{"gene":"RPL15","run_date":"2026-06-10T06:43:37","timeline":{"discoveries":[{"year":2013,"finding":"RPL15 is required both for 60S ribosomal subunit formation and for efficient cleavage of internal transcribed spacer 1 (ITS1) during pre-rRNA processing. Cells from patients with RPL15 deletions show defective pre-ribosomal RNA processing.","method":"Array-CGH identification of deletion, pre-rRNA processing analysis in patient-derived cells","journal":"Human genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional pre-rRNA processing assay in patient cells, single lab, two orthogonal approaches (genetic deletion + rRNA processing readout)","pmids":["23812780"],"is_preprint":false},{"year":2018,"finding":"RPL15 mutations (truncating and missense) cause defective pre-rRNA processing, reduced 60S ribosomal subunit formation, severe proliferation defects, elevated TP53 activity, and increased apoptosis in erythroblast cells, establishing RPL15 as integral to 60S subunit biogenesis and the TP53 stress-response pathway in hematopoietic cells.","method":"In vitro pre-rRNA processing assays, ribosome profile analysis, red cell culture assays with primary erythroblasts, flow cytometry for apoptosis and TP53 activity","journal":"Haematologica","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (rRNA processing, subunit profiling, proliferation, apoptosis, TP53 assays) in patient-derived primary cells, independently replicating findings from PMID:23812780","pmids":["29599205"],"is_preprint":false},{"year":2022,"finding":"Topotecan (TPT) directly binds RPL15 and inhibits pre-ribosomal subunit formation. TPT binding to RPL15 disrupts RPL15–RPL4 protein interactions and decreases RPL4 stability; CDK12 activity can recover RPL4 stability. RPL15 knockdown induces DAMP secretion and activates cGAS-STING-mediated antitumor immune responses independent of TOP1.","method":"Co-immunoprecipitation (RPL15–RPL4 interaction), drug-binding assay, RPL15 knockdown in B16-F10 murine melanoma model, DAMP secretion assay, flow cytometry for CTL/Treg populations, CDK12 activity rescue experiment","journal":"Journal of immunology (Baltimore, Md. : 1950)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP for RPL15-RPL4, in vivo knockdown with defined immune readouts, single lab with multiple orthogonal methods","pmids":["35725272"],"is_preprint":false},{"year":2022,"finding":"RPL15 knockdown activates the RPs-MDM2-p53 pathway: RPL15 silencing affects the interaction between p53, MDM2, and RPL5/RPL11 (assessed by co-immunoprecipitation and cycloheximide chase), leading to p53 stabilization, cell cycle arrest, and suppression of HCC cell proliferation, invasion, and migration.","method":"RPL15 knockdown/overexpression, co-immunoprecipitation of p53/MDM2/RPL5/RPL11, cycloheximide chase assay, Western blot for p53/p21/CDK2/Cyclin E1/EMT markers, xenograft model","journal":"Cancer cell international","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and CHX chase for pathway placement, single lab, multiple orthogonal methods including in vivo xenograft","pmids":["35410346"],"is_preprint":false},{"year":2023,"finding":"In yeast, depletion of eL15 (RPL15 ortholog) causes defective processing of 27SA3 to 27SBS pre-rRNA and impaired 27SB processing to mature 25S and 5.8S rRNAs, efficient turnover of de novo-formed 27S pre-rRNAs, blocked nucleocytoplasmic export of pre-60S particles, and disrupted assembly of neighboring ribosomal proteins eL8 and eL36 and associated A3- and B-factor assembly factors. eL15 assembly is a prerequisite for shaping domain I of 5.8S/25S rRNA within early pre-60S particles.","method":"In vivo depletion of eL15 in S. cerevisiae, polysome profiling, Northern blot pre-rRNA processing analysis, mass spectrometry composition of pre-60S particles, nucleocytoplasmic export assay","journal":"Journal of molecular biology","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — multiple orthogonal methods (polysome profiling, Northern blot, MS-based particle composition, export assay) in yeast ortholog system with rigorous controls","pmids":["37865285"],"is_preprint":false},{"year":2025,"finding":"A