{"gene":"SHPRH","run_date":"2026-06-10T07:46:31","timeline":{"discoveries":[{"year":2006,"finding":"SHPRH physically interacts with the Rad6-Rad18 and Mms2-Ubc13 complexes and functions as a ubiquitin E3 ligase indispensable for Mms2-Ubc13-dependent K63-linked polyubiquitylation of PCNA, establishing it as a functional homolog of yeast Rad5 in error-free postreplicational repair.","method":"Co-immunoprecipitation, in vitro ubiquitylation assay, active-site analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro ubiquitylation assay plus physical interaction studies, replicated in a parallel independent study (PMID:17130289)","pmids":["17108083"],"is_preprint":false},{"year":2006,"finding":"SHPRH associates with PCNA, RAD18, and UBC13 and promotes MMS-induced PCNA polyubiquitination; shRNA-mediated reduction of SHPRH increases MMS sensitivity and enhances genomic instability, placing SHPRH in the Rad5-orthologous error-free bypass branch of postreplication repair.","method":"Co-immunoprecipitation, stable shRNA knockdown, genomic instability assays, MMS sensitivity assay","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, loss-of-function with defined cellular phenotype, replicated independently (PMID:17108083)","pmids":["17130289"],"is_preprint":false},{"year":2008,"finding":"HLTF, a second RAD5 homolog, interacts with UBC13, PCNA, and SHPRH and promotes K63-linked PCNA polyubiquitination cooperatively with SHPRH; reduction of either SHPRH or HLTF enhances spontaneous mutagenesis.","method":"Co-immunoprecipitation, siRNA knockdown, mutagenesis assay, MEF chromosome break analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, multiple knockdown phenotypes, replicated across labs","pmids":["18719106"],"is_preprint":false},{"year":2011,"finding":"SHPRH and HLTF act in a damage-specific manner: MMS promotes HLTF degradation and enables SHPRH interactions with Rad18 and polymerase κ for error-free bypass, while UV activates HLTF-dependent TLS and HLTF inhibits SHPRH function. This demonstrates coordinated regulation of the two main PRR branches by these RAD5 homologs.","method":"Co-immunoprecipitation, siRNA knockdown, mutagenesis assay, TLS polymerase recruitment assay","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods, functional epistasis established, published in high-impact peer-reviewed journal","pmids":["21396873"],"is_preprint":false},{"year":2011,"finding":"In Shprh/Hltf double-mutant mouse embryonic fibroblasts, PCNA polyubiquitination (PCNA-Ubn) is not abolished, demonstrating that an alternative E3 ligase exists for PCNA polyubiquitination beyond SHPRH and HLTF.","method":"Double-knockout mouse genetics, immunoblot for PCNA ubiquitination, B-cell somatic hypermutation assay","journal":"DNA repair","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic double knockout with clean biochemical readout (PCNA-Ubn), in vivo model","pmids":["21269891"],"is_preprint":false},{"year":2013,"finding":"The NMR solution structure of the SHPRH PHD domain adopts a canonical PHD-finger fold with a central two-stranded anti-parallel β-sheet flanked by interleaved zinc-binding sites; NMR titration experiments reveal that the SHPRH PHD domain does NOT specifically interact with H3-derived peptides regardless of K4 methylation status, suggesting an alternative function for this domain.","method":"Solution NMR structure determination, NMR titration experiments with histone H3 peptides","journal":"Journal of biomolecular NMR","confidence":"High","confidence_rationale":"Tier 1 / Moderate — NMR structure with functional validation (binding experiments), single lab","pmids":["23907177"],"is_preprint":false},{"year":2017,"finding":"SHPRH localizes to ribosomal DNA (rDNA) promoters in nucleoli and facilitates rRNA transcription; the PHD domain mediates interaction with histone H3 when H3K4 is not trimethylated. SHPRH enrichment at the rDNA promoter requires CHD4, is inhibited by rapamycin (mTOR inhibition), starvation, or actinomycin D, and SHPRH physically interacts with the RNA Polymerase I complex.","method":"ChIP (chromatin immunoprecipitation), co-immunoprecipitation with RNA Pol I, shRNA knockdown, pharmacological inhibition (rapamycin, actinomycin D), subcellular fractionation/localization","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Moderate — ChIP, Co-IP with Pol I complex, multiple orthogonal perturbations in one study","pmids":["28400511"],"is_preprint":false},{"year":2019,"finding":"SHPRH is a nucleosome-stimulated ATPase that binds equally well to dsDNA and nucleosome core particles but shows preference for nucleosomes with extranucleosomal DNA; nucleosomes (but not dsDNA) strongly stimulate SHPRH ATPase activity without conventional remodeling. SHPRH also functions as a potent nucleosome E3-ubiquitin ligase active with at least 7 different E2 enzymes; with UBE2D1, it catalyzes branched polyubiquitin linkages and linkages associated with DNA repair factor recruitment or proteasomal degradation.","method":"In vitro ATPase assay, nucleosome binding assay, E2 enzyme screen (26 E2s), in vitro ubiquitylation assay, mass spectrometry of ubiquitin linkages","journal":"Epigenetics & chromatin","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstituted in vitro biochemical assays with multiple substrates and mass spectrometry validation, single lab","pmids":["31434570"],"is_preprint":false},{"year":2019,"finding":"TonEBP (tonicity-responsive enhancer-binding