{"gene":"POLR2B","run_date":"2026-06-10T06:43:35","timeline":{"discoveries":[{"year":2000,"finding":"Ssu72 physically interacts with purified RNA Pol II (containing Rpb2 subunit) as shown by co-immunoprecipitation, and a suppressor allele rpb2-100 (R512C in homology block D) was identified that genetically interacts with ssu72-2, defining Ssu72 as a factor that physically and functionally interacts with the RNAP II core machinery during transcription initiation.","method":"Genetic suppressor screen, co-immunoprecipitation with purified RNAP II, in vivo transcription assays","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal genetic and biochemical evidence from a single lab using two orthogonal methods (suppressor genetics + co-IP)","pmids":["11046131"],"is_preprint":false},{"year":2003,"finding":"Mutations in the RPB2 subunit of RNA Pol II (along with SPT5 and TFIIS) cause increased utilization of internal and upstream poly(A) sites in yeast, indicating that transcriptional elongation defects leading to pausing or arrest can be visualized in vivo through poly(A) site choice, and that RPB2 plays a role in transcriptional elongation efficiency.","method":"In vivo gene expression analysis using poly(A) site reporter genes in yeast rpb2 mutants","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo functional assay with rpb2 mutants and multiple gene targets, single lab","pmids":["14560031"],"is_preprint":false},{"year":2004,"finding":"A functional interaction between the B-finger domain of TFIIB and the lobe domain of Rpb2 (G369S substitution) was identified through genetic suppressor analysis. The Rpb2 lobe-jaw region defines a novel role for Rpb2 in transcription start site selection, and the sua7-3 rpb2-101 double mutant was sensitive to 6-azauracil in vivo and to NTP substrate depletion in vitro, linking Rpb2 lobe domain to elongation as well.","method":"Genetic suppressor screen, in vitro promoter-specific run-on transcription, abortive initiation assay, 6-azauracil sensitivity","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis plus multiple in vitro transcription assays in single lab","pmids":["15082791"],"is_preprint":false},{"year":2005,"finding":"The highly conserved Rpb2 residue E791 (human RNAP II; E836 in yeast) is involved in NTP and Mg(B) binding at the active center. The E791A substitution mutant showed impaired transcription activity at low NTP concentrations both in vitro and in vivo, and decreased NTP polymerization and transcript cleavage activities at low Mg concentrations, implicating this residue in NTP-Mg(B) loading into the active site.","method":"Affinity purification of human RNAP II E791A mutant, in vitro transcription assay, in vivo transcription assay, transcript cleavage assay at varying Mg and NTP concentrations","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — active-site mutagenesis with both in vitro enzymatic assays and in vivo validation in human RNAP II","pmids":["15886393"],"is_preprint":false},{"year":2011,"finding":"Deletion of the RPB2 flap loop (Δ873-884) in human RNAP II, which removes the TFIIB interaction interface, had no detectable effect on global transcription initiation, RNAP II occupancy, promoter escape, productive elongation, promoter binding, abortive initiation, TFIIS-stimulated transcript cleavage, or NELF/DSIF inhibition in vitro. A modest effect on elongation and pausing was suppressed by TFIIF.","method":"Genome-wide ChIP-seq in HEK293 cells expressing flap loop deletion mutant, in vitro transcription and abortive initiation assays, TFIIS cleavage assay, NELF/DSIF pausing assay","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — mutagenesis with multiple orthogonal in vitro and genome-wide in vivo assays; negative result rigorously established","pmids":["21670157"],"is_preprint":false},{"year":1998,"finding":"Two-hybrid analysis using S. pombe RNAP II subunits mapped an Rpb3 contact site within the conserved region H of Rpb2 (the beta-subunit homology region conserved among all RNA polymerases), establishing a direct physical interaction between Rpb2 and Rpb3 within the polymerase complex.","method":"Yeast two-hybrid assay with fission yeast Rpb2 fragments","journal":"Molecular & general genetics","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single two-hybrid mapping study, no orthogonal validation","pmids":["9738888"],"is_preprint":false},{"year":2001,"finding":"Two-hybrid mapping in S. pombe identified the Rpb2-Rpb3 interaction site at the C-terminal region of Rpb2, within amino acids 902–989 (encoded by base 2701–2966 of Rpb2 cDNA).","method":"Yeast two-hybrid assay with Rpb2 fragment constructs","journal":"Wei sheng wu xue bao (Acta Microbiologica Sinica)","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single two-hybrid mapping, single lab, no orthogonal validation","pmids":["12552808"],"is_preprint":false},{"year":2013,"finding":"UV damage regulates alternative polyadenylation of the RPB2 gene in yeast: in normally growing cells, the proximal poly(A) site is preferentially used; after UV damage and transcription recovery, the distal poly(A) site is preferentially used. The 3'UTR of RPB2 is sufficient for this regulation, and the rate of transcription elongation (not initiation) is an important determinant of poly(A) site choice.","method":"RNA analysis of poly(A) site usage before and after UV damage, 3'UTR reporter constructs, mRNA stability assays","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple experimental approaches (reporter constructs, stability assays, transcription rate manipulation) from single lab","pmids":["23355614"],"is_preprint":false},{"year":2022,"finding":"Rtr1 is directly required for assembly of the two largest RNAP II subunits (Rpb1 and Rpb2). Deletion of RTR1 leads to cytoplasmic clumping of RNAP II subunits. Rtr1 coordinates with Gpn3 and Npa3 assembly factors; overexpression of RTR1 suppresses cytoplasmic clump formation of RNAP II subunit in a gpn3-9 mutant. This function does not require Rtr1 phosphatase