PROTAC degrader (SN38-PROTAC) conjugating SN-38 to pomalidomide induces ubiquitin-mediated proteasomal degradation of RPL15 (but not TOP1), confirming that RPL15 is the relevant target for DAMP secretion and cGAS-STING activation in dendritic cells, and sensitizing tumors to anti-PD-1 therapy in a STING-dependent manner.","method":"PROTAC synthesis, Western blot for RPL15/TOP1 degradation, ubiquitin pathway dependency assay, DAMP secretion assay, cGAS-STING reporter in dendritic cells, in vivo B16-F10 mouse model with STING-deficient controls","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — selective PROTAC degradation confirms target identity, mechanistic cGAS-STING pathway placement confirmed in STING-KO mice, single lab","pmids":["41276688"],"is_preprint":false},{"year":2025,"finding":"Free (ribosome-unbound) RPL15 (eL15) is ADP-ribosylated (PARylated) by PARP1 and PARP2 in vitro, identifying it as one of the major targets of 60S ribosomal protein PARylation; ribosome-bound RPL15 was not detectably modified.","method":"In vitro ADP-ribosylation assay with radiolabeled NAD+, using isolated ribosomal subunit proteins and recombinant PARP1/PARP2; free vs. ribosome-bound protein comparison","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 / Weak — in vitro biochemical assay, single lab, preprint not yet peer-reviewed, single method","pmids":["bio_10.1101_2025.09.15.676193"],"is_preprint":true}],"current_model":"RPL15 (eL15) is a large ribosomal subunit protein essential for early 60S subunit biogenesis: it is required for pre-rRNA processing (including ITS1 cleavage and 27SA3→27SB→25S/5.8S rRNA maturation), assembly of neighboring ribosomal proteins (eL8, eL36), recruitment of A3- and B-factor assembly complexes, and nucleocytoplasmic export of pre-60S particles; loss of RPL15 activates the RPs-MDM2-p53 stress pathway leading to p53 stabilization and apoptosis; topotecan directly binds RPL15 and disrupts RPL15–RPL4 interactions, triggering ribosomal stress and DAMP secretion that activates cGAS-STING antitumor immunity; and free RPL15 can be ADP-ribosylated by PARP1/PARP2 in vitro."},"narrative":{"mechanistic_narrative":"RPL15 (eL15) is a large ribosomal subunit protein essential for early 60S subunit biogenesis, where it acts during pre-rRNA processing and pre-60S particle assembly [PMID:29599205, PMID:37865285]. RPL15 is required for efficient cleavage of internal transcribed spacer 1 (ITS1) and for the 27SA3→27SB→25S/5.8S maturation steps of the large-subunit rRNA, and its loss causes turnover of nascent 27S pre-rRNAs and reduced 60S formation [PMID:23812780, PMID:29599205, PMID:37865285]. Within the assembling particle, RPL15/eL15 incorporation is a prerequisite for shaping domain I of the 5.8S/25S rRNA, for the assembly of neighboring ribosomal proteins eL8 and eL36, for recruitment of A3- and B-factor assembly complexes, and for nucleocytoplasmic export of pre-60S particles [PMID:37865285]. Disruption of RPL15 function triggers ribosomal stress: loss activates the RPs-MDM2-p53 pathway through RPL5/RPL11, stabilizing p53 to drive cell-cycle arrest and apoptosis, a response evident as severe proliferation defects and elevated TP53 activity in erythroblasts and as suppressed proliferation, invasion, and migration in hepatocellular carcinoma cells [PMID:29599205, PMID:35410346]. Topotecan directly binds RPL15 and disrupts the RPL15–RPL4 interaction while inhibiting pre-ribosomal subunit formation, and depletion of RPL15 induces DAMP secretion that activates cGAS-STING-mediated antitumor immunity in a manner that is independent of TOP1 and dependent on STING [PMID:35725272, PMID:41276688].","teleology":[{"year":2013,"claim":"Established that RPL15 is needed for human large-subunit biogenesis by linking patient deletions to a specific pre-rRNA processing defect rather than a generic loss.","evidence":"Array-CGH deletion mapping and pre-rRNA processing analysis in patient-derived cells","pmids":["23812780"],"confidence":"Medium","gaps":["Did not define which assembly factors or neighboring proteins depend on RPL15","Causal mechanism linking processing