protein) is recruited to DNA damage sites with bulky adducts and sequentially recruits E3 ubiquitin ligase SHPRH and then deubiquitinase USP1 to DNA damage sites, regulating the dynamics of PCNA polyubiquitination; the Rel-homology domain of TonEBP is essential for interaction with SHPRH and USP1.","method":"Co-immunoprecipitation, laser micro-irradiation with live-cell imaging of recruitment, siRNA knockdown, immunoblot for PCNA ubiquitination","journal":"iScience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, live-cell recruitment assay, knockdown phenotype; single lab, multiple methods","pmids":["31376680"],"is_preprint":false},{"year":2020,"finding":"SHPRH contains a functional APIM (PCNA-interacting) motif, and nuclear localization of SHPRH depends on direct interaction with PCNA through this motif; mutation of APIM alters mutation spectra.","method":"APIM mutagenesis, nuclear localization assay, mutation spectrum analysis (SupF assay), overexpression","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — APIM mutagenesis with localization and functional readout; single lab","pmids":["31973093"],"is_preprint":false},{"year":2020,"finding":"SHPRH knockout cells show reduced activation of checkpoint kinase CHK2 and MCM2 after MMS treatment, identifying SHPRH as a regulator of the CHK2-dependent DNA damage response branch.","method":"CRISPR/gene knockout, immunoblot for CHK2 and MCM2 phosphorylation, drug sensitivity assays","journal":"Biomolecules","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — knockout with defined biochemical readout (CHK2/MCM2 phosphorylation); single lab, single method per endpoint","pmids":["32192191"],"is_preprint":false},{"year":2022,"finding":"SHPRH interacts with MMR protein MLH1 (but not MSH2); this interaction is conserved from yeast Rad5, where a MIP-box motif mediates Mlh1 interaction. Depletion of SHPRH results in mild resistance to alkylating agents, suggesting a role in MMR-associated apoptotic response.","method":"Co-immunoprecipitation, yeast two-hybrid, bioinformatic MIP-box identification, siRNA knockdown with drug sensitivity assay","journal":"Frontiers in cell and developmental biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP confirmed interaction, functional knockdown assay; single lab","pmids":["35784486"],"is_preprint":false},{"year":2021,"finding":"KMT2B (lysine methyltransferase 2B) elevates SHPRH expression via H3K4me3 modification at the SHPRH promoter; SHPRH in turn modulates FYN ubiquitination, promoting FYN protein degradation.","method":"ChIP for H3K4me3 at SHPRH promoter, co-immunoprecipitation, ubiquitination assay, overexpression/knockdown with immunoblot","journal":"Cellular signalling","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP and ubiquitination assay; single lab, multiple methods","pmids":["34606908"],"is_preprint":false},{"year":2024,"finding":"Cryo-EM structure of SHPRH bound to the nucleosome at 2.8 Å resolution confirms that SHPRH interacts with the H2A-H2B acidic patch of the nucleosome, reveals that the SHPRH ATPase engages a different nucleosomal DNA location than other SF2-type ATPases, and clarifies the structural roles of SHPRH domains in nucleosome recognition.","method":"Cryo-EM structure determination at 2.8 Å, AlphaFold-Multimer prediction","journal":"bioRxiv (preprint) / also published as PMID:38979307","confidence":"High","confidence_rationale":"Tier 1 / Moderate — high-resolution cryo-EM structure with structural validation; single lab but rigorous structural method","pmids":["38979307"],"is_preprint":true},{"year":2026,"finding":"SHPRH directly ubiquitinates unmodified PCNA at Lys164 using the Ube2D family of E2-conjugating enzymes (not only Ubc13-Mms2), acting robustly on both free and DNA-bound PCNA; this activity requires the SHPRH HIRAN domain for PCNA interaction.","method":"In vitro ubiquitylation assay with purified components, mass spectrometry to confirm Lys164 modification, HIRAN domain mutagenesis/deletion","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstituted in vitro biochemical assay with mutagenesis of HIRAN domain and mass spectrometry validation; single lab","pmids":["41990034"],"is_preprint":false},{"year":2018,"finding":"SHPRH-146aa, a protein encoded by circ-SHPRH via IRES-driven translation, protects full-length SHPRH from ubiquitin-proteasome degradation, thereby stabilizing SHPRH and allowing SHPRH to function as an E3 ligase ubiquitinating PCNA to suppress glioblastoma cell proliferation.","method":"Overexpression/knockdown in glioblastoma cell lines, in vivo xenograft, co-immunoprecipitation, ubiquitin proteasome degradation assay, PCNA ubiquitination immunoblot","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, functional overexpression/knockdown in vitro and in vivo; this discovery concerns the circRNA-encoded peptide's effect on full-length SHPRH protein stability, which is mechanistically about the canonical SHPRH protein","pmids":["29343848"],"is_preprint":false}],"current_model":"SHPRH is a SWI2/SNF2-family E3 ubiquitin ligase and RAD5 ortholog that promotes error-free DNA damage tolerance by catalyzing K63-linked polyubiquitination of PCNA at Lys164 (using Ubc13-Mms2 or Ube2D E2s), requires its HIRAN domain for PCNA interaction, binds the nucleosome acidic patch and extranucleosomal DNA through a distinct ATPase engagement mode, localizes to rDNA promoters via its PHD domain to facilitate mTOR-dependent rRNA transcription through interaction with RNA Polymerase I, is recruited to DNA damage sites by TonEBP which coordinates SHPRH and USP1 activity, acts in a damage-type-specific manner relative to its paralog HLTF (MMS favors SHPRH/polymerase κ; UV favors HLTF/TLS), and also interacts with MLH1 and can ubiquitinate FYN, reflecting roles beyond replication fork bypass."