catalytic activity.","method":"Genetic suppressor analysis, co-immunoprecipitation, fluorescence microscopy of RNAP II subunit localization, analysis of catalytically inactive Rtr1 mutant","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis combined with localization microscopy and biochemical co-IP, single lab","pmids":["36190433"],"is_preprint":false},{"year":2024,"finding":"TANGO6 associates with COPI vesicles and captures RPB2 in the cis-Golgi during G1 phase, then carries RPB2 to the ER and subsequently to the nucleus. Functional disruption of TANGO6 causes cytoplasmic accumulation of RPB2 and G1 cell cycle arrest. Conditional depletion or overexpression of TANGO6 in mouse hematopoietic stem cells compromises or expands hematopoiesis.","method":"Co-immunoprecipitation, live cell fluorescence imaging, cell cycle analysis (PI staining), conditional knockout/overexpression in mouse hematopoietic stem cells","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP, live imaging of RPB2 trafficking, functional rescue experiments; single lab but multiple orthogonal methods","pmids":["38490996"],"is_preprint":false},{"year":2023,"finding":"Knockdown of POLR2B (RPB2) by shRNA suppresses GBM tumor cell growth in vitro and in vivo in a xenograft model, and RNA sequencing identified DDIT4 (DNA damage-inducible transcript 4) as a downstream transcriptional target regulated by RPB2 in glioblastoma cells.","method":"shRNA knockdown, cell proliferation assay, xenograft mouse model, RNA sequencing, GO and GSEA analysis","journal":"Biochemical and biophysical research communications","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, knockdown phenotype with downstream target identification but limited mechanistic pathway resolution","pmids":["37423037"],"is_preprint":false},{"year":2025,"finding":"A gain-of-function mutation in the Rpb2 subunit of Pol II (rpb2-N44Y in S. pombe) reduces RNAi-dependent heterochromatin. The heterochromatin defects of rpb2-N44Y require Elongator Protein 1 (Elp1); loss of Elp1 (but not other Elongator subunits such as Elp3) suppresses these defects independently of the mcm5s2U34 tRNA modification, revealing an Elp1 chromatin function downstream of or parallel to Rpb2.","method":"CRISPR site-directed mutagenesis, genetic epistasis with elp1/elp3 deletions, siRNA quantification, heterochromatin reporter assays in S. pombe","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR mutagenesis with genetic epistasis and multiple molecular readouts; preprint, single lab","pmids":[],"is_preprint":true},{"year":2025,"finding":"Cryo-EM mapping showed that IWS1 short linear motifs (SLiMs) in its intrinsically disordered C-terminal region directly contact the RPB2 lobe domain of Pol II. Functional assays demonstrated that IWS1 interactions with the RPB2 lobe and ELOF1 are required for IWS1-dependent transcription elongation stimulation, while IWS1 recruitment depends on the RPB1 jaw domain and downstream DNA binding.","method":"Cryo-electron microscopy structure determination, deletion mutagenesis of SLiMs, in vitro transcription elongation assays","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure plus functional mutagenesis assays; preprint, single lab","pmids":[],"is_preprint":true},{"year":2025,"finding":"In yeast, Prp40 (not U1-70K) is the predominant mediator of U1 snRNP interaction with Pol II; multiple domains of Prp40 interact with Pol II independently of the CTD. In humans, the cryo-EM structure showed that U1-70K RRM domain directly interacts with the RPB2 subunit of Pol II, mediating co-transcriptional splicing coupling.","method":"Co-immunoprecipitation in yeast, cryo-EM structure of human U1 snRNP–Pol II complex, domain deletion analysis","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 / Weak — preprint; the RPB2-specific finding in humans is based on a referenced prior cryo-EM structure, with primary new data in yeast using co-IP","pmids":[],"is_preprint":true}],"current_model":"POLR2B (RPB2) encodes the second-largest subunit of RNA polymerase II; it contributes to the catalytic active site (E791 coordinates NTP-Mg(B) loading), participates in transcription start site selection via its lobe domain (interacting functionally with TFIIB B-finger), influences transcriptional elongation efficiency (with rpb2 mutations causing increased upstream poly(A) site usage), forms a direct subunit–subunit interface with Rpb3 via its conserved region H/C-terminus, undergoes TANGO6/COPI-vesicle-mediated nuclear import from the Golgi during G1 phase, and requires Rtr1 (coordinating with Gpn3 and Npa3) for proper cytoplasmic assembly with Rpb1 before nuclear import; additionally, a gain-of-function Rpb2 variant (N44Y) disrupts RNAi-dependent heterochromatin through an Elp1-dependent pathway."},"narrative":{"mechanistic_narrative":"POLR2B (RPB2) is the second-largest subunit of RNA polymerase II and contributes directly to the enzyme's catalytic core and to multiple steps of mRNA synthesis [PMID:15886393]. Its conserved active-center residue E791 (E836 in yeast) coordinates NTP and Mg(B) loading, and substitution to alanine impairs both NTP polymerization and intrinsic transcript cleavage at limiting Mg/NTP concentrations [PMID:15886393]. Beyond catalysis, the Rpb2 lobe domain functionally interacts with the TFIIB B-finger to govern transcription start-site selection and to support elongation [PMID:15082791], and rpb2 mutations alter transcriptional elongation efficiency that is read out in vivo as shifts toward upstream and internal poly(A) site usage [PMID:14560031, PMID:23355614]. The lobe surface also serves as a docking platform for the elongation factor IWS1, whose short linear motifs contact this region to stimulate elongation. RPB2 forms a direct subunit-subunit interface with Rpb3 through its conserved C-terminal region [PMID:9738888, PMID:12552808]. Before functioning in the nucleus, RPB2 is assembled with Rpb1 in the cytoplasm in a process requiring the assembly factor Rtr1 acting with Gpn3 and Npa3 [PMID:36190433], and