defect to disease phenotype not resolved"]},{"year":2018,"claim":"Connected RPL15 loss-of-function mutations to the TP53 stress-response axis in hematopoietic cells, explaining how a biogenesis defect produces a proliferation/apoptosis phenotype.","evidence":"In vitro pre-rRNA processing, ribosome profiling, primary erythroblast culture, and flow cytometry for apoptosis and TP53 activity","pmids":["29599205"],"confidence":"High","gaps":["Did not delineate the molecular bridge between RPL15 deficiency and p53 activation","Cell-type specificity of erythroid sensitivity not mechanistically explained"]},{"year":2022,"claim":"Placed RPL15 in the canonical ribosomal-stress p53 circuit by showing its silencing alters p53/MDM2 engagement of RPL5/RPL11 to stabilize p53.","evidence":"RPL15 knockdown/overexpression with Co-IP of p53/MDM2/RPL5/RPL11, cycloheximide chase, and xenograft in HCC cells","pmids":["35410346"],"confidence":"Medium","gaps":["Whether RPL15 acts upstream of free RPL5/RPL11 release versus directly on MDM2 not distinguished","Direct binding partners within the RPL5/RPL11-MDM2 complex not mapped"]},{"year":2022,"claim":"Identified RPL15 as a direct drug target whose perturbation links ribosomal stress to innate antitumor immunity, separating this activity from topoisomerase inhibition.","evidence":"Drug-binding assay, reciprocal Co-IP of RPL15–RPL4, RPL15 knockdown in B16-F10 melanoma, DAMP secretion and immune-cell flow cytometry, CDK12 rescue","pmids":["35725272"],"confidence":"Medium","gaps":["Structural basis of topotecan–RPL15 binding not defined","Mechanism linking RPL4 destabilization to DAMP secretion not resolved"]},{"year":2023,"claim":"Defined the mechanistic role of eL15 in 60S assembly, showing it gates 27S pre-rRNA processing, neighboring protein incorporation, assembly factor recruitment, and pre-60S export.","evidence":"Yeast eL15 depletion with polysome profiling, Northern blot, MS analysis of pre-60S particle composition, and nucleocytoplasmic export assay","pmids":["37865285"],"confidence":"High","gaps":["Conservation of every step in human cells not directly demonstrated","Order of eL15 incorporation relative to eL8/eL36 within the assembly hierarchy not fully resolved"]},{"year":2025,"claim":"Confirmed via selective degradation that RPL15 itself, not TOP1, is the effector for DAMP secretion and cGAS-STING activation, and showed therapeutic synergy with anti-PD-1.","evidence":"SN38-PROTAC inducing ubiquitin-proteasomal RPL15 degradation, DAMP and cGAS-STING reporter assays in dendritic cells, and B16-F10 model with STING-deficient controls","pmids":["41276688"],"confidence":"Medium","gaps":["The signaling intermediates between RPL15 degradation and STING activation are undefined","Whether immune activation requires the biogenesis defect or a separate RPL15 function is unclear"]},{"year":2025,"claim":"Identified free RPL15 as a major substrate of PARP1/PARP2 ADP-ribosylation, raising the possibility of a post-translational regulatory layer on ribosome-unbound RPL15.","evidence":"In vitro ADP-ribosylation assay with radiolabeled NAD+ comparing free versus ribosome-bound subunit proteins (preprint)","pmids":["bio_10.1101_2025.09.15.676193"],"confidence":"Low","gaps":["In vitro only; not confirmed in cells or peer-reviewed","Functional consequence of PARylation on RPL15 unknown","Modified residues not mapped"]},{"year":null,"claim":"It remains unknown how RPL15-dependent biogenesis defects mechanistically couple to the distinct downstream outputs of p53 stabilization versus cGAS-STING immune activation, and whether post-translational modification of free RPL15 regulates these branches.","evidence":"","pmids":[],"confidence":"Low","gaps":["No unified model integrating biogenesis, p53 stress, and innate immune signaling","No structural data on RPL15 within human pre-60S particles or drug complexes"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[1,4]},{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[4]}],"localization":[{"term_id":"GO:0005840","term_label":"ribosome","supporting_discovery_ids":[1,4]},{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[4]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,1,4]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[1,4]}],"complexes":["60S