},"narrative":{"mechanistic_narrative":"SHPRH is a SWI2/SNF2-family E3 ubiquitin ligase and human RAD5 ortholog that operates in error-free DNA damage tolerance by catalyzing K63-linked polyubiquitination of PCNA at Lys164, thereby channeling stalled replication into the error-free postreplication-repair branch [PMID:17108083, PMID:17130289]. It assembles with the Rad6-Rad18 and Mms2-Ubc13 complexes and binds PCNA to drive MMS-induced PCNA polyubiquitination, with its loss increasing alkylation sensitivity and genomic instability [PMID:17108083, PMID:17130289]. SHPRH and its paralog HLTF act in a damage-specific division of labor: MMS triggers HLTF degradation and licenses SHPRH-Rad18-polymerase κ error-free bypass, whereas UV favors HLTF-dependent translesion synthesis [PMID:21396873]. SHPRH directly ubiquitinates Lys164 of both free and DNA-bound PCNA using either Ubc13-Mms2 or the Ube2D E2 family, an activity that requires its HIRAN domain for PCNA engagement [PMID:41990034]. Biochemically it is a nucleosome-stimulated ATPase that binds the H2A-H2B acidic patch and extranucleosomal DNA through a distinct SF2 ATPase engagement mode, and acts as a promiscuous nucleosome E3 ligase capable of branched and degradation-associated ubiquitin linkages [PMID:31434570, PMID:38979307]. Beyond replication-fork bypass, SHPRH localizes through its PHD domain to nucleolar rDNA promoters, where it interacts with RNA Polymerase I to support mTOR-dependent rRNA transcription [PMID:28400511], is recruited to bulky-adduct damage sites by TonEBP that sequentially coordinates SHPRH and the deubiquitinase USP1 [PMID:31376680], and additionally engages the mismatch-repair protein MLH1 and ubiquitinates the kinase FYN [PMID:35784486, PMID:34606908].","teleology":[{"year":2006,"claim":"Established that human SHPRH is the functional RAD5 ortholog providing the missing E3 activity for the error-free branch of postreplication repair, answering how K63-linked PCNA polyubiquitination is achieved in human cells.","evidence":"Co-IP with Rad6-Rad18 and Mms2-Ubc13, in vitro ubiquitylation and active-site analysis, plus reciprocal Co-IP with knockdown phenotypes in a parallel study","pmids":["17108083","17130289"],"confidence":"High","gaps":["Did not define which domain mediates PCNA recognition","Did not resolve substrate specificity versus other E3s"]},{"year":2008,"claim":"Showed SHPRH is not the sole RAD5 homolog, revealing HLTF as a cooperating E3 in PCNA polyubiquitination and raising the question of functional redundancy versus specialization.","evidence":"Co-IP of HLTF with UBC13/PCNA/SHPRH, siRNA knockdown, mutagenesis and MEF chromosome-break assays","pmids":["18719106"],"confidence":"High","gaps":["Mechanism distinguishing SHPRH from HLTF unresolved at this stage"]},{"year":2011,"claim":"Resolved how the two RAD5 homologs are coordinated, demonstrating damage-type-specific partitioning of postreplication-repair branches, and showed an additional E3 must exist for PCNA polyubiquitination.","evidence":"Co-IP, siRNA, TLS-polymerase recruitment and mutagenesis assays; independently, Shprh/Hltf double-knockout MEFs retaining PCNA-Ubn","pmids":["21396873","21269891"],"confidence":"High","gaps":["Identity of the alternative PCNA E3 ligase not determined","Molecular trigger for damage-specific HLTF degradation not fully defined"]},{"year":2013,"claim":"Determined the SHPRH PHD-domain fold and found it does not read H3K4-methylated peptides, redirecting the search for the domain's function.","evidence":"Solution NMR structure and NMR titrations with H3 peptides","pmids":["23907177"],"confidence":"High","gaps":["Positive binding partner of the PHD domain not identified in this study"]},{"year":2017,"claim":"Uncovered a replication-independent nucleolar role, placing SHPRH at rDNA promoters to support mTOR-dependent rRNA transcription and assigning a function to its PHD domain.","evidence":"ChIP, Co-IP with RNA Pol I, shRNA, and pharmacological inhibition (rapamycin, actinomycin D)","pmids":["28400511"],"confidence":"High","gaps":["Whether E3 ligase activity is required for rRNA transcription not established","Direct substrate at the rDNA promoter not identified"]},{"year":2019,"claim":"Defined SHPRH's intrinsic biochemistry as a nucleosome-stimulated ATPase and broad nucleosome E3 ligase, indicating activity extends beyond PCNA and the Ubc13-Mms2 pair.","evidence":"Reconstituted ATPase and nucleosome-binding assays, 26-E2 screen, in vitro ubiquitylation and MS of ubiquitin linkages","pmids":["31434570"],"confidence":"High","gaps":["Physiological nucleosomal substrates not identified","In vivo relevance of branched linkages not tested"]},{"year":2019,"claim":"Identified an upstream recruiter, TonEBP, that sequentially delivers SHPRH and then USP1 to bulky-adduct lesions, explaining how PCNA ubiquitination dynamics are timed.","evidence":"Co-IP, laser micro-irradiation live-cell recruitment, siRNA, PCNA-ubiquitination immunoblot","pmids":["31376680"],"confidence":"Medium","gaps":["Single lab; reciprocal structural mapping of the SHPRH-TonEBP