is then trafficked from the cis-Golgi through the ER to the nucleus by TANGO6 in association with COPI vesicles during G1, a route whose disruption causes cytoplasmic accumulation of RPB2 and G1 arrest [PMID:38490996]. Distinct rpb2 alleles also couple the polymerase to chromatin and RNA-processing outputs, including RNAi-dependent heterochromatin maintenance via an Elp1-dependent pathway.","teleology":[{"year":1998,"claim":"Establishing how the two largest polymerase subunits are held together: mapping where Rpb2 contacts Rpb3 defined a physical anchor point within the assembled enzyme.","evidence":"Yeast two-hybrid mapping of S. pombe Rpb2 fragments against Rpb3","pmids":["9738888"],"confidence":"Low","gaps":["Single two-hybrid mapping with no orthogonal or structural validation","Interface boundaries refined only in later mapping work","Does not address functional consequence of disrupting the interface"]},{"year":2000,"claim":"Connecting Rpb2 to initiation-stage regulators: a genetically interacting rpb2 allele and co-purification placed Ssu72 in physical and functional contact with the polymerase core.","evidence":"Genetic suppressor screen plus co-IP with purified RNAP II in yeast","pmids":["11046131"],"confidence":"Medium","gaps":["Does not define the direct Rpb2 surface contacted","Mechanism of the functional interaction during initiation not resolved"]},{"year":2001,"claim":"Refining the Rpb2-Rpb3 interface to a defined C-terminal segment localized the subunit contact within Rpb2 primary sequence.","evidence":"Yeast two-hybrid mapping with Rpb2 fragment constructs in S. pombe","pmids":["12552808"],"confidence":"Low","gaps":["Single-method mapping without structural confirmation","No test of whether the segment is necessary for assembly in vivo"]},{"year":2003,"claim":"Linking Rpb2 to elongation in vivo: rpb2 mutations shifted poly(A) site choice, showing that elongation defects manifest as altered 3'-end processing.","evidence":"In vivo poly(A) site reporter analysis in yeast rpb2 mutants","pmids":["14560031"],"confidence":"Medium","gaps":["Does not pinpoint which Rpb2 structural element drives the elongation defect","Indirect readout of elongation through poly(A) choice"]},{"year":2004,"claim":"Defining a start-site selection role: the Rpb2 lobe domain was shown to interact functionally with the TFIIB B-finger and to also affect elongation.","evidence":"Genetic suppressor analysis, in vitro run-on and abortive initiation assays, 6-azauracil sensitivity in yeast","pmids":["15082791"],"confidence":"Medium","gaps":["Physical contact inferred genetically rather than structurally at the time","Quantitative contribution to start-site scanning not isolated"]},{"year":2005,"claim":"Assigning a catalytic role to a specific Rpb2 residue: E791 was shown to be required for NTP-Mg(B) loading, defining Rpb2's contribution to the active center.","evidence":"Human RNAP II E791A mutant purification with in vitro and in vivo transcription and transcript cleavage assays","pmids":["15886393"],"confidence":"High","gaps":["Does not resolve full coordination geometry of the active site","Other Rpb2 active-center residues not systematically tested here"]},{"year":2011,"claim":"Testing the necessity of the Rpb2 flap loop/TFIIB interface: its deletion left global initiation, elongation, pausing, and cleavage largely intact, bounding the functional importance of this surface.","evidence":"Flap loop deletion mutant in HEK293 with genome-wide ChIP-seq and multiple in vitro transcription assays","pmids":["21670157"],"confidence":"High","gaps":["A modest pausing effect was masked by TFIIF, leaving its physiological relevance unresolved","Negative result does not exclude redundant contributions from adjacent surfaces"]},{"year":2013,"claim":"Tying elongation rate to stress-responsive 3'-end choice: UV damage redirected RPB2 poly(A) site usage, showing elongation rate governs poly(A) selection on the RPB2 gene itself.","evidence":"Poly(A) site usage analysis, 3'UTR reporters, and mRNA stability assays before/after UV in yeast","pmids":["23355614"],"confidence":"Medium","gaps":["Whether this autoregulatory behavior generalizes to RPB2 protein levels not established","Molecular sensor coupling damage to elongation rate not identified"]},{"year":2022,"claim":"Defining cytoplasmic assembly of the enzyme: Rtr1, with Gpn3 and Npa3, was shown to be required for proper assembly of Rpb1 and Rpb2 prior to nuclear import.","evidence":"Genetic suppressor analysis, co-IP, localization microscopy, and a catalytically dead Rtr1 mutant in yeast","pmids":["36190433"],"confidence":"Medium","gaps":["Order and stoichiometry of assembly intermediates not fully resolved","How phosphatase-independent Rtr1 promotes assembly mechanistically unclear"]},{"year":2023,"claim":"Connecting RPB2 to tumor cell growth: POLR2B knockdown suppressed glioblastoma proliferation and altered a downstream target, linking the subunit to a disease-relevant transcriptional output.","evidence":"shRNA knockdown, proliferation and xenograft assays, RNA-seq identifying DDIT4 in GBM cells","pmids":["37423037"],"confidence":"Low","gaps":["Phenotype may reflect general loss of polymerase function rather than specific regulation","DDIT4 regulation not shown to be direct","Single lab without rescue controls"]},{"year":2024,"claim":"Resolving how RPB2 reaches the nucleus: TANGO6 captures RPB2 in the cis-Golgi via COPI vesicles and routes it through the ER to the nucleus during G1.","evidence":"Co-IP, live-cell imaging of RPB2 trafficking, cell cycle analysis, and conditional TANGO6 perturbation in mouse HSCs","pmids":["38490996"],"confidence":"Medium","gaps":["How RPB2 is loaded onto the Golgi/COPI route is not defined","Relationship between this trafficking step and cytoplasmic Rtr1-dependent assembly not integrated"]},{"year":2025,"claim":"Extending Rpb2 function to chromatin silencing: a gain-of-function rpb2-N44Y allele impaired RNAi-dependent heterochromatin through an Elp1-dependent pathway.","evidence":"CRISPR