ribosomal subunit","pre-60S particle"],"partners":["RPL4","RPL5","RPL11","MDM2","TP53","EL8","EL36"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P61313","full_name":"Large ribosomal subunit protein eL15","aliases":["60S ribosomal protein L15"],"length_aa":204,"mass_kda":24.1,"function":"Component of the large ribosomal subunit. The ribosome is a large ribonucleoprotein complex responsible for the synthesis of proteins in the cell","subcellular_location":"Cytoplasm","url":"https://www.uniprot.org/uniprotkb/P61313/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/RPL15","classification":"Common Essential","n_dependent_lines":317,"n_total_lines":317,"dependency_fraction":1.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"RACK1","stoichiometry":10.0},{"gene":"RPL10A","stoichiometry":10.0},{"gene":"RPL11","stoichiometry":10.0},{"gene":"RPL35","stoichiometry":10.0},{"gene":"RPL5","stoichiometry":10.0},{"gene":"RPS16","stoichiometry":10.0},{"gene":"SRP68","stoichiometry":10.0},{"gene":"SRP9","stoichiometry":10.0},{"gene":"DRG1","stoichiometry":4.0},{"gene":"ENY2","stoichiometry":4.0}],"url":"https://opencell.sf.czbiohub.org/search/RPL15","total_profiled":1310},"omim":[{"mim_id":"615550","title":"DIAMOND-BLACKFAN ANEMIA 12; DBA12","url":"https://www.omim.org/entry/615550"},{"mim_id":"614900","title":"DIAMOND-BLACKFAN ANEMIA 11; DBA11","url":"https://www.omim.org/entry/614900"},{"mim_id":"613309","title":"DIAMOND-BLACKFAN ANEMIA 10; DBA10","url":"https://www.omim.org/entry/613309"},{"mim_id":"613308","title":"DIAMOND-BLACKFAN ANEMIA 9; DBA9","url":"https://www.omim.org/entry/613308"},{"mim_id":"612563","title":"DIAMOND-BLACKFAN ANEMIA 8; DBA8","url":"https://www.omim.org/entry/612563"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/RPL15"},"hgnc":{"alias_symbol":["RPL10","RPLY10","RPYL10","EC45","L15","eL15"],"prev_symbol":[]},"alphafold":{"accession":"P61313","domains":[{"cath_id":"3.40.1120.10","chopping":"2-172","consensus_level":"high","plddt":96.8122,"start":2,"end":172},{"cath_id":"-","chopping":"176-204","consensus_level":"medium","plddt":92.931,"start":176,"end":204}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P61313","model_url":"https://alphafold.ebi.ac.uk/files/AF-P61313-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P61313-F1-predicted_aligned_error_v6.png","plddt_mean":96.19},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=RPL15","jax_strain_url":"https://www.jax.org/strain/search?query=RPL15"},"sequence":{"accession":"P61313","fasta_url":"https://rest.uniprot.org/uniprotkb/P61313.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P61313/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P61313"}},"corpus_meta":[{"pmid":"23812780","id":"PMC_23812780","title":"Novel deletion of RPL15 identified by array-comparative genomic hybridization in Diamond-Blackfan anemia.","date":"2013","source":"Human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/23812780","citation_count":91,"is_preprint":false},{"pmid":"27612557","id":"PMC_27612557","title":"Inhibition of Aurora kinases induces apoptosis and autophagy via AURKB/p70S6K/RPL15 axis in human leukemia cells.","date":"2016","source":"Cancer letters","url":"https://pubmed.ncbi.nlm.nih.gov/27612557","citation_count":38,"is_preprint":false},{"pmid":"29599205","id":"PMC_29599205","title":"Recurring mutations in RPL15 are linked to hydrops fetalis and treatment independence in Diamond-Blackfan anemia.","date":"2018","source":"Haematologica","url":"https://pubmed.ncbi.nlm.nih.gov/29599205","citation_count":26,"is_preprint":false},{"pmid":"1581570","id":"PMC_1581570","title":"Characterization of rps17, rp19 and rpl15: three nucleus-encoded plastid ribosomal protein genes.","date":"1992","source":"Plant molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/1581570","citation_count":25,"is_preprint":false},{"pmid":"35410346","id":"PMC_35410346","title":"RPL15 promotes hepatocellular carcinoma progression via regulation of RPs-MDM2-p53 