB7 remains with polymerase through early elongation and extends to late elongation under stress, indicating a role in stabilizing transcribing polymerase.","evidence":"TAP-ChIP with qPCR across transcribed regions in human cells","pmids":["17848138"],"confidence":"Medium","gaps":["Single-lab ChIP without orthogonal occupancy method","Stress-induced retention mechanism undefined"]},{"year":2013,"claim":"Defined the elongation stage at which RPB7 leaves polymerase, showing dissociation coincides with Ser2-CTD phosphorylation and recruitment of elongation factors.","evidence":"Quantitative mass spectrometry with Rpb7-tagged Pol II immunoprecipitation","pmids":["23418395"],"confidence":"Medium","gaps":["Trigger for dissociation not causally established","Reciprocity with reassociation during reinitiation untested here"]},{"year":2023,"claim":"Assigned RPB7 a repressive role in transcription-coupled nucleotide excision repair through distinct domain contacts, partly via the Spt4/Spt5 pathway and partly independently.","evidence":"Genetic epistasis with double mutants, TCR and UV-sensitivity assays in S. cerevisiae","pmids":["37380080"],"confidence":"Medium","gaps":["Molecular mechanism of repression not biochemically defined","Relevance to human TCR not tested"]},{"year":2025,"claim":"Established RPB7 as a stabilizer of the largest subunit RPB1, acting through CTDP1 recruitment and antagonism of Cullin-3 ubiquitylation, and required for reinitiation and RNA processing.","evidence":"RPB7 depletion, co-IP, Cullin-3 ubiquitylation assay, reinitiation and RNA-processing assays in human cells","pmids":["40038320"],"confidence":"Medium","gaps":["Not independently replicated","Direct CTDP1-RPB1 dephosphorylation step not reconstituted","Whether stabilization is constitutive or signal-regulated unknown"]},{"year":null,"claim":"How RPB7's RNA exit-channel positioning, elongation-stage dissociation/reassociation, and its roles in RPB1 stabilization and RNA processing are integrated into a single regulated transcription cycle remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking RNA contact, CTD-phosphorylation-dependent dissociation, and reinitiation","Signal-dependent control of RPB7 abundance versus assembly not integrated"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[7,12,13]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[7]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0,3,4]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[4,7,17]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[9,18]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[4,7,17,18]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[10,22]},{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[21]}],"complexes":["RNA polymerase II","RPB4/RPB7 subcomplex"],"partners":["POLR2D","POLR2A","POLR2F","CTDP1","VHL","EWSR1","NRD1","RARA"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P62487","full_name":"DNA-directed RNA polymerase II subunit RPB7","aliases":["DNA-directed RNA polymerase II subunit G","RNA polymerase II 19 kDa subunit","RPB19"],"length_aa":172,"mass_kda":19.3,"function":"Core component of RNA polymerase II (Pol II), a DNA-dependent RNA polymerase which synthesizes mRNA precursors and many functional non-coding RNAs using the four ribonucleoside triphosphates as substrates. Pol II is the central component of the basal RNA polymerase II transcription machinery. It is composed of mobile elements that move relative to each other. POLR2G/RPB7 is part of a subcomplex with POLR2D/RPB4 that binds to a pocket formed by POLR2A/RPB1, POLR2B/RPB2 and POLR2F/RPABC2 at the base of the clamp element. The POLR2D/RPB4-POLR2G/RPB7 subcomplex seems to lock the clamp via POLR2G/RPB7 in the closed conformation thus preventing double-stranded DNA to enter the active site cleft. The POLR2D/RPB4-POLR2G/RPB7 subcomplex binds single-stranded DNA and RNA","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/P62487/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/POLR2G","classification":"Common Essential","n_dependent_lines":1197,"n_total_lines":1208,"dependency_fraction":0.9908940397350994},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000168002","cell_line_id":"CID000702","localizations":[{"compartment":"nucleoplasm","grade":3}],"interactors":[{"gene":"POLR2A","stoichiometry":10.0},{"gene":"POLR2B","stoichiometry":10.0},{"gene":"POLR2C","stoichiometry":10.0},{"gene":"POLR2D","stoichiometry":10.0},{"gene":"POLR2E","stoichiometry":10.0},{"gene":"GTF2B","stoichiometry":10.0},{"gene":"POLR2K","stoichiometry":10.0},{"gene":"GTF2F1","stoichiometry":4.0},{"gene":"POLR2F","stoichiometry":4.0},{"gene":"POLR2H","stoichiometry":4.0}],"url":"https://opencell.sf.czbiohub.org/target/CID000702","total_profiled":1310},"omim":[{"mim_id":"619801","title":"RNA POLYMERASE III, SUBUNIT H; POLR3H","url":"https://www.omim.org/entry/619801"},{"mim_id":"609881","title":"RNA POLYMERASE II, SUBUNIT J2; POLR2J2","url":"https://www.omim.org/entry/609881"},{"mim_id":"606017","title":"POLYMERASE II, RNA, SUBUNIT D; POLR2D","url":"https://www.omim.org/entry/606017"},{"mim_id":"602013","title":"POLYMERASE II, RNA, SUBUNIT G; POLR2G","url":"https://www.omim.org/entry/602013"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/POLR2G"},"hgnc":{"alias_symbol":["hRPB19","hsRPB7","RPB7"],"prev_symbol":[]},"alphafold":{"accession":"P62487","domains":[{"cath_id":"3.30.1490.120","chopping":"1-76","consensus_level":"high","plddt":95.6337,"start":1,"end":76},{"cath_id":"2.40.50.140","chopping":"82-169","consensus_level":"high","plddt":95.401,"start":82,"end":169}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P62487","model_url":"https://alphafold.ebi.ac.uk/files/AF-P62487-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P62487-F1-predicted_aligned_error_v6.png","plddt_mean":95.62},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=POLR2G","jax_strain_url":"https://www.jax.org/strain/search?query=POLR2G"},"sequence":{"accession":"P62487","fasta_url":"https://rest.uniprot.org/uniprotkb/P62487.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P62487/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P62487"}},"corpus_meta":[{"pmid":"16204182","id":"PMC_16204182","title":"RNA 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RPB7 association with Pol II is dependent on RPB4, as polymerase purified from rpb4Δ cells also lacks RPB7.\",\n      \"method\": \"Gene deletion, yeast genetics, polymerase purification\",\n      \"journal\": \"Yeast\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean genetic knockouts with defined viability phenotypes, replicated in multiple follow-up studies\",\n      \"pmids\": [\"8488730\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"Human hsRPB7 is a functional ortholog of yeast RPB7: it assembles into a complete 12-subunit yeast Pol II complex (shown by immunoprecipitation) and rescues the essential RPB7 deletion at moderate temperatures. However, hsRPB7 shows reduced affinity for yeast RPB4 compared to yeast RPB7, explaining failure to fully complement at temperature extremes.\",\n      \"method\": \"Complementation assay, immunoprecipitation of Pol II from yeast expressing hsRPB7, yeast two-hybrid\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — complementation, immunoprecipitation, and two-hybrid results are mutually consistent and replicated by later studies\",\n      \"pmids\": [\"7579693\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Oncogenic EWS-Fli1 specifically interacts with hsRPB7 (the human Pol II seventh subunit) via the EWS amino terminus; this interaction was identified by yeast two-hybrid and confirmed by in vitro immunoprecipitation. hsRPB7 copurifies with EWS-Fli1 but not with wild-type Fli1 from nuclear extracts. Overexpression of hsRPB7 specifically enhances gene activation by EWS-chimeric transcription factors.\",\n      \"method\": \"Yeast two-hybrid, in vitro immunoprecipitation, nuclear extract copurification, overexpression transactivation assay\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — yeast two-hybrid confirmed by reciprocal co-IP and nuclear extract copurification plus functional overexpression assay in single study\",\n      \"pmids\": [\"9704926\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Human hsRPB4 strongly and specifically interacts with hsRPB7 in yeast and mammalian cells and copurifies with intact Pol II. Structure-function mapping defined the interaction interface between hsRPB4 and hsRPB7. hsRPB4 does not interact with yeast RPB7, showing species-specific divergence while retaining partial complementation of rpb4− yeast phenotypes.\",\n      \"method\": \"Yeast two-hybrid, coimmunoprecipitation in mammalian cells, polymerase copurification, truncation/deletion mapping\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP in two cell systems plus copurification with Pol II and domain mapping\",\n      \"pmids\": [\"9528765\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Cryo-EM difference mapping showed that Rpb4/Rpb7 form part of the floor of the DNA-binding cleft of yeast Pol II. Surface plasmon resonance demonstrated that Rpb4/Rpb7 stabilize a minimal pre-initiation complex containing promoter DNA, TBP, TFIIB and polymerase, suggesting a role in coupling DNA entry into the active center cleft to cleft closure.\",\n      \"method\": \"Cryo-EM 3D reconstruction, difference mapping, surface plasmon resonance\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure with SPR functional validation in a single rigorous study\",\n      \"pmids\": [\"9545247\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Ligand-free RARα interacts with hsRPB7 through its DNA-binding domain (in the absence of retinoic acid); RA addition disrupts the interaction. Overexpressed hRARα (no RA) represses Pol II-dependent activators (AP-1, GR), and this repression is relieved by co-transfected hsRPB7, indicating that unliganded RARα sequesters hsRPB7 to repress transcription.\",\n      \"method\": \"Yeast two-hybrid, truncation analysis, transient transfection repression/rescue assays\",\n      \"journal\": \"Endocrine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — two-hybrid plus functional rescue assay, single lab, no reciprocal co-IP\",\n      \"pmids\": [\"10484292\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Rpb7 can associate with Pol II lacking Rpb4 (Pol IIΔ4) when overexpressed, demonstrated by two reciprocal coimmunoprecipitation experiments. This Rpb4-independent interaction supports transcription under mild stress, establishing that a major role of Rpb4 is to augment the affinity of Rpb7 for Pol II.\",\n      \"method\": \"Suppressor screen, reciprocal coimmunoprecipitation, transcription assays in rpb4Δ cells\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP with functional (transcription) readout and genetic suppressor data converging on same conclusion\",\n      \"pmids\": [\"10082533\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"The purified Rpb4/Rpb7 complex binds single-stranded DNA and RNA via an OB-fold motif in Rpb7. A small deletion in the OB-fold abolished nucleic acid binding without disrupting the Rpb4/Rpb7 complex or its association with Pol II, yet destroyed transcription activity. A separate OB-fold deletion blocked transcription but not nucleic acid binding, indicating the OB-fold mediates both nucleic acid binding and protein-protein interactions required for initiation. Template competition showed Rpb4/Rpb7 is not needed for stable polymerase recruitment but acts at a post-recruitment step.\",\n      \"method\": \"Purified complex binding assays (ssDNA/RNA), deletion mutagenesis, reconstituted transcription assay, template competition\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution with mutagenesis and multiple orthogonal functional assays in one study\",\n      \"pmids\": [\"11087726\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Crystal structure of the archaeal RPB7/RPB4 homologs (Methanococcus jannaschii subunits E and F) was determined, revealing an elongated two-domain structure for subunit E with two potential RNA-binding motifs, and subunit F wrapping around subunit E at the domain interface. A model was proposed in which the RNA-binding face of RPB7 contacts the nascent RNA transcript.\",\n      \"method\": \"X-ray crystallography\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure of archaeal ortholog complex, foundational structural study widely replicated\",\n      \"pmids\": [\"11741548\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"pVHL directly binds to hsRPB7 through its beta-domain; naturally occurring beta-domain mutations decrease this interaction. Introduction of wild-type pVHL into VHL-mutant renal carcinoma cells promotes ubiquitylation and proteasomal degradation of hsRPB7, reduces its nuclear accumulation, and suppresses hsRPB7-induced VEGF promoter transactivation and VEGF secretion.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, ubiquitylation assay, proteasome inhibitor experiments, luciferase reporter assay, ELISA\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct binding confirmed by co-IP, ubiquitylation demonstrated biochemically, functional consequence shown by reporter and VEGF secretion assays; multiple orthogonal methods\",\n      \"pmids\": [\"12912922\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"In fission yeast, Rpb7 directly interacts in vitro with Seb1 (the S. pombe homolog of Nrd1, an RNA-binding protein for 3′-end formation of snRNA/snoRNA). Two-hybrid and in vitro binding identified Glu166/Asp167 near the Rpb7 C-terminus as critical for the interaction. S. cerevisiae Rpb7 also interacts with Nrd1, showing evolutionary conservation. This suggests Rpb7 anchors an RNA-processing factor to the Pol II apparatus.\",\n      \"method\": \"Two-hybrid screen, in vitro direct binding, site-directed mutagenesis\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro direct binding plus two-hybrid with mutagenesis, single lab\",\n      \"pmids\": [\"12907709\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"A conditional mutation in the shared Rpb6 subunit (Q100R) causes selective loss of Rpb4 and Rpb7 from purified Pol II. Interaction experiments support a direct association between Rpb6 and Rpb4, identifying Rpb6 as one contact point through which the Rpb4/Rpb7 subcomplex associates with the Pol II core.\",\n      \"method\": \"Genetic screen, polymerase purification, interaction experiments (pulldown/co-IP)\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — purification of mutant polymerase plus direct interaction experiments, single lab\",\n      \"pmids\": [\"12697831\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"UV cross-linking of uridine analogs in the first 6 nt of nascent RNA demonstrates that the emerging transcript contacts Rpb7 in Pol II elongation complexes carrying 26–32 nt transcripts; this interaction is greatly reduced at 41–43 nt and absent at 125 nt, establishing groove 1 (pointing toward Rpb4/Rpb7) as the nascent RNA exit path.\",\n      \"method\": \"UV cross-linking with uridine analogs on reconstituted elongation complexes\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstituted complexes with defined transcript lengths and site-specific cross-linking; directly identifies RNA-Rpb7 contact\",\n      \"pmids\": [\"16327806\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Crystal structure of the human Rpb4/Rpb7 heterodimer was determined at 2.7 Å. Site-directed mutagenesis of conserved surface residues in Rpb7 (and archaeal subunit E), combined with EMSA RNA-binding assays, identified an elongated surface spanning the OB-fold B4–B5 loop and extending to the N-terminal domain as the RNA-binding interface.\",\n      \"method\": \"X-ray crystallography, site-directed mutagenesis, EMSA\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure at 2.7 Å with mutagenesis-validated RNA-binding surface, multiple orthogonal methods\",\n      \"pmids\": [\"16282592\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Fission yeast Rpb7 has a specific role in transcription of centromeric pre-siRNA precursors required for RNAi-directed heterochromatin formation. The rpb7-G150D point mutation specifically impairs centromeric pre-siRNA transcription (and thus RNAi silencing) without equivalent effects seen with other Pol II subunit mutants.\",\n      \"method\": \"Point mutation analysis, centromeric transcription assays, RNAi/heterochromatin reporter assays\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — defined point mutation with specific transcriptional and silencing phenotypes, single lab\",\n      \"pmids\": [\"16204182\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"In fission yeast, Rpb7 interacts with both GAPDH and actin. GAPDH was identified as an Rpb7-binding protein by two-hybrid screen and confirmed by affinity purification from S. pombe extract using an Rpb4/Rpb7-coupled column. Actin was identified as a Pol II-associated protein and shown to interact with Rpb7.\",\n      \"method\": \"Two-hybrid screen, affinity purification (Rpb4/Rpb7-coupled column pull-down)\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, two-hybrid plus single affinity-pull-down without reciprocal co-IP\",\n      \"pmids\": [\"15620689\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Mutations in the N-terminal RNP-like domain of S. cerevisiae Rpb7 impair stress responses (high-temperature growth, sporulation) and increase Rpb7's dependence on Rpb4 for Pol II interaction. RNA polymerase pulldown assays revealed two crucial interaction points between the Rpb4/Rpb7 subcomplex and the core: the N-terminal RNP-like domain of Rpb7 and the partially ordered N-terminus of Rpb4 (contacting Rpb2).\",\n      \"method\": \"Mutagenesis, RNA polymerase pulldown assay, complementation analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo pulldown with defined mutants identifying two interaction points, single lab\",\n      \"pmids\": [\"18056993\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"In Pyrococcus furiosus, the archaeal RPB7 ortholog E' (subunit E') stimulates Pol II core enzyme activity at low temperatures and promotes open complex formation (bubble formation at multiple positions in the transcription bubble), as shown by permanganate footprinting of reconstituted transcription complexes. Subunit F did not significantly contribute to catalytic activity.\",\n      \"method\": \"Reconstitution of archaeal RNA polymerase, permanganate footprinting, in vitro transcription at varying temperatures\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — fully reconstituted system with permanganate footprinting and kinetic analyses; rigorous in vitro mechanistic dissection\",\n      \"pmids\": [\"17311916\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"ChIP experiments in live human cells show that Rpb7 remains associated with Pol II during early elongation (not solely at initiation as previously believed), and under stress conditions Rpb7 association extends to late elongation stages, suggesting a role in stabilizing transcribing polymerase throughout the transcription cycle.\",\n      \"method\": \"TAP-ChIP (chromatin immunoprecipitation) with quantitative PCR across promoter-proximal and downstream transcribed regions in human cells\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP in living human cells with multiple gene loci and stress conditions, single lab\",\n      \"pmids\": [\"17848138\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Pull-down assays confirmed direct in vitro interaction between the first 57 residues of the EWS N-terminal activation domain (EAD) and hsRPB7, forming a stable complex.\",\n      \"method\": \"In vitro pull-down assay with recombinant proteins\",\n      \"journal\": \"Molecular biology reports\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, single in vitro pull-down method only\",\n      \"pmids\": [\"18607770\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Quantitative proteomics demonstrated that Rpb4/Rpb7 dissociate from Pol II upon interaction with transcriptional elongation-associated proteins recruited to the hyperphosphorylated CTD (Ser2-P form). Pol II isolated through Rpb7 is depleted in Ser2-CTD phosphorylation, indicating Rpb4/7 are dispensable during specific elongation stages.\",\n      \"method\": \"Quantitative proteomics (mass spectrometry), immunoprecipitation via Rpb7-tagged Pol II\",\n      \"journal\": \"Molecular & cellular proteomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — quantitative MS with orthogonal co-IP approach, single lab\",\n      \"pmids\": [\"23418395\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Rpb7 functions as a repressor of transcription-coupled nucleotide excision repair (TCR) in S. cerevisiae. The Rpb7 region interacting with the KOW3 domain of Spt5 represses TCR through a common mechanism with Spt4/Spt5. The Rpb7 regions contacting Rpb4 and/or core RNAPII repress TCR largely independently of Spt4/Spt5, as shown by synergistic enhancement of TCR derepression when combined with spt4Δ.\",\n      \"method\": \"Genetic epistasis (double mutants), TCR assays at specific gene loci, UV sensitivity assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with defined domain mutations and multiple gene loci, single lab\",\n      \"pmids\": [\"37380080\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"RPB7 stabilizes RPB1 (the largest Pol II subunit): depletion of RPB7 destabilizes RPB1 via the E3 ubiquitin ligase Cullin 3. Stabilization depends on the loop regions of RPB7, CDK9, the CTD and linker region of RPB1. RPB7 also interacts with the phosphatase CTDP1, which is required for RPB1 stability. RPB7 is essential for Pol II reinitiation, engages RNA-processing factors, and localizes to the RNA exit channel; its absence compromises RNA processing.\",\n      \"method\": \"RPB7 depletion experiments, co-immunoprecipitation (RPB7-CTDP1 interaction), ubiquitylation assay (Cullin 3), reinitiation assays, RNA processing assays, localization data\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (depletion, co-IP, ubiquitylation, functional assays) in single study, not yet independently replicated\",\n      \"pmids\": [\"40038320\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Mapping of the Rpb4–Rpb7 interaction interface by two-hybrid analysis with truncation mutants showed that both the amino- and carboxy-terminal domains of Rpb7 are required for interaction with Rpb4, and that N-terminal deletions of Rpb4 (beyond 49 residues) progressively abolish interaction with Rpb7.\",\n      \"method\": \"Yeast two-hybrid with truncation mutants, computational structural analysis\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — two-hybrid only, single lab, no in vitro validation\",\n      \"pmids\": [\"15913559\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"POLR2G (RPB7/hsRPB7) encodes the seventh subunit of RNA polymerase II that forms an obligate heterodimer with RPB4; this subcomplex protrudes from the Pol II core at the RNA exit groove (groove 1), contacts the nascent transcript through an OB-fold RNA-binding domain, stabilizes the RPB1 largest subunit via recruitment of the CTDP1 phosphatase (countering Cullin 3-mediated ubiquitylation), mediates a post-recruitment step in transcription initiation (open-complex formation and/or clamp closure), remains associated with elongating polymerase through early elongation and dissociates during late elongation, represses transcription-coupled nucleotide excision repair, couples RNA processing to transcription, and serves as a direct binding target/effector for oncoproteins (EWS-Fli1), tumor suppressors (pVHL, which ubiquitylates it for proteasomal degradation), and nuclear receptors (unliganded RARα), thereby linking core transcription machinery to signaling and post-transcriptional gene regulation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"POLR2G (RPB7/hsRPB7) encodes the essential seventh subunit of RNA polymerase II, which forms an obligate heterodimer with RPB4 that contacts the Pol II core and is required for cell viability [#0, #3]. The subcomplex sits in the floor of the DNA-binding cleft and stabilizes a minimal pre-initiation complex of promoter DNA, TBP, TFIIB and polymerase, acting at a post-recruitment step rather than in stable polymerase recruitment; RPB7's OB-fold mediates both nucleic-acid binding and the protein-protein contacts required for initiation, and the subunit promotes open-complex formation [#4, #7, #17]. RPB7 lines the RNA exit path (groove 1): the nascent transcript cross-links to RPB7 during early elongation and the contact is lost as the transcript lengthens, consistent with RPB7 marking the RNA exit channel [#12, #13]. RPB7 stays associated with elongating polymerase through early elongation and dissociates upon recruitment of Ser2-phosphorylated CTD elongation factors during late elongation [#18, #20]. Through these contacts RPB7 couples transcription to RNA processing by anchoring 3'-end/processing factors such as Nrd1/Seb1 to the polymerase [#10, #22], and it stabilizes the largest subunit RPB1 by recruiting the CTDP1 phosphatase and countering Cullin-3-mediated ubiquitylation [#22]. Beyond core transcription, RPB7 is a binding target for the oncogenic EWS-Fli1 fusion, whose EWS N-terminus contacts RPB7 to enhance EWS-chimera-driven transactivation [#2], and for the tumor suppressor pVHL, which binds RPB7 through its beta-domain and targets it for ubiquitylation and proteasomal degradation, thereby limiting RPB7-driven VEGF promoter activation [#9]. RPB7 also represses transcription-coupled nucleotide excision repair via domain contacts overlapping with the Spt4/Spt5 pathway [#21].\",\n  \"teleology\": [\n    {\n      \"year\": 1993,\n      \"claim\": \"Established that RPB7 is an essential, RPB4-dependent component of Pol II, distinguishing it from the dispensable RPB4 and defining the architecture of the subcomplex's attachment to the core.