mutagenesis, genetic epistasis with elp1/elp3 deletions, siRNA and heterochromatin reporter assays in S. pombe (preprint)","pmids":[],"confidence":"Medium","gaps":["Preprint, single lab","Direct biochemical link between Rpb2-N44Y and Elp1 chromatin function not established"]},{"year":2025,"claim":"Structurally defining elongation- and splicing-factor docking on Rpb2: cryo-EM placed IWS1 SLiMs and the U1-70K RRM on the RPB2 lobe domain.","evidence":"Cryo-EM of Pol II complexes plus SLiM deletion and in vitro elongation assays; co-IP in yeast (preprints)","pmids":[],"confidence":"Medium","gaps":["Preprints, single labs","Human U1-70K-RPB2 contact partly relies on a referenced prior structure","Functional importance of the splicing contact in cells not established"]},{"year":null,"claim":"How cytoplasmic Rtr1-dependent assembly, Golgi/COPI-TANGO6 trafficking, and nuclear catalytic function are temporally and mechanistically integrated into a single biogenesis pathway for RPB2 remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking assembly, trafficking, and nuclear loading","Regulation of RPB2 biogenesis across the cell cycle incompletely defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140098","term_label":"catalytic activity, acting on RNA","supporting_discovery_ids":[3]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[3]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[3,5]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[9]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[8]},{"term_id":"GO:0005794","term_label":"Golgi apparatus","supporting_discovery_ids":[9]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[3,2,1]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[1,7,13]}],"complexes":["RNA polymerase II"],"partners":["RPB3","TFIIB","IWS1","TANGO6","RTR1","SSU72","U1-70K"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P30876","full_name":"DNA-directed RNA polymerase II subunit RPB2","aliases":["3'-5' exoribonuclease","DNA-directed RNA polymerase II 140 kDa polypeptide","DNA-directed RNA polymerase II subunit B","RNA polymerase II subunit 2","RNA polymerase II subunit B2","RNA-directed RNA polymerase II subunit RPB2"],"length_aa":1174,"mass_kda":133.9,"function":"Catalytic 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 (By similarity) (PubMed:27193682, PubMed:30190596, PubMed:9852112). Pol II-mediated transcription cycle proceeds through transcription initiation, transcription elongation and transcription termination stages. During transcription initiation, Pol II pre-initiation complex (PIC) is recruited to DNA promoters, with focused-type promoters containing either the initiator (Inr) element, or the TATA-box found in cell-type specific genes and dispersed-type promoters that often contain hypomethylated CpG islands usually found in housekeeping genes. Once the polymerase has escaped from the promoter it enters the elongation phase during which RNA is actively polymerized, based on complementarity with the template DNA strand. Transcription termination involves the release of the RNA transcript and polymerase from the DNA (PubMed:27193682, PubMed:30190596, PubMed:9852112). Forms Pol II active center together with the largest subunit POLR2A/RPB1. Appends one nucleotide at a time to the 3' end of the nascent RNA, with POLR2A/RPB1 most likely contributing a Mg(2+)-coordinating DxDGD motif and POLR2B/RPB2 participating in the coordination of a second Mg(2+) ion and providing lysine residues believed to facilitate Watson-Crick base pairing between the incoming nucleotide and template base. Typically, Mg(2+) ions direct a 5' nucleoside triphosphate to form a phosphodiester bond with the 3' hydroxyl of the preceding nucleotide of the nascent RNA, with the elimination of pyrophosphate. The reversible pyrophosphorolysis can occur at high pyrophosphate concentrations (By similarity) (PubMed:30190596, PubMed:9852112). Can proofread the nascent RNA transcript by means of a 3' -> 5' exonuclease activity. If a ribonucleotide is mis-incorporated, backtracks along the template DNA and cleaves the phosphodiester bond releasing the mis-incorporated 5'-ribonucleotide (By similarity) (PubMed:8381534) RNA-dependent RNA polymerase that catalyzes the extension of a non-coding RNA (ncRNA) at the 3'-end using the four ribonucleoside triphosphates as substrates. An internal ncRNA sequence near the 3'-end serves as a template in a single-round Pol II-mediated RNA polymerization reaction. May decrease the stability of ncRNAs that repress Pol II-mediated gene transcription","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/P30876/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/POLR2B","classification":"Common Essential","n_dependent_lines":1207,"n_total_lines":1208,"dependency_fraction":0.9991721854304636},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000047315","cell_line_id":"CID000697","localizations":[{"compartment":"nuclear_punctae","grade":3},{"compartment":"nucleoplasm","grade":3},{"compartment":"cytoplasmic","grade":1}],"interactors":[{"gene":"GTF2F1","stoichiometry":10.0},{"gene":"MED19","stoichiometry":10.0},{"gene":"POLR2A","stoichiometry":10.0},{"gene":"POLR2I","stoichiometry":10.0},{"gene":"POLR2K","stoichiometry":10.0},{"gene":"GTF2F2","stoichiometry":10.0},{"gene":"POLR2E","stoichiometry":10.0},{"gene":"POLR2C","stoichiometry":10.0},{"gene":"POLR2H","stoichiometry":10.0},{"gene":"POLR2J3;POLR2J;POLR2J2","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/target/CID000697","total_profiled":1310},"omim":[{"mim_id":"621545","title":"GPN-LOOP GTPase 3; GPN3","url":"https://www.omim.org/entry/621545"},{"mim_id":"621544","title":"GPN-LOOP GTPase 2; GPN2","url":"https://www.omim.org/entry/621544"},{"mim_id":"614366","title":"POLYMERASE III, RNA, SUBUNIT B; POLR3B","url":"https://www.omim.org/entry/614366"},{"mim_id":"609881","title":"RNA POLYMERASE II, SUBUNIT J2; POLR2J2","url":"https://www.omim.org/entry/609881"},{"mim_id":"603075","title":"MACULAR DEGENERATION, AGE-RELATED, 