signaling pathway.","date":"2022","source":"Cancer cell international","url":"https://pubmed.ncbi.nlm.nih.gov/35410346","citation_count":14,"is_preprint":false},{"pmid":"35725272","id":"PMC_35725272","title":"Identification of RPL15 60S Ribosomal Protein as a Novel Topotecan Target Protein That Correlates with DAMP Secretion and Antitumor Immune Activation.","date":"2022","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/35725272","citation_count":14,"is_preprint":false},{"pmid":"32572886","id":"PMC_32572886","title":"Circ-RPL15/miR-146b-3p/VEGFA feedback loop is responsible for triggering proliferation and migration in glioma.","date":"2020","source":"European review for medical and pharmacological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/32572886","citation_count":12,"is_preprint":false},{"pmid":"35117718","id":"PMC_35117718","title":"The miR-567/RPL15/TGF-β/Smad axis inhibits the stem-like properties and chemo-resistance of gastric cancer cells.","date":"2020","source":"Translational cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/35117718","citation_count":8,"is_preprint":false},{"pmid":"11129599","id":"PMC_11129599","title":"A cDNA encoding ribosomal protein RPL15 from the desiccation-tolerant bryophyte Tortula ruralis: mRNA transcripts are stably maintained in desiccated and rehydrated gametophytes.","date":"2000","source":"Bioscience, biotechnology, and biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/11129599","citation_count":6,"is_preprint":false},{"pmid":"37431269","id":"PMC_37431269","title":"Decreased expression of RPL15 and RPL18 exacerbated the calcification of valve interstitial cells during aortic valve calcification.","date":"2023","source":"Cell biology international","url":"https://pubmed.ncbi.nlm.nih.gov/37431269","citation_count":4,"is_preprint":false},{"pmid":"37865285","id":"PMC_37865285","title":"The Role of Ribosomal Proteins eL15 and eL36 in the Early Steps of Yeast 60S Ribosomal Subunit Assembly.","date":"2023","source":"Journal of molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/37865285","citation_count":3,"is_preprint":false},{"pmid":"7733938","id":"PMC_7733938","title":"The Thermoplasma acidophilum rpl15 gene encodes a homologue of eukaryotic ribosomal proteins L15/YL10.","date":"1995","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/7733938","citation_count":2,"is_preprint":false},{"pmid":"41276688","id":"PMC_41276688","title":"A selective RPL15 PROTAC degrader enhances anti-PD-1 immunotherapy in a murine melanoma tumor model.","date":"2025","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/41276688","citation_count":1,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.09.15.676193","title":"Interplay between human ribosomal proteins, PARP1, PARP2, HPF1 and histones","date":"2025-09-17","source":"bioRxiv","url":"https://doi.org/10.1101/2025.09.15.676193","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":9507,"output_tokens":2205,"usd":0.030798,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9154,"output_tokens":2935,"usd":0.059573,"stage2_stop_reason":"end_turn"},"total_usd":0.090371,"stage1_batch_id":"msgbatch_01CNoDbkChHEkgrJnL7BwtGJ","stage2_batch_id":"msgbatch_016Y4x9E9Wg5VQDANj9quSQD","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2013,\n      \"finding\": \"RPL15 is required both for 60S ribosomal subunit formation and for efficient cleavage of internal transcribed spacer 1 (ITS1) during pre-rRNA processing. Cells from patients with RPL15 deletions show defective pre-ribosomal RNA processing.\",\n      \"method\": \"Array-CGH identification of deletion, pre-rRNA processing analysis in patient-derived cells\",\n      \"journal\": \"Human genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional pre-rRNA processing assay in patient cells, single lab, two orthogonal approaches (genetic deletion + rRNA processing readout)\",\n      \"pmids\": [\"23812780\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"RPL15 mutations (truncating and missense) cause defective pre-rRNA processing, reduced 60S ribosomal subunit formation, severe proliferation defects, elevated TP53 activity, and increased apoptosis in erythroblast cells, establishing RPL15 as integral to 60S subunit biogenesis and the TP53 stress-response pathway in hematopoietic cells.