{"gene":"DPPA5","run_date":"2026-06-09T23:54:42","timeline":{"discoveries":[{"year":2014,"finding":"DPPA5 reduces endoplasmic reticulum (ER) stress and apoptosis in hematopoietic stem cells (HSCs): ectopic expression of Dppa5 followed by in vitro culture increased HSC reconstitution capacity, while knockdown of Dppa5 impaired long-term reconstitution ability due to elevated ER stress levels, establishing a direct mechanistic link between Dppa5 and ER stress regulation in HSCs.","method":"Ectopic overexpression and knockdown (loss-of-function) in HSCs with functional readouts of ER stress levels, apoptosis, and in vivo bone marrow reconstitution assays","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal gain- and loss-of-function experiments with defined cellular and in vivo phenotypes, supported by chemical chaperone rescue confirming ER stress as the mechanism","pmids":["24882002"],"is_preprint":false},{"year":2015,"finding":"DPPA5 directly interacts with NANOG protein and stabilizes it via a post-transcriptional mechanism, increasing NANOG protein levels without affecting NANOG mRNA, thereby enhancing pluripotency and reprogramming efficiency in human PSCs.","method":"Co-immunoprecipitation, protein stability assays, quantitative RT-PCR, DPPA5 overexpression in hPSCs with NANOG protein level measurement","journal":"Stem cells (Dayton, Ohio)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP plus protein stability assay plus qRT-PCR in single lab, orthogonal methods supporting post-transcriptional stabilization","pmids":["26661329"],"is_preprint":false},{"year":2024,"finding":"The ubiquitin E3 ligase FBXO9 targets DPPA5 for ubiquitylation and proteasomal degradation; FBXO9 silencing decreases proteasomal degradation of DPPA5, thereby facilitating induction of pluripotency, identifying FBXO9 as a writer of ubiquitin on DPPA5.","method":"RNAi screen during cellular reprogramming; FBXO9 silencing with measurement of DPPA5 protein levels and pluripotency induction efficiency","journal":"Stem cells (Dayton, Ohio)","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — single lab, RNAi-based loss-of-function with proteasomal degradation readout, no in vitro ubiquitylation reconstitution described in abstract","pmids":["38227647"],"is_preprint":false},{"year":2007,"finding":"DPPA5 encodes a protein with an atypical KH RNA-binding domain, shared with related family members (KHDC1a, KHDC1b, ECAT1, OOEP), establishing that DPPA5 belongs to a eutherian-specific gene family of KH-domain RNA-binding proteins expressed in oocytes and/or embryonic stem cells.","method":"Sequence/domain analysis combined with expression profiling (structural characterization of protein family)","journal":"Genomics","confidence":"Medium","confidence_rationale":"Tier 3 / Strong — domain architecture established by multiple sequence analyses replicated across species; functional RNA-binding activity not directly tested in this study","pmids":["17913455"],"is_preprint":false},{"year":2017,"finding":"DPPA5 protein is recruited to stress granules (SGs) in human induced pluripotent stem cells upon sodium arsenite or heat shock treatment, identifying DPPA5 as a novel component of SGs in hiPSCs.","method":"Immunofluorescence/protein localization analysis in hiPSCs under stress conditions (sodium arsenite, heat shock, hydrogen peroxide)","journal":"PloS one","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, single method (co-localization with known SG markers), no functional consequence of SG recruitment tested","pmids":["28746394"],"is_preprint":false},{"year":2008,"finding":"DPPA5 (ESG1) protein was identified in two isoforms (with and without N-terminal acetylation) in the nuclear proteome of mouse embryonic stem cells and embryonic germ cells, establishing its nuclear localization in pluripotent cells.","method":"Subcellular fractionation followed by 2D gel electrophoresis and MALDI-TOF-MS/nano-LC-MS/MS identification","journal":"Electrophoresis","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab proteomics identification; nuclear localization established but no functional consequence directly tested","pmids":["18449859"],"is_preprint":false},{"year":2006,"finding":"Targeted disruption of the mouse ESG1 (Dppa5) gene showed that ESG1-knockout mice develop normally and are fertile, and ESG1-/- ES cells show normal morphology, proliferation, and differentiation, demonstrating that ESG1/Dppa5 is dispensable for ES cell self-renewal and germ cell establishment despite its specific expression in these cells.","method":"Homologous recombination gene knockout in mouse ES cells and generation of knockout mice; Northern and Western blot confirmation of absence of ESG1; phenotypic characterization","journal":"BMC developmental biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean genetic knockout with multiple confirmatory methods (two independent homologous recombination events, blastocyst-derived KO ES cells), confirmed absence of