interface limited","How the SHPRH-to-USP1 switch is triggered unresolved"]},{"year":2020,"claim":"Linked SHPRH nuclear localization to a PCNA-interacting APIM motif and connected it to the CHK2 damage-response branch, broadening its regulatory reach.","evidence":"APIM mutagenesis with localization and SupF mutation-spectrum assays; CRISPR knockout with CHK2/MCM2 phosphorylation immunoblots","pmids":["31973093","32192191"],"confidence":"Medium","gaps":["How APIM and HIRAN-mediated PCNA contacts are coordinated unclear","Whether CHK2 effect is direct or downstream of PCNA ubiquitination not resolved"]},{"year":2021,"claim":"Extended SHPRH substrates and regulation beyond replication, showing KMT2B drives SHPRH expression and SHPRH ubiquitinates FYN for degradation.","evidence":"ChIP for H3K4me3 at the SHPRH promoter, Co-IP, ubiquitination assays and overexpression/knockdown","pmids":["34606908"],"confidence":"Medium","gaps":["Direct versus indirect ubiquitination of FYN not fully separated","Single lab"]},{"year":2022,"claim":"Demonstrated a conserved SHPRH-MLH1 interaction via a Rad5-derived MIP-box motif, implicating SHPRH in mismatch-repair-associated drug responses.","evidence":"Co-IP, yeast two-hybrid, bioinformatic MIP-box identification, siRNA with drug sensitivity","pmids":["35784486"],"confidence":"Medium","gaps":["Functional consequence of the MLH1 interaction for MMR mechanistically thin","Single lab"]},{"year":2024,"claim":"Provided the structural basis for SHPRH-nucleosome recognition, showing acidic-patch engagement and a non-canonical SF2 ATPase contact with nucleosomal DNA.","evidence":"Cryo-EM structure at 2.8 Å with AlphaFold-Multimer prediction","pmids":["38979307"],"confidence":"High","gaps":["Does not capture a SHPRH-PCNA or SHPRH-E2 complex","Catalytic cycle on a true substrate not visualized"]},{"year":2026,"claim":"Established the direct biochemical mechanism of PCNA modification, showing SHPRH ubiquitinates unmodified PCNA at Lys164 via the Ube2D family in a HIRAN-dependent manner on free and DNA-bound PCNA.","evidence":"Reconstituted in vitro ubiquitylation, MS confirmation of Lys164, HIRAN domain mutagenesis/deletion","pmids":["41990034"],"confidence":"High","gaps":["In vivo contribution of Ube2D versus Ubc13-Mms2 not quantified","Regulation of HIRAN-PCNA engagement at forks not addressed"]},{"year":null,"claim":"The integration of SHPRH's replication-fork, nucleolar transcription, and non-PCNA substrate (FYN, MLH1) activities into a single regulatory logic remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking nucleosome-ATPase activity to PCNA versus rDNA functions","Physiological nucleosomal ubiquitination substrates unidentified","Determinants choosing Ube2D versus Ubc13-Mms2 E2 usage in cells unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[0,1,7,14]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,7,14,12]},{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[7]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[7]},{"term_id":"GO:0042393","term_label":"histone binding","supporting_discovery_ids":[6]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[9]},{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[6]},{"term_id":"GO:0000228","term_label":"nuclear chromosome","supporting_discovery_ids":[7,13]}],"pathway":[{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[0,1,3]},{"term_id":"R-HSA-69306","term_label":"DNA Replication","supporting_discovery_ids":[1,14]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[6]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,12]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[7,13]}],"complexes":[],"partners":["PCNA","RAD18","UBC13","HLTF","MLH1","TONEBP","USP1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q149N8","full_name":"E3 ubiquitin-protein ligase SHPRH","aliases":["RING-type E3 ubiquitin transferase SHPRH","SNF2, histone-linker, PHD and RING finger domain-containing helicase"],"length_aa":1683,"mass_kda":193.1,"function":"E3 ubiquitin-protein ligase involved in DNA repair. Upon genotoxic stress, accepts ubiquitin from the UBE2N-UBE2V2 E2 complex and transfers it to 'Lys-164' of PCNA which had been monoubiquitinated by UBE2A/B-RAD18, promoting the formation of non-canonical poly-ubiquitin chains linked through 'Lys-63'","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q149N8/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/SHPRH","classification":"Not Classified","n_dependent_lines":3,"n_total_lines":1208,"dependency_fraction":0.0024834437086092716},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"FKBP5","stoichiometry":0.2},{"gene":"HIST2H2BE","stoichiometry":0.2},{"gene":"PARP1","stoichiometry":0.2},{"gene":"RBBP4","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/SHPRH","total_profiled":1310},"omim":[{"mim_id":"608048","title":"SNF2, LINKER HISTONE, PHD FINGER, RING FINGER, AND HELICASE DOMAINS-CONTAINING PROTEIN; SHPRH","url":"https://www.omim.org/entry/608048"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"},{"location":"Nuclear membrane","reliability":"Approved"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/SHPRH"},"hgnc":{"alias_symbol":["FLJ90837","KIAA2023","bA545I5.2"],"prev_symbol":[]},"alphafold":{"accession":"Q149N8","domains":[{"cath_id":"-","chopping":"446-498","consensus_level":"high","plddt":83.2896,"start":446,"end":498},{"cath_id":"3.40.50.300","chopping":"918-930_937-989_1509-1677","consensus_level":"high","plddt":78.0496,"start":918,"end":1677},{"cath_id":"-","chopping":"1154-1168_1182-1213_1226-1263_1291-1346","consensus_level":"medium","plddt":87.6098,"start":1154,"end":1346},{"cath_id":"3.30.40.10","chopping":"1428-1493","consensus_level":"medium","plddt":78.6745,"start":1428,"end":1493}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q149N8","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q149N8-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q149N8-F1-predicted_aligned_error_v6.png","plddt_mean":70.69},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=SHPRH","jax_strain_url":"https://www.jax.org/strain/search?query=SHPRH"},"sequence":{"accession":"Q149N8","fasta_url":"https://rest.uniprot.org/uniprotkb/Q149N8.