\",\n      \"evidence\": \"Gene deletion and polymerase purification in S. cerevisiae\",\n      \"pmids\": [\"8488730\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the molecular contacts anchoring RPB7 to the core\", \"Did not establish the biochemical function of RPB7\"]\n    },\n    {\n      \"year\": 1995,\n      \"claim\": \"Showed functional conservation by demonstrating the human ortholog assembles into and rescues yeast Pol II, validating yeast as a model for human RPB7 mechanism.\",\n      \"evidence\": \"Complementation, immunoprecipitation and two-hybrid in yeast expressing hsRPB7\",\n      \"pmids\": [\"7579693\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Reduced affinity for yeast RPB4 left interface determinants undefined\", \"No human-cell functional assay\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Mapped the obligate RPB4–RPB7 heterodimer interface in human cells and placed the subcomplex in the floor of the DNA-binding cleft, where it stabilizes the pre-initiation complex.\",\n      \"evidence\": \"Reciprocal co-IP and domain mapping in mammalian cells; cryo-EM difference mapping and SPR in yeast\",\n      \"pmids\": [\"9528765\", \"9545247\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism linking cleft positioning to clamp closure inferred not proven\", \"Atomic-resolution interface not yet resolved at this stage\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Identified RPB7 as a direct target of the oncogenic EWS-Fli1 fusion, linking the core transcription subunit to oncogenic transactivation.\",\n      \"evidence\": \"Yeast two-hybrid, in vitro co-IP, nuclear-extract copurification and overexpression transactivation assay\",\n      \"pmids\": [\"9704926\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Endogenous relevance in Ewing sarcoma cells not established\", \"Mechanism by which RPB7 enhances chimera activity unresolved\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Defined RPB7's OB-fold as the dual nucleic-acid-binding and initiation-competent element and showed the subcomplex acts at a post-recruitment step, not in stable polymerase loading.\",\n      \"evidence\": \"Purified-complex binding assays, OB-fold deletion mutagenesis, reconstituted transcription and template competition\",\n      \"pmids\": [\"11087726\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Precise post-recruitment step (open complex vs clamp closure) not separated\", \"In vivo confirmation of OB-fold requirement absent\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Provided the first atomic view of the RPB7/RPB4 fold from the archaeal homolog, predicting an RNA-binding face that contacts the nascent transcript.\",\n      \"evidence\": \"X-ray crystallography of Methanococcus jannaschii subunits E and F\",\n      \"pmids\": [\"11741548\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"RNA contact was modeled, not observed\", \"Archaeal divergence from eukaryotic Pol II left untested\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Connected RPB7 to tumor-suppressor regulation by showing pVHL binds and targets RPB7 for ubiquitin-proteasome degradation, controlling RPB7-driven VEGF activation.\",\n      \"evidence\": \"Two-hybrid, co-IP, ubiquitylation and proteasome-inhibitor assays, luciferase reporter and ELISA in renal carcinoma cells\",\n      \"pmids\": [\"12912922\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether degradation affects assembled Pol II or only free RPB7 unclear\", \"Direct E3 ligase chemistry not reconstituted\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Implicated RPB7 in coupling transcription to RNA processing by identifying a conserved interaction with the 3'-end formation factor Nrd1/Seb1, and identified RPB6 as a core attachment point for the subcomplex.\",\n      \"evidence\": \"Two-hybrid, in vitro direct binding and site-directed mutagenesis (Seb1/Nrd1); conditional Rpb6 mutant and interaction assays\",\n      \"pmids\": [\"12907709\", \"12697831\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No reciprocal co-IP for the Nrd1 interaction in mammalian cells\", \"Functional consequence for processing not directly measured here\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Localized the nascent RNA exit path to RPB7 and resolved the human heterodimer crystal structure with a mutagenesis-validated RNA-binding surface.