protein, assessed multiple phenotypic readouts","pmids":["16504174"],"is_preprint":false},{"year":2007,"finding":"The DPPA5 promoter CpG island is densely methylated and the gene is silenced in normal somatic tissues, but is unmethylated and expressed in testis and sperm; treatment with a DNA demethylating agent or loss of DNMT1 and/or DNMT3b reactivated DPPA5 expression, establishing DNA methylation as the primary mechanism of tissue-specific silencing of DPPA5.","method":"Methylated CpG island amplification/microarray, bisulfite cloning and sequencing, treatment with DNA demethylating agent, analysis of DNMT1/DNMT3b-deficient cell lines","journal":"PLoS genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (bisulfite sequencing, demethylating agent, DNMT-deficient cells) in single study establishing epigenetic silencing mechanism","pmids":["17967063"],"is_preprint":false}],"current_model":"DPPA5 is a eutherian-specific KH-domain RNA-binding protein expressed in pluripotent stem cells and germ cells whose expression is controlled by promoter CpG methylation; it reduces endoplasmic reticulum stress to support hematopoietic stem cell reconstitution, stabilizes NANOG protein post-transcriptionally to promote pluripotency and reprogramming, is itself targeted for ubiquitylation and proteasomal degradation by the E3 ligase FBXO9, localizes to the nucleus and to stress granules under cellular stress, and is dispensable for mouse ES cell self-renewal and germ cell development under normal conditions."},"narrative":{"mechanistic_narrative":"DPPA5 is a eutherian-specific KH-domain RNA-binding protein restricted to pluripotent stem cells and germ cells, where it acts to buffer cellular stress and reinforce the pluripotency network [PMID:17913455, PMID:24882002]. In hematopoietic stem cells it lowers endoplasmic reticulum stress and apoptosis, with gain of function enhancing reconstitution capacity and knockdown impairing long-term reconstitution through elevated ER stress [PMID:24882002]. In pluripotent cells it physically binds NANOG and stabilizes the protein post-transcriptionally—raising NANOG protein without changing its mRNA—thereby promoting pluripotency and reprogramming efficiency [PMID:26661329]. DPPA5 abundance is itself controlled by the E3 ubiquitin ligase FBXO9, which targets it for proteasomal degradation; relieving this degradation facilitates induction of pluripotency [PMID:38227647]. Its tissue-restricted expression is enforced by promoter CpG island methylation, with silencing in somatic tissues reversed by demethylation or loss of DNMT1/DNMT3b [PMID:17967063]. Despite this expression pattern, genetic ablation in mouse shows DPPA5 is dispensable for ES cell self-renewal and germ cell development under normal conditions [PMID:16504174].","teleology":[{"year":2006,"claim":"Established whether DPPA5, despite its highly specific expression, is genetically required for the pluripotent and germ cell compartments where it is found.","evidence":"Homologous recombination knockout of mouse Esg1/Dppa5 in ES cells and mice with phenotypic characterization","pmids":["16504174"],"confidence":"High","gaps":["Does not test redundancy with paralogous KH-domain family members","No challenge or stress condition tested where loss might manifest a phenotype"]},{"year":2007,"claim":"Defined how DPPA5's restriction to germline/pluripotent tissues is enforced, identifying promoter CpG methylation as the silencing mechanism in soma.","evidence":"Bisulfite sequencing, demethylating agent, and DNMT1/DNMT3b-deficient cell analysis","pmids":["17967063"],"confidence":"Medium","gaps":["Does not identify the transcription factors driving expression in unmethylated states","Functional role of expressed protein not addressed"]},{"year":2007,"claim":"Placed DPPA5 in a defined molecular class by showing it carries an atypical KH RNA-binding domain shared across a eutherian-specific oocyte/ES-cell gene family.","evidence":"Sequence and domain analysis with expression profiling across family members","pmids":["17913455"],"confidence":"Medium","gaps":["RNA-binding activity not directly demonstrated","No RNA targets identified"]},{"year":2008,"claim":"Localized the endogenous protein, establishing nuclear residence in pluripotent cells and the existence of N-terminally acetylated and unmodified isoforms.","evidence":"Subcellular fractionation with 2D gel and mass spectrometry of mouse ES/EG cell nuclear proteome","pmids":["18449859"],"confidence":"Low","gaps":["Single-lab proteomics without functional follow-up","Significance of acetylated isoform unknown","Does not exclude cytoplasmic pools"]},{"year":2014,"claim":"Assigned a cellular function by linking DPPA5 to ER stress control and stem cell maintenance beyond the pluripotent compartment.","evidence":"Reciprocal overexpression/knockdown in HSCs with ER stress, apoptosis, and in vivo reconstitution readouts plus chemical chaperone rescue","pmids":["24882002"],"confidence":"High","gaps":["Molecular