\",\n      \"method\": \"In vitro pre-rRNA processing assays, ribosome profile analysis, red cell culture assays with primary erythroblasts, flow cytometry for apoptosis and TP53 activity\",\n      \"journal\": \"Haematologica\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (rRNA processing, subunit profiling, proliferation, apoptosis, TP53 assays) in patient-derived primary cells, independently replicating findings from PMID:23812780\",\n      \"pmids\": [\"29599205\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Topotecan (TPT) directly binds RPL15 and inhibits pre-ribosomal subunit formation. TPT binding to RPL15 disrupts RPL15–RPL4 protein interactions and decreases RPL4 stability; CDK12 activity can recover RPL4 stability. RPL15 knockdown induces DAMP secretion and activates cGAS-STING-mediated antitumor immune responses independent of TOP1.\",\n      \"method\": \"Co-immunoprecipitation (RPL15–RPL4 interaction), drug-binding assay, RPL15 knockdown in B16-F10 murine melanoma model, DAMP secretion assay, flow cytometry for CTL/Treg populations, CDK12 activity rescue experiment\",\n      \"journal\": \"Journal of immunology (Baltimore, Md. : 1950)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP for RPL15-RPL4, in vivo knockdown with defined immune readouts, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"35725272\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"RPL15 knockdown activates the RPs-MDM2-p53 pathway: RPL15 silencing affects the interaction between p53, MDM2, and RPL5/RPL11 (assessed by co-immunoprecipitation and cycloheximide chase), leading to p53 stabilization, cell cycle arrest, and suppression of HCC cell proliferation, invasion, and migration.\",\n      \"method\": \"RPL15 knockdown/overexpression, co-immunoprecipitation of p53/MDM2/RPL5/RPL11, cycloheximide chase assay, Western blot for p53/p21/CDK2/Cyclin E1/EMT markers, xenograft model\",\n      \"journal\": \"Cancer cell international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and CHX chase for pathway placement, single lab, multiple orthogonal methods including in vivo xenograft\",\n      \"pmids\": [\"35410346\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In yeast, depletion of eL15 (RPL15 ortholog) causes defective processing of 27SA3 to 27SBS pre-rRNA and impaired 27SB processing to mature 25S and 5.8S rRNAs, efficient turnover of de novo-formed 27S pre-rRNAs, blocked nucleocytoplasmic export of pre-60S particles, and disrupted assembly of neighboring ribosomal proteins eL8 and eL36 and associated A3- and B-factor assembly factors. eL15 assembly is a prerequisite for shaping domain I of 5.8S/25S rRNA within early pre-60S particles.\",\n      \"method\": \"In vivo depletion of eL15 in S. cerevisiae, polysome profiling, Northern blot pre-rRNA processing analysis, mass spectrometry composition of pre-60S particles, nucleocytoplasmic export assay\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — multiple orthogonal methods (polysome profiling, Northern blot, MS-based particle composition, export assay) in yeast ortholog system with rigorous controls\",\n      \"pmids\": [\"37865285\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A PROTAC degrader (SN38-PROTAC) conjugating SN-38 to pomalidomide induces ubiquitin-mediated proteasomal degradation of RPL15 (but not TOP1), confirming that RPL15 is the relevant target for DAMP secretion and cGAS-STING activation in dendritic cells, and sensitizing tumors to anti-PD-1 therapy in a STING-dependent manner.\",\n      \"method\": \"PROTAC synthesis, Western blot for RPL15/TOP1 degradation, ubiquitin pathway dependency assay, DAMP secretion assay, cGAS-STING reporter in dendritic cells, in vivo B16-F10 mouse model with STING-deficient controls\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — selective PROTAC degradation confirms target identity, mechanistic cGAS-STING pathway placement confirmed in STING-KO mice, single lab\",\n      \"pmids\": [\"41276688\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Free (ribosome-unbound) RPL15 (eL15) is ADP-ribosylated (PARylated) by PARP1 and PARP2 in vitro, identifying it as one of the major targets of 60S ribosomal protein PARylation; ribosome-bound RPL15 was not detectably modified.