1; ARMD1","url":"https://www.omim.org/entry/603075"}],"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/POLR2B"},"hgnc":{"alias_symbol":["RPB2"],"prev_symbol":[]},"alphafold":{"accession":"P30876","domains":[{"cath_id":"-","chopping":"62-194_395-470","consensus_level":"high","plddt":88.3726,"start":62,"end":470},{"cath_id":"3.90.1110.10","chopping":"199-390","consensus_level":"high","plddt":87.4489,"start":199,"end":390},{"cath_id":"2.40.270.10","chopping":"713-807_930-1049","consensus_level":"medium","plddt":95.2731,"start":713,"end":1049},{"cath_id":"2.40.50.150","chopping":"808-875_886-926","consensus_level":"medium","plddt":91.129,"start":808,"end":926},{"cath_id":"3.90.1800.10","chopping":"1077-1164","consensus_level":"medium","plddt":93.9309,"start":1077,"end":1164}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P30876","model_url":"https://alphafold.ebi.ac.uk/files/AF-P30876-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P30876-F1-predicted_aligned_error_v6.png","plddt_mean":89.94},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=POLR2B","jax_strain_url":"https://www.jax.org/strain/search?query=POLR2B"},"sequence":{"accession":"P30876","fasta_url":"https://rest.uniprot.org/uniprotkb/P30876.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P30876/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P30876"}},"corpus_meta":[{"pmid":"15737578","id":"PMC_15737578","title":"Improving phylogenetic inference of mushrooms with RPB1 and RPB2 nucleotide sequences (Inocybe; Agaricales).","date":"2005","source":"Molecular phylogenetics and evolution","url":"https://pubmed.ncbi.nlm.nih.gov/15737578","citation_count":379,"is_preprint":false},{"pmid":"23357352","id":"PMC_23357352","title":"Phylogenetic analyses of RPB1 and RPB2 support a middle Cretaceous origin for a clade comprising all agriculturally and medically important fusaria.","date":"2013","source":"Fungal genetics and biology : FG & B","url":"https://pubmed.ncbi.nlm.nih.gov/23357352","citation_count":280,"is_preprint":false},{"pmid":"15288074","id":"PMC_15288074","title":"Contribution of RPB2 to multilocus phylogenetic studies of the euascomycetes (Pezizomycotina, Fungi) with special emphasis on the lichen-forming Acarosporaceae and evolution of polyspory.","date":"2004","source":"Molecular phylogenetics and evolution","url":"https://pubmed.ncbi.nlm.nih.gov/15288074","citation_count":277,"is_preprint":false},{"pmid":"17081773","id":"PMC_17081773","title":"Contributions of rpb2 and tef1 to the phylogeny of mushrooms and allies (Basidiomycota, Fungi).","date":"2006","source":"Molecular phylogenetics and evolution","url":"https://pubmed.ncbi.nlm.nih.gov/17081773","citation_count":262,"is_preprint":false},{"pmid":"16085431","id":"PMC_16085431","title":"Lower level relationships in the mushroom genus Cortinarius (Basidiomycota, Agaricales): a comparison of RPB1, RPB2, and ITS phylogenies.","date":"2005","source":"Molecular phylogenetics and evolution","url":"https://pubmed.ncbi.nlm.nih.gov/16085431","citation_count":84,"is_preprint":false},{"pmid":"15904853","id":"PMC_15904853","title":"Evolutionary relationships of the cup-fungus genus Peziza and Pezizaceae inferred from multiple nuclear genes: RPB2, beta-tubulin, and LSU rDNA.","date":"2005","source":"Molecular phylogenetics and 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during transcription initiation.\",\n      \"method\": \"Genetic suppressor screen, co-immunoprecipitation with purified RNAP II, in vivo transcription assays\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal genetic and biochemical evidence from a single lab using two orthogonal methods (suppressor genetics + co-IP)\",\n      \"pmids\": [\"11046131\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Mutations in the RPB2 subunit of RNA Pol II (along with SPT5 and TFIIS) cause increased utilization of internal and upstream poly(A) sites in yeast, indicating that transcriptional elongation defects leading to pausing or arrest can be visualized in vivo through poly(A) site choice, and that RPB2 plays a role in transcriptional elongation efficiency.\",\n      \"method\": \"In vivo gene expression analysis using poly(A) site reporter genes in yeast rpb2 mutants\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo functional assay with rpb2 mutants and multiple gene targets, single lab\",\n      \"pmids\": [\"14560031\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"A functional interaction between the B-finger domain of TFIIB and the lobe domain of Rpb2 (G369S substitution) was identified through genetic suppressor analysis. The Rpb2 lobe-jaw region defines a novel role for Rpb2 in transcription start site selection, and the sua7-3 rpb2-101 double mutant was sensitive to 6-azauracil in vivo and to NTP substrate depletion in vitro, linking Rpb2 lobe domain to elongation as well.\",\n      \"method\": \"Genetic suppressor screen, in vitro promoter-specific run-on transcription, abortive initiation assay, 6-azauracil sensitivity\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis plus multiple in vitro transcription assays in single lab\",\n      \"pmids\": [\"15082791\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The highly conserved Rpb2 residue E791 (human RNAP II; E836 in yeast) is involved in NTP and Mg(B) binding at the active center. The E791A substitution mutant showed impaired transcription activity at low NTP concentrations both in vitro and in vivo, and decreased NTP polymerization and transcript cleavage activities at low Mg concentrations, implicating this residue in NTP-Mg(B) loading into the active site.\",\n      \"method\": \"Affinity purification of human RNAP II E791A mutant, in vitro transcription assay, in vivo transcription assay, transcript cleavage assay at varying Mg and NTP concentrations\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — active-site mutagenesis with both in vitro enzymatic assays and in vivo validation in human RNAP II\",\n      \"pmids\": [\"15886393\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Deletion of the RPB2 flap loop (Δ873-884) in human RNAP II, which removes the TFIIB interaction interface, had no detectable effect on global transcription initiation, RNAP II occupancy, promoter escape, productive elongation, promoter binding, abortive initiation, TFIIS-stimulated transcript cleavage, or NELF/DSIF inhibition in vitro. A modest effect on elongation and pausing was suppressed by TFIIF.