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q149N8/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q149N8"}},"corpus_meta":[{"pmid":"29343848","id":"PMC_29343848","title":"A 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\"Co-immunoprecipitation, in vitro ubiquitylation assay, active-site analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro ubiquitylation assay plus physical interaction studies, replicated in a parallel independent study (PMID:17130289)\",\n      \"pmids\": [\"17108083\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"SHPRH associates with PCNA, RAD18, and UBC13 and promotes MMS-induced PCNA polyubiquitination; shRNA-mediated reduction of SHPRH increases MMS sensitivity and enhances genomic instability, placing SHPRH in the Rad5-orthologous error-free bypass branch of postreplication repair.\",\n      \"method\": \"Co-immunoprecipitation, stable shRNA knockdown, genomic instability assays, MMS sensitivity assay\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, loss-of-function with defined cellular phenotype, replicated independently (PMID:17108083)\",\n      \"pmids\": [\"17130289\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"HLTF, a second RAD5 homolog, interacts with UBC13, PCNA, and SHPRH and promotes K63-linked PCNA polyubiquitination cooperatively with SHPRH; reduction of either SHPRH or HLTF enhances spontaneous mutagenesis.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, mutagenesis assay, MEF chromosome break analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, multiple knockdown phenotypes, replicated across labs\",\n      \"pmids\": [\"18719106\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"SHPRH and HLTF act in a damage-specific manner: MMS promotes HLTF degradation and enables SHPRH interactions with Rad18 and polymerase κ for error-free bypass, while UV activates HLTF-dependent TLS and HLTF inhibits SHPRH function. This demonstrates coordinated regulation of the two main PRR branches by these RAD5 homologs.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, mutagenesis assay, TLS polymerase recruitment assay\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods, functional epistasis established, published in high-impact peer-reviewed journal\",\n      \"pmids\": [\"21396873\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"In Shprh/Hltf double-mutant mouse embryonic fibroblasts, PCNA polyubiquitination (PCNA-Ubn) is not abolished, demonstrating that an alternative E3 ligase exists for PCNA polyubiquitination beyond SHPRH and HLTF.\",\n      \"method\": \"Double-knockout mouse genetics, immunoblot for PCNA ubiquitination, B-cell somatic hypermutation assay\",\n      \"journal\": \"DNA repair\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic double knockout with clean biochemical readout (PCNA-Ubn), in vivo model\",\n      \"pmids\": [\"21269891\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"The NMR solution structure of the SHPRH PHD domain adopts a canonical PHD-finger fold with a central two-stranded anti-parallel β-sheet flanked by interleaved zinc-binding sites; NMR titration experiments reveal that the SHPRH PHD domain does NOT specifically interact with H3-derived peptides regardless of K4 methylation status, suggesting an alternative function for this domain.\",\n      \"method\": \"Solution NMR structure determination, NMR titration experiments with histone H3 peptides\",\n      \"journal\": \"Journal of biomolecular NMR\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — NMR structure with functional validation (binding experiments), single lab\",\n      \"pmids\": [\"23907177\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"SHPRH localizes to ribosomal DNA (rDNA) promoters in nucleoli and facilitates rRNA transcription; the PHD domain mediates interaction with histone H3 when H3K4 is not trimethylated. SHPRH enrichment at the rDNA promoter requires CHD4, is inhibited by rapamycin (mTOR inhibition), starvation, or actinomycin D, and SHPRH physically interacts with the RNA Polymerase I complex.\",\n      \"method\": \"ChIP (chromatin immunoprecipitation), co-immunoprecipitation with RNA Pol I, shRNA knockdown, pharmacological inhibition (rapamycin, actinomycin D), subcellular fractionation/localization\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP, Co-IP with Pol I complex, multiple orthogonal perturbations in one study\",\n      \"pmids\": [\"28400511\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"SHPRH is a nucleosome-stimulated ATPase that binds equally well to dsDNA and nucleosome core particles but shows preference for nucleosomes with extranucleosomal DNA; nucleosomes (but not dsDNA) strongly stimulate SHPRH ATPase activity without conventional remodeling. SHPRH also functions as a potent nucleosome E3-ubiquitin ligase active with at least 7 different E2 enzymes; with UBE2D1, it catalyzes branched polyubiquitin linkages and linkages associated with DNA repair factor recruitment or proteasomal degradation.