\",\n      \"evidence\": \"Site-specific UV cross-linking on reconstituted elongation complexes; 2.7 Å human Rpb4/Rpb7 crystal structure with mutagenesis and EMSA\",\n      \"pmids\": [\"16327806\", \"16282592\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional role of the transient RNA contact in elongation not defined\", \"Dynamics of contact loss at longer transcripts not mechanistically explained\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Demonstrated a catalytic-stimulatory role by showing the archaeal RPB7 ortholog promotes open-complex (bubble) formation and stimulates core polymerase activity.\",\n      \"evidence\": \"Reconstituted archaeal RNA polymerase, permanganate footprinting and in vitro transcription\",\n      \"pmids\": [\"17311916\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Generalization to eukaryotic Pol II not directly tested\", \"Mechanistic link between RNA binding and bubble formation unresolved\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Revised the initiation-only view by showing RPB7 remains with polymerase through early elongation and extends to late elongation under stress, indicating a role in stabilizing transcribing polymerase.\",\n      \"evidence\": \"TAP-ChIP with qPCR across transcribed regions in human cells\",\n      \"pmids\": [\"17848138\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab ChIP without orthogonal occupancy method\", \"Stress-induced retention mechanism undefined\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Defined the elongation stage at which RPB7 leaves polymerase, showing dissociation coincides with Ser2-CTD phosphorylation and recruitment of elongation factors.\",\n      \"evidence\": \"Quantitative mass spectrometry with Rpb7-tagged Pol II immunoprecipitation\",\n      \"pmids\": [\"23418395\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Trigger for dissociation not causally established\", \"Reciprocity with reassociation during reinitiation untested here\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Assigned RPB7 a repressive role in transcription-coupled nucleotide excision repair through distinct domain contacts, partly via the Spt4/Spt5 pathway and partly independently.\",\n      \"evidence\": \"Genetic epistasis with double mutants, TCR and UV-sensitivity assays in S. cerevisiae\",\n      \"pmids\": [\"37380080\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular mechanism of repression not biochemically defined\", \"Relevance to human TCR not tested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Established RPB7 as a stabilizer of the largest subunit RPB1, acting through CTDP1 recruitment and antagonism of Cullin-3 ubiquitylation, and required for reinitiation and RNA processing.\",\n      \"evidence\": \"RPB7 depletion, co-IP, Cullin-3 ubiquitylation assay, reinitiation and RNA-processing assays in human cells\",\n      \"pmids\": [\"40038320\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Not independently replicated\", \"Direct CTDP1-RPB1 dephosphorylation step not reconstituted\", \"Whether stabilization is constitutive or signal-regulated unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How RPB7's RNA exit-channel positioning, elongation-stage dissociation/reassociation, and its roles in RPB1 stabilization and RNA processing are integrated into a single regulated transcription cycle remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking RNA contact, CTD-phosphorylation-dependent dissociation, and reinitiation\", \"Signal-dependent control of RPB7 abundance versus assembly not integrated\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [7, 12, 13]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 3, 4]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [4, 7, 17]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [9, 18]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [4, 7, 17, 18]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [10, 22]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [21]}\n    ],\n    \"complexes\": [\n      \"RNA polymerase II\",\n      \"RPB4/RPB7 subcomplex\"\n    ],\n    \"partners\": [\n      \"POLR2D\",\n      \"POLR2A\",\n      \"POLR2F\",\n      \"CTDP1\",\n      \"VHL\",\n      \"EWSR1\",\n      \"NRD1\",\n      \"RARA\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}