\",\n      \"method\": \"Genome-wide ChIP-seq in HEK293 cells expressing flap loop deletion mutant, in vitro transcription and abortive initiation assays, TFIIS cleavage assay, NELF/DSIF pausing assay\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mutagenesis with multiple orthogonal in vitro and genome-wide in vivo assays; negative result rigorously established\",\n      \"pmids\": [\"21670157\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Two-hybrid analysis using S. pombe RNAP II subunits mapped an Rpb3 contact site within the conserved region H of Rpb2 (the beta-subunit homology region conserved among all RNA polymerases), establishing a direct physical interaction between Rpb2 and Rpb3 within the polymerase complex.\",\n      \"method\": \"Yeast two-hybrid assay with fission yeast Rpb2 fragments\",\n      \"journal\": \"Molecular & general genetics\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single two-hybrid mapping study, no orthogonal validation\",\n      \"pmids\": [\"9738888\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Two-hybrid mapping in S. pombe identified the Rpb2-Rpb3 interaction site at the C-terminal region of Rpb2, within amino acids 902–989 (encoded by base 2701–2966 of Rpb2 cDNA).\",\n      \"method\": \"Yeast two-hybrid assay with Rpb2 fragment constructs\",\n      \"journal\": \"Wei sheng wu xue bao (Acta Microbiologica Sinica)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single two-hybrid mapping, single lab, no orthogonal validation\",\n      \"pmids\": [\"12552808\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"UV damage regulates alternative polyadenylation of the RPB2 gene in yeast: in normally growing cells, the proximal poly(A) site is preferentially used; after UV damage and transcription recovery, the distal poly(A) site is preferentially used. The 3'UTR of RPB2 is sufficient for this regulation, and the rate of transcription elongation (not initiation) is an important determinant of poly(A) site choice.\",\n      \"method\": \"RNA analysis of poly(A) site usage before and after UV damage, 3'UTR reporter constructs, mRNA stability assays\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple experimental approaches (reporter constructs, stability assays, transcription rate manipulation) from single lab\",\n      \"pmids\": [\"23355614\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Rtr1 is directly required for assembly of the two largest RNAP II subunits (Rpb1 and Rpb2). Deletion of RTR1 leads to cytoplasmic clumping of RNAP II subunits. Rtr1 coordinates with Gpn3 and Npa3 assembly factors; overexpression of RTR1 suppresses cytoplasmic clump formation of RNAP II subunit in a gpn3-9 mutant. This function does not require Rtr1 phosphatase catalytic activity.\",\n      \"method\": \"Genetic suppressor analysis, co-immunoprecipitation, fluorescence microscopy of RNAP II subunit localization, analysis of catalytically inactive Rtr1 mutant\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis combined with localization microscopy and biochemical co-IP, single lab\",\n      \"pmids\": [\"36190433\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"TANGO6 associates with COPI vesicles and captures RPB2 in the cis-Golgi during G1 phase, then carries RPB2 to the ER and subsequently to the nucleus. Functional disruption of TANGO6 causes cytoplasmic accumulation of RPB2 and G1 cell cycle arrest. Conditional depletion or overexpression of TANGO6 in mouse hematopoietic stem cells compromises or expands hematopoiesis.\",\n      \"method\": \"Co-immunoprecipitation, live cell fluorescence imaging, cell cycle analysis (PI staining), conditional knockout/overexpression in mouse hematopoietic stem cells\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP, live imaging of RPB2 trafficking, functional rescue experiments; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"38490996\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Knockdown of POLR2B (RPB2) by shRNA suppresses GBM tumor cell growth in vitro and in vivo in a xenograft model, and RNA sequencing identified DDIT4 (DNA damage-inducible transcript 4) as a downstream transcriptional target regulated by RPB2 in glioblastoma cells.\",\n      \"method\": \"shRNA knockdown, cell proliferation assay, xenograft mouse model, RNA sequencing, GO and GSEA analysis\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, knockdown phenotype with downstream target identification but limited mechanistic pathway resolution\",\n      \"pmids\": [\"37423037\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A gain-of-function mutation in the Rpb2 subunit of Pol II (rpb2-N44Y in S. pombe) reduces RNAi-dependent heterochromatin. The heterochromatin defects of rpb2-N44Y require Elongator Protein 1 (Elp1); loss of Elp1 (but not other Elongator subunits such as Elp3) suppresses these defects independently of the mcm5s2U34 tRNA modification, revealing an Elp1 chromatin function downstream of or parallel to Rpb2.\",\n      \"method\": \"CRISPR site-directed mutagenesis, genetic epistasis with elp1/elp3 deletions, siRNA quantification, heterochromatin reporter assays in S. pombe\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR mutagenesis with genetic epistasis and multiple molecular readouts; preprint, single lab\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Cryo-EM mapping showed that IWS1 short linear motifs (SLiMs) in its intrinsically disordered C-terminal region directly contact the RPB2 lobe domain of Pol II. Functional assays demonstrated that IWS1 interactions with the RPB2 lobe and ELOF1 are required for IWS1-dependent transcription elongation stimulation, while IWS1 recruitment depends on the RPB1 jaw domain and downstream DNA binding.\",\n      \"method\": \"Cryo-electron microscopy structure determination, deletion mutagenesis of SLiMs, in vitro transcription elongation assays\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure plus functional mutagenesis assays; preprint, single lab\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In yeast, Prp40 (not U1-70K) is the predominant mediator of U1 snRNP interaction with Pol II; multiple domains of Prp40 interact with Pol II independently of the CTD. In humans, the cryo-EM structure showed that U1-70K RRM domain directly interacts with the RPB2 subunit of Pol II, mediating co-transcriptional splicing coupling.\",\n      \"method\": \"Co-immunoprecipitation in yeast, cryo-EM structure of human U1 snRNP–Pol II complex, domain deletion analysis\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — preprint; the RPB2-specific finding in humans is based on a referenced prior cryo-EM structure, with primary new data in yeast using co-IP\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"POLR2B (RPB2) encodes the second-largest subunit of RNA polymerase II; it contributes to the catalytic active site (E791 coordinates NTP-Mg(B) loading), participates in transcription start site selection via its lobe domain (interacting functionally with TFIIB B-finger), influences transcriptional elongation efficiency (with rpb2 mutations causing increased upstream poly(A) site usage), forms a direct subunit–subunit interface with Rpb3 via its conserved region H/C-terminus, undergoes TANGO6/COPI-vesicle-mediated nuclear import from the Golgi during G1 phase, and requires Rtr1 (coordinating with Gpn3 and Npa3) for proper cytoplasmic assembly with Rpb1 before nuclear import; additionally, a gain-of-function Rpb2 variant (N44Y) disrupts RNAi-dependent heterochromatin through an Elp1-dependent pathway.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"POLR2B (RPB2) is the second-largest subunit of RNA polymerase II and contributes directly to the enzyme's catalytic core and to multiple steps of mRNA synthesis [#3]. Its conserved active-center residue E791 (E836 in yeast) coordinates NTP and Mg(B) loading, and substitution to alanine impairs both NTP polymerization and intrinsic transcript cleavage at limiting Mg/NTP concentrations [#3]. Beyond catalysis, the Rpb2 lobe domain functionally interacts with the TFIIB B-finger to govern transcription start-site selection and to support elongation [#2], and rpb2 mutations alter transcriptional elongation efficiency that is read out in vivo as shifts toward upstream and internal poly(A) site usage [#1, #7]. The lobe surface also serves as a docking platform for the elongation factor IWS1, whose short linear motifs contact this region to stimulate elongation [#12]. RPB2 forms a direct subunit-subunit interface with Rpb3 through its conserved C-terminal region [#5, #6]. Before functioning in the nucleus, RPB2 is assembled with Rpb1 in the cytoplasm in a process requiring the assembly factor Rtr1 acting with Gpn3 and Npa3 [#8], and is then trafficked from the cis-Golgi through the ER to the nucleus by TANGO6 in association with COPI vesicles during G1, a route whose disruption causes cytoplasmic accumulation of RPB2 and G1 arrest [#9]. Distinct rpb2 alleles also couple the polymerase to chromatin and RNA-processing outputs, including RNAi-dependent heterochromatin maintenance via an Elp1-dependent pathway [#11].\"\n,\n  \"teleology\": [\n    {\n      \"year\": 1998,\n      \"claim\": \"Establishing how the two largest polymerase subunits are held together: mapping where Rpb2 contacts Rpb3 defined a physical anchor point within the assembled enzyme.\",\n      \"evidence\": \"Yeast two-hybrid mapping of S. pombe Rpb2 fragments against Rpb3\",\n      \"pmids\": [\"9738888\"],\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Single two-hybrid mapping with no orthogonal or structural validation\", \"Interface boundaries refined only in later mapping work\", \"Does not address functional consequence of disrupting the interface\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Connecting Rpb2 to initiation-stage regulators: a genetically interacting rpb2 allele and co-purification placed Ssu72 in physical and functional contact with the polymerase core.\",\n      \"evidence\": \"Genetic suppressor screen plus co-IP with purified RNAP II in yeast\",\n      \"pmids\": [\"11046131\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Does not define the direct Rpb2 surface contacted\", \"Mechanism of the functional interaction during initiation not resolved\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Refining the Rpb2-Rpb3 interface to a defined C-terminal segment localized the subunit contact within Rpb2 primary sequence.\",\n      \"evidence\": \"Yeast two-hybrid mapping with Rpb2 fragment constructs in S. pombe\",\n      \"pmids\": [\"12552808\"],\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Single-method mapping without structural confirmation\", \"No test of whether the segment is necessary for assembly in vivo\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Linking Rpb2 to elongation in vivo: rpb2 mutations shifted poly(A) site choice, showing that elongation defects manifest as altered 3'-end processing.\",\n      \"evidence\": \"In vivo poly(A) site reporter analysis in yeast rpb2 mutants\",\n      \"pmids\": [\"14560031\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Does not pinpoint which Rpb2 structural element drives the elongation defect\", \"Indirect readout of elongation through poly(A) choice\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Defining a start-site selection role: the Rpb2 lobe domain was shown to interact functionally with the TFIIB B-finger and to also affect elongation.