\",\n      \"method\": \"In vitro ATPase assay, nucleosome binding assay, E2 enzyme screen (26 E2s), in vitro ubiquitylation assay, mass spectrometry of ubiquitin linkages\",\n      \"journal\": \"Epigenetics & chromatin\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstituted in vitro biochemical assays with multiple substrates and mass spectrometry validation, single lab\",\n      \"pmids\": [\"31434570\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"TonEBP (tonicity-responsive enhancer-binding protein) is recruited to DNA damage sites with bulky adducts and sequentially recruits E3 ubiquitin ligase SHPRH and then deubiquitinase USP1 to DNA damage sites, regulating the dynamics of PCNA polyubiquitination; the Rel-homology domain of TonEBP is essential for interaction with SHPRH and USP1.\",\n      \"method\": \"Co-immunoprecipitation, laser micro-irradiation with live-cell imaging of recruitment, siRNA knockdown, immunoblot for PCNA ubiquitination\",\n      \"journal\": \"iScience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, live-cell recruitment assay, knockdown phenotype; single lab, multiple methods\",\n      \"pmids\": [\"31376680\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"SHPRH contains a functional APIM (PCNA-interacting) motif, and nuclear localization of SHPRH depends on direct interaction with PCNA through this motif; mutation of APIM alters mutation spectra.\",\n      \"method\": \"APIM mutagenesis, nuclear localization assay, mutation spectrum analysis (SupF assay), overexpression\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — APIM mutagenesis with localization and functional readout; single lab\",\n      \"pmids\": [\"31973093\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"SHPRH knockout cells show reduced activation of checkpoint kinase CHK2 and MCM2 after MMS treatment, identifying SHPRH as a regulator of the CHK2-dependent DNA damage response branch.\",\n      \"method\": \"CRISPR/gene knockout, immunoblot for CHK2 and MCM2 phosphorylation, drug sensitivity assays\",\n      \"journal\": \"Biomolecules\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — knockout with defined biochemical readout (CHK2/MCM2 phosphorylation); single lab, single method per endpoint\",\n      \"pmids\": [\"32192191\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"SHPRH interacts with MMR protein MLH1 (but not MSH2); this interaction is conserved from yeast Rad5, where a MIP-box motif mediates Mlh1 interaction. Depletion of SHPRH results in mild resistance to alkylating agents, suggesting a role in MMR-associated apoptotic response.\",\n      \"method\": \"Co-immunoprecipitation, yeast two-hybrid, bioinformatic MIP-box identification, siRNA knockdown with drug sensitivity assay\",\n      \"journal\": \"Frontiers in cell and developmental biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP confirmed interaction, functional knockdown assay; single lab\",\n      \"pmids\": [\"35784486\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"KMT2B (lysine methyltransferase 2B) elevates SHPRH expression via H3K4me3 modification at the SHPRH promoter; SHPRH in turn modulates FYN ubiquitination, promoting FYN protein degradation.\",\n      \"method\": \"ChIP for H3K4me3 at SHPRH promoter, co-immunoprecipitation, ubiquitination assay, overexpression/knockdown with immunoblot\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP and ubiquitination assay; single lab, multiple methods\",\n      \"pmids\": [\"34606908\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Cryo-EM structure of SHPRH bound to the nucleosome at 2.8 Å resolution confirms that SHPRH interacts with the H2A-H2B acidic patch of the nucleosome, reveals that the SHPRH ATPase engages a different nucleosomal DNA location than other SF2-type ATPases, and clarifies the structural roles of SHPRH domains in nucleosome recognition.\",\n      \"method\": \"Cryo-EM structure determination at 2.8 Å, AlphaFold-Multimer prediction\",\n      \"journal\": \"bioRxiv (preprint) / also published as PMID:38979307\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — high-resolution cryo-EM structure with structural validation; single lab but rigorous structural method\",\n      \"pmids\": [\"38979307\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"SHPRH directly ubiquitinates unmodified PCNA at Lys164 using the Ube2D family of E2-conjugating enzymes (not only Ubc13-Mms2), acting robustly on both free and DNA-bound PCNA; this activity requires the SHPRH HIRAN domain for PCNA interaction.\",\n      \"method\": \"In vitro ubiquitylation assay with purified components, mass spectrometry to confirm Lys164 modification, HIRAN domain mutagenesis/deletion\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstituted in vitro biochemical assay with mutagenesis of HIRAN domain and mass spectrometry validation; single lab\",\n      \"pmids\": [\"41990034\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"SHPRH-146aa, a protein encoded by circ-SHPRH via IRES-driven translation, protects full-length SHPRH from ubiquitin-proteasome degradation, thereby stabilizing SHPRH and allowing SHPRH to function as an E3 ligase ubiquitinating PCNA to suppress glioblastoma cell proliferation.