\",\n      \"method\": \"In vitro ADP-ribosylation assay with radiolabeled NAD+, using isolated ribosomal subunit proteins and recombinant PARP1/PARP2; free vs. ribosome-bound protein comparison\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — in vitro biochemical assay, single lab, preprint not yet peer-reviewed, single method\",\n      \"pmids\": [\"bio_10.1101_2025.09.15.676193\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"RPL15 (eL15) is a large ribosomal subunit protein essential for early 60S subunit biogenesis: it is required for pre-rRNA processing (including ITS1 cleavage and 27SA3→27SB→25S/5.8S rRNA maturation), assembly of neighboring ribosomal proteins (eL8, eL36), recruitment of A3- and B-factor assembly complexes, and nucleocytoplasmic export of pre-60S particles; loss of RPL15 activates the RPs-MDM2-p53 stress pathway leading to p53 stabilization and apoptosis; topotecan directly binds RPL15 and disrupts RPL15–RPL4 interactions, triggering ribosomal stress and DAMP secretion that activates cGAS-STING antitumor immunity; and free RPL15 can be ADP-ribosylated by PARP1/PARP2 in vitro.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"RPL15 (eL15) is a large ribosomal subunit protein essential for early 60S subunit biogenesis, where it acts during pre-rRNA processing and pre-60S particle assembly [#1, #4]. RPL15 is required for efficient cleavage of internal transcribed spacer 1 (ITS1) and for the 27SA3\\u219227SB\\u219225S/5.8S maturation steps of the large-subunit rRNA, and its loss causes turnover of nascent 27S pre-rRNAs and reduced 60S formation [#0, #1, #4]. Within the assembling particle, RPL15/eL15 incorporation is a prerequisite for shaping domain I of the 5.8S/25S rRNA, for the assembly of neighboring ribosomal proteins eL8 and eL36, for recruitment of A3- and B-factor assembly complexes, and for nucleocytoplasmic export of pre-60S particles [#4]. Disruption of RPL15 function triggers ribosomal stress: loss activates the RPs-MDM2-p53 pathway through RPL5/RPL11, stabilizing p53 to drive cell-cycle arrest and apoptosis, a response evident as severe proliferation defects and elevated TP53 activity in erythroblasts and as suppressed proliferation, invasion, and migration in hepatocellular carcinoma cells [#1, #3]. Topotecan directly binds RPL15 and disrupts the RPL15\\u2013RPL4 interaction while inhibiting pre-ribosomal subunit formation, and depletion of RPL15 induces DAMP secretion that activates cGAS-STING-mediated antitumor immunity in a manner that is independent of TOP1 and dependent on STING [#2, #5].\",\n  \"teleology\": [\n    {\n      \"year\": 2013,\n      \"claim\": \"Established that RPL15 is needed for human large-subunit biogenesis by linking patient deletions to a specific pre-rRNA processing defect rather than a generic loss.\",\n      \"evidence\": \"Array-CGH deletion mapping and pre-rRNA processing analysis in patient-derived cells\",\n      \"pmids\": [\"23812780\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not define which assembly factors or neighboring proteins depend on RPL15\", \"Causal mechanism linking processing defect to disease phenotype not resolved\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Connected RPL15 loss-of-function mutations to the TP53 stress-response axis in hematopoietic cells, explaining how a biogenesis defect produces a proliferation/apoptosis phenotype.\",\n      \"evidence\": \"In vitro pre-rRNA processing, ribosome profiling, primary erythroblast culture, and flow cytometry for apoptosis and TP53 activity\",\n      \"pmids\": [\"29599205\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not delineate the molecular bridge between RPL15 deficiency and p53 activation\", \"Cell-type specificity of erythroid sensitivity not mechanistically explained\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Placed RPL15 in the canonical ribosomal-stress p53 circuit by showing its silencing alters p53/MDM2 engagement of RPL5/RPL11 to stabilize p53.\",\n      \"evidence\": \"RPL15 knockdown/overexpression with Co-IP of p53/MDM2/RPL5/RPL11, cycloheximide chase, and xenograft in HCC cells\",\n      \"pmids\": [\"35410346\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether RPL15 acts upstream of free RPL5/RPL11 release versus directly on MDM2 not distinguished\", \"Direct binding partners within the RPL5/RPL11-MDM2 complex not mapped\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified RPL15 as a direct drug target whose perturbation links ribosomal stress to innate antitumor immunity, separating this activity from topoisomerase inhibition.\",\n      \"evidence\": \"Drug-binding assay, reciprocal Co-IP of RPL15\\u2013RPL4, RPL15 knockdown in B16-F10 melanoma, DAMP secretion and immune-cell flow cytometry, CDK12 rescue\",\n      \"pmids\": [\"35725272\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of topotecan\\u2013RPL15 binding not defined\", \"Mechanism linking RPL4 destabilization to DAMP secretion not resolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined the mechanistic role of eL15 in 60S assembly, showing it gates 27S pre-rRNA processing, neighboring protein incorporation, assembly factor recruitment, and pre-60S export.\",\n      \"evidence\": \"Yeast eL15 depletion with polysome profiling, Northern blot, MS analysis of pre-60S particle composition, and nucleocytoplasmic export assay\",\n      \"pmids\": [\"37865285\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conservation of every step in human cells not directly demonstrated\", \"Order of eL15 incorporation relative to eL8/eL36 within the assembly hierarchy not fully resolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Confirmed via selective degradation that RPL15 itself, not TOP1, is the effector for DAMP secretion and cGAS-STING activation, and showed therapeutic synergy with anti-PD-1.\",\n      \"evidence\": \"SN38-PROTAC inducing ubiquitin-proteasomal RPL15 degradation, DAMP and cGAS-STING reporter assays in dendritic cells, and B16-F10 model with STING-deficient controls\",\n      \"pmids\": [\"41276688\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The signaling intermediates between RPL15 degradation and STING activation are undefined\", \"Whether immune activation requires the biogenesis defect or a separate RPL15 function is unclear\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified free RPL15 as a major substrate of PARP1/PARP2 ADP-ribosylation, raising the possibility of a post-translational regulatory layer on ribosome-unbound RPL15.\",\n      \"evidence\": \"In vitro ADP-ribosylation assay with radiolabeled NAD+ comparing free versus ribosome-bound subunit proteins (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.09.15.676193\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"In vitro only; not confirmed in cells or peer-reviewed\", \"Functional consequence of PARylation on RPL15 unknown\", \"Modified residues not mapped\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unknown how RPL15-dependent biogenesis defects mechanistically couple to the distinct downstream outputs of p53 stabilization versus cGAS-STING immune activation, and whether post-translational modification of free RPL15 regulates these branches.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No unified model integrating biogenesis, p53 stress, and innate immune signaling\", \"No structural data on RPL15 within human pre-60S particles or drug complexes\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [1, 4]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005840\", \"supporting_discovery_ids\": [1, 4]},\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 1, 4]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [1, 4]}\n    ],\n    \"complexes\": [\"60S ribosomal subunit\", \"pre-60S particle\"],\n    \"partners\": [\"RPL4\", \"RPL5\", \"RPL11\", \"MDM2\", \"TP53\", \"eL8\", \"eL36\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}