\",\n      \"evidence\": \"Genetic suppressor analysis, in vitro run-on and abortive initiation assays, 6-azauracil sensitivity in yeast\",\n      \"pmids\": [\"15082791\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Physical contact inferred genetically rather than structurally at the time\", \"Quantitative contribution to start-site scanning not isolated\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Assigning a catalytic role to a specific Rpb2 residue: E791 was shown to be required for NTP-Mg(B) loading, defining Rpb2's contribution to the active center.\",\n      \"evidence\": \"Human RNAP II E791A mutant purification with in vitro and in vivo transcription and transcript cleavage assays\",\n      \"pmids\": [\"15886393\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Does not resolve full coordination geometry of the active site\", \"Other Rpb2 active-center residues not systematically tested here\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Testing the necessity of the Rpb2 flap loop/TFIIB interface: its deletion left global initiation, elongation, pausing, and cleavage largely intact, bounding the functional importance of this surface.\",\n      \"evidence\": \"Flap loop deletion mutant in HEK293 with genome-wide ChIP-seq and multiple in vitro transcription assays\",\n      \"pmids\": [\"21670157\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"A modest pausing effect was masked by TFIIF, leaving its physiological relevance unresolved\", \"Negative result does not exclude redundant contributions from adjacent surfaces\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Tying elongation rate to stress-responsive 3'-end choice: UV damage redirected RPB2 poly(A) site usage, showing elongation rate governs poly(A) selection on the RPB2 gene itself.\",\n      \"evidence\": \"Poly(A) site usage analysis, 3'UTR reporters, and mRNA stability assays before/after UV in yeast\",\n      \"pmids\": [\"23355614\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Whether this autoregulatory behavior generalizes to RPB2 protein levels not established\", \"Molecular sensor coupling damage to elongation rate not identified\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defining cytoplasmic assembly of the enzyme: Rtr1, with Gpn3 and Npa3, was shown to be required for proper assembly of Rpb1 and Rpb2 prior to nuclear import.\",\n      \"evidence\": \"Genetic suppressor analysis, co-IP, localization microscopy, and a catalytically dead Rtr1 mutant in yeast\",\n      \"pmids\": [\"36190433\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Order and stoichiometry of assembly intermediates not fully resolved\", \"How phosphatase-independent Rtr1 promotes assembly mechanistically unclear\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Connecting RPB2 to tumor cell growth: POLR2B knockdown suppressed glioblastoma proliferation and altered a downstream target, linking the subunit to a disease-relevant transcriptional output.\",\n      \"evidence\": \"shRNA knockdown, proliferation and xenograft assays, RNA-seq identifying DDIT4 in GBM cells\",\n      \"pmids\": [\"37423037\"],\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Phenotype may reflect general loss of polymerase function rather than specific regulation\", \"DDIT4 regulation not shown to be direct\", \"Single lab without rescue controls\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Resolving how RPB2 reaches the nucleus: TANGO6 captures RPB2 in the cis-Golgi via COPI vesicles and routes it through the ER to the nucleus during G1.\",\n      \"evidence\": \"Co-IP, live-cell imaging of RPB2 trafficking, cell cycle analysis, and conditional TANGO6 perturbation in mouse HSCs\",\n      \"pmids\": [\"38490996\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"How RPB2 is loaded onto the Golgi/COPI route is not defined\", \"Relationship between this trafficking step and cytoplasmic Rtr1-dependent assembly not integrated\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extending Rpb2 function to chromatin silencing: a gain-of-function rpb2-N44Y allele impaired RNAi-dependent heterochromatin through an Elp1-dependent pathway.\",\n      \"evidence\": \"CRISPR mutagenesis, genetic epistasis with elp1/elp3 deletions, siRNA and heterochromatin reporter assays in S. pombe (preprint)\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Preprint, single lab\", \"Direct biochemical link between Rpb2-N44Y and Elp1 chromatin function not established\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Structurally defining elongation- and splicing-factor docking on Rpb2: cryo-EM placed IWS1 SLiMs and the U1-70K RRM on the RPB2 lobe domain.\",\n      \"evidence\": \"Cryo-EM of Pol II complexes plus SLiM deletion and in vitro elongation assays; co-IP in yeast (preprints)\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Preprints, single labs\", \"Human U1-70K-RPB2 contact partly relies on a referenced prior structure\", \"Functional importance of the splicing contact in cells not established\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How cytoplasmic Rtr1-dependent assembly, Golgi/COPI-TANGO6 trafficking, and nuclear catalytic function are temporally and mechanistically integrated into a single biogenesis pathway for RPB2 remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"No unified model linking assembly, trafficking, and nuclear loading\", \"Regulation of RPB2 biogenesis across the cell cycle incompletely defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140098\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [3, 5]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [9]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [8]},\n      {\"term_id\": \"GO:0005794\", \"supporting_discovery_ids\": [9]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [3, 2, 1]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [1, 7, 13]}\n    ],\n    \"complexes\": [\n      \"RNA polymerase II\"\n    ],\n    \"partners\": [\n      \"RPB3\",\n      \"TFIIB\",\n      \"IWS1\",\n      \"TANGO6\",\n      \"RTR1\",\n      \"SSU72\",\n      \"U1-70K\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}