\",\n      \"method\": \"Overexpression/knockdown in glioblastoma cell lines, in vivo xenograft, co-immunoprecipitation, ubiquitin proteasome degradation assay, PCNA ubiquitination immunoblot\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, functional overexpression/knockdown in vitro and in vivo; this discovery concerns the circRNA-encoded peptide's effect on full-length SHPRH protein stability, which is mechanistically about the canonical SHPRH protein\",\n      \"pmids\": [\"29343848\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SHPRH is a SWI2/SNF2-family E3 ubiquitin ligase and RAD5 ortholog that promotes error-free DNA damage tolerance by catalyzing K63-linked polyubiquitination of PCNA at Lys164 (using Ubc13-Mms2 or Ube2D E2s), requires its HIRAN domain for PCNA interaction, binds the nucleosome acidic patch and extranucleosomal DNA through a distinct ATPase engagement mode, localizes to rDNA promoters via its PHD domain to facilitate mTOR-dependent rRNA transcription through interaction with RNA Polymerase I, is recruited to DNA damage sites by TonEBP which coordinates SHPRH and USP1 activity, acts in a damage-type-specific manner relative to its paralog HLTF (MMS favors SHPRH/polymerase κ; UV favors HLTF/TLS), and also interacts with MLH1 and can ubiquitinate FYN, reflecting roles beyond replication fork bypass.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"SHPRH is a SWI2/SNF2-family E3 ubiquitin ligase and human RAD5 ortholog that operates in error-free DNA damage tolerance by catalyzing K63-linked polyubiquitination of PCNA at Lys164, thereby channeling stalled replication into the error-free postreplication-repair branch [#0, #1]. It assembles with the Rad6-Rad18 and Mms2-Ubc13 complexes and binds PCNA to drive MMS-induced PCNA polyubiquitination, with its loss increasing alkylation sensitivity and genomic instability [#0, #1]. SHPRH and its paralog HLTF act in a damage-specific division of labor: MMS triggers HLTF degradation and licenses SHPRH-Rad18-polymerase \\u03ba error-free bypass, whereas UV favors HLTF-dependent translesion synthesis [#3]. SHPRH directly ubiquitinates Lys164 of both free and DNA-bound PCNA using either Ubc13-Mms2 or the Ube2D E2 family, an activity that requires its HIRAN domain for PCNA engagement [#14]. Biochemically it is a nucleosome-stimulated ATPase that binds the H2A-H2B acidic patch and extranucleosomal DNA through a distinct SF2 ATPase engagement mode, and acts as a promiscuous nucleosome E3 ligase capable of branched and degradation-associated ubiquitin linkages [#7, #13]. Beyond replication-fork bypass, SHPRH localizes through its PHD domain to nucleolar rDNA promoters, where it interacts with RNA Polymerase I to support mTOR-dependent rRNA transcription [#6], is recruited to bulky-adduct damage sites by TonEBP that sequentially coordinates SHPRH and the deubiquitinase USP1 [#8], and additionally engages the mismatch-repair protein MLH1 and ubiquitinates the kinase FYN [#11, #12].\"\n  ,\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Established that human SHPRH is the functional RAD5 ortholog providing the missing E3 activity for the error-free branch of postreplication repair, answering how K63-linked PCNA polyubiquitination is achieved in human cells.\",\n      \"evidence\": \"Co-IP with Rad6-Rad18 and Mms2-Ubc13, in vitro ubiquitylation and active-site analysis, plus reciprocal Co-IP with knockdown phenotypes in a parallel study\",\n      \"pmids\": [\"17108083\", \"17130289\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define which domain mediates PCNA recognition\", \"Did not resolve substrate specificity versus other E3s\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Showed SHPRH is not the sole RAD5 homolog, revealing HLTF as a cooperating E3 in PCNA polyubiquitination and raising the question of functional redundancy versus specialization.\",\n      \"evidence\": \"Co-IP of HLTF with UBC13/PCNA/SHPRH, siRNA knockdown, mutagenesis and MEF chromosome-break assays\",\n      \"pmids\": [\"18719106\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism distinguishing SHPRH from HLTF unresolved at this stage\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Resolved how the two RAD5 homologs are coordinated, demonstrating damage-type-specific partitioning of postreplication-repair branches, and showed an additional E3 must exist for PCNA polyubiquitination.\",\n      \"evidence\": \"Co-IP, siRNA, TLS-polymerase recruitment and mutagenesis assays; independently, Shprh/Hltf double-knockout MEFs retaining PCNA-Ubn\",\n      \"pmids\": [\"21396873\", \"21269891\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of the alternative PCNA E3 ligase not determined\", \"Molecular trigger for damage-specific HLTF degradation not fully defined\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Determined the SHPRH PHD-domain fold and found it does not read H3K4-methylated peptides, redirecting the search for the domain's function.\",\n      \"evidence\": \"Solution NMR structure and NMR titrations with H3 peptides\",\n      \"pmids\": [\"23907177\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Positive binding partner of the PHD domain not identified in this study\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Uncovered a replication-independent nucleolar role, placing SHPRH at rDNA promoters to support mTOR-dependent rRNA transcription and assigning a function to its PHD domain.\",\n      \"evidence\": \"ChIP, Co-IP with RNA Pol I, shRNA, and pharmacological inhibition (rapamycin, actinomycin D)\",\n      \"pmids\": [\"28400511\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether E3 ligase activity is required for rRNA transcription not established\", \"Direct substrate at the rDNA promoter not identified\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Defined SHPRH's intrinsic biochemistry as a nucleosome-stimulated ATPase and broad nucleosome E3 ligase, indicating activity extends beyond PCNA and the Ubc13-Mms2 pair.\",\n      \"evidence\": \"Reconstituted ATPase and nucleosome-binding assays, 26-E2 screen, in vitro ubiquitylation and MS of ubiquitin linkages\",\n      \"pmids\": [\"31434570\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological nucleosomal substrates not identified\", \"In vivo relevance of branched linkages not tested\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified an upstream recruiter, TonEBP, that sequentially delivers SHPRH and then USP1 to bulky-adduct lesions, explaining how PCNA ubiquitination dynamics are timed.\",\n      \"evidence\": \"Co-IP, laser micro-irradiation live-cell recruitment, siRNA, PCNA-ubiquitination immunoblot\",\n      \"pmids\": [\"31376680\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab; reciprocal structural mapping of the SHPRH-TonEBP interface limited\", \"How the SHPRH-to-USP1 switch is triggered unresolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Linked SHPRH nuclear localization to a PCNA-interacting APIM motif and connected it to the CHK2 damage-response branch, broadening its regulatory reach.\",\n      \"evidence\": \"APIM mutagenesis with localization and SupF mutation-spectrum assays; CRISPR knockout with CHK2/MCM2 phosphorylation immunoblots\",\n      \"pmids\": [\"31973093\", \"32192191\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"How APIM and HIRAN-mediated PCNA contacts are coordinated unclear\", \"Whether CHK2 effect is direct or downstream of PCNA ubiquitination not resolved\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Extended SHPRH substrates and regulation beyond replication, showing KMT2B drives SHPRH expression and SHPRH ubiquitinates FYN for degradation.\",\n      \"evidence\": \"ChIP for H3K4me3 at the SHPRH promoter, Co-IP, ubiquitination assays and overexpression/knockdown\",\n      \"pmids\": [\"34606908\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct versus indirect ubiquitination of FYN not fully separated\", \"Single lab\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Demonstrated a conserved SHPRH-MLH1 interaction via a Rad5-derived MIP-box motif, implicating SHPRH in mismatch-repair-associated drug responses.\",\n      \"evidence\": \"Co-IP, yeast two-hybrid, bioinformatic MIP-box identification, siRNA with drug sensitivity\",\n      \"pmids\": [\"35784486\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of the MLH1 interaction for MMR mechanistically thin\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Provided the structural basis for SHPRH-nucleosome recognition, showing acidic-patch engagement and a non-canonical SF2 ATPase contact with nucleosomal DNA.\",\n      \"evidence\": \"Cryo-EM structure at 2.8 \\u00c5 with AlphaFold-Multimer prediction\",\n      \"pmids\": [\"38979307\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not capture a SHPRH-PCNA or SHPRH-E2 complex\", \"Catalytic cycle on a true substrate not visualized\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Established the direct biochemical mechanism of PCNA modification, showing SHPRH ubiquitinates unmodified PCNA at Lys164 via the Ube2D family in a HIRAN-dependent manner on free and DNA-bound PCNA.\",\n      \"evidence\": \"Reconstituted in vitro ubiquitylation, MS confirmation of Lys164, HIRAN domain mutagenesis/deletion\",\n      \"pmids\": [\"41990034\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo contribution of Ube2D versus Ubc13-Mms2 not quantified\", \"Regulation of HIRAN-PCNA engagement at forks not addressed\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The integration of SHPRH's replication-fork, nucleolar transcription, and non-PCNA substrate (FYN, MLH1) activities into a single regulatory logic remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking nucleosome-ATPase activity to PCNA versus rDNA functions\", \"Physiological nucleosomal ubiquitination substrates unidentified\", \"Determinants choosing Ube2D versus Ubc13-Mms2 E2 usage in cells unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [0, 1, 7, 14]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 7, 14, 12]},\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"GO:0042393\", \"supporting_discovery_ids\": [6]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [9]},\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"GO:0000228\", \"supporting_discovery_ids\": [7, 13]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [0, 1, 3]},\n      {\"term_id\": \"R-HSA-69306\", \"supporting_discovery_ids\": [1, 14]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 12]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [7, 13]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"PCNA\", \"RAD18\", \"UBC13\", \"HLTF\", \"MLH1\", \"TonEBP\", \"USP1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}