mechanism connecting DPPA5 to the unfolded protein response not resolved","Direct RNA or protein effectors in the ER stress pathway not identified"]},{"year":2015,"claim":"Identified a direct molecular partner and mechanism, showing DPPA5 stabilizes NANOG protein to reinforce pluripotency.","evidence":"Co-immunoprecipitation, protein stability assays, and qRT-PCR in human PSCs with reprogramming readouts","pmids":["26661329"],"confidence":"Medium","gaps":["Domain mediating the DPPA5-NANOG interaction not mapped","Mechanism of stabilization (e.g., blocking degradation) not defined","Reciprocal validation in additional systems absent"]},{"year":2017,"claim":"Extended DPPA5 localization to a stress-responsive compartment, identifying it as a stress granule component upon oxidative and heat stress.","evidence":"Immunofluorescence co-localization with SG markers in hiPSCs under arsenite and heat shock","pmids":["28746394"],"confidence":"Low","gaps":["Single co-localization method without functional consequence tested","Relationship to its ER stress role unclear"]},{"year":2024,"claim":"Defined how DPPA5 protein levels are regulated, identifying FBXO9-mediated ubiquitylation and proteasomal turnover as a control point during reprogramming.","evidence":"RNAi screen during reprogramming with FBXO9 silencing and DPPA5 protein/pluripotency readouts","pmids":["38227647"],"confidence":"Medium","gaps":["No in vitro ubiquitylation reconstitution to prove direct ligase activity","Ubiquitylation site on DPPA5 not mapped"]},{"year":null,"claim":"Whether DPPA5's RNA-binding KH domain has bona fide RNA targets that mechanistically connect its roles in ER stress buffering, NANOG stabilization, and stress granule recruitment remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No direct RNA targets identified","Unifying biochemical mechanism across phenotypes not established","Whether nuclear, stress granule, and stabilizing functions reflect distinct activities is unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[3]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[5]}],"pathway":[{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[0,4]}],"complexes":[],"partners":["NANOG","FBXO9"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"A6NC42","full_name":"Developmental pluripotency-associated 5 protein","aliases":["Embryonal stem cell-specific gene 1 protein","ESG-1"],"length_aa":116,"mass_kda":13.5,"function":"Involved in the maintenance of embryonic stem (ES) cell pluripotency. Dispensable for self-renewal of pluripotent ES cells and establishment of germ cells. Associates with specific target mRNAs (By similarity)","subcellular_location":"Cytoplasm","url":"https://www.uniprot.org/uniprotkb/A6NC42/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/DPPA5","classification":"Not Classified","n_dependent_lines":228,"n_total_lines":1208,"dependency_fraction":0.18874172185430463},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/DPPA5","total_profiled":1310},"omim":[{"mim_id":"611689","title":"OOCYTE-EXPRESSED PROTEIN; OOEP","url":"https://www.omim.org/entry/611689"},{"mim_id":"611688","title":"KH DOMAIN-CONTAINING PROTEIN 1; KHDC1","url":"https://www.omim.org/entry/611688"},{"mim_id":"611687","title":"KHDC3-LIKE PROTEIN, SUBCORTICAL MATERNAL COMPLEX MEMBER; KHDC3L","url":"https://www.omim.org/entry/611687"},{"mim_id":"611111","title":"DEVELOPMENTAL PLURIPOTENCY-ASSOCIATED GENE 5; DPPA5","url":"https://www.omim.org/entry/611111"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Uncertain","locations":[{"location":"Cell Junctions","reliability":"Uncertain"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in single","driving_tissues":[{"tissue":"testis","ntpm":2.0}],"url":"https://www.proteinatlas.org/search/DPPA5"},"hgnc":{"alias_symbol":["Esg1"],"prev_symbol":[]},"alphafold":{"accession":"A6NC42","domains":[{"cath_id":"3.30.1370.10","chopping":"17-108","consensus_level":"high","plddt":92.6973,"start":17,"end":108}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/A6NC42","model_url":"https://alphafold.ebi.ac.uk/files/AF-A6NC42-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-A6NC42-F1-predicted_aligned_error_v6.png","plddt_mean":88.44},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=DPPA5","jax_strain_url":"https://www.jax.org/strain/search?query=DPPA5"},"sequence":{"accession":"A6NC42","fasta_url":"https://rest.uniprot.org/uniprotkb/A6NC42.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/A6NC42/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/A6NC42"}},"corpus_meta":[{"pmid":"17967063","id":"PMC_17967063","title":"Genome-wide profiling of DNA methylation reveals a class of normally methylated CpG island promoters.","date":"2007","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/17967063","citation_count":275,"is_preprint":false},{"pmid":"16081659","id":"PMC_16081659","title":"Primary differentiation in the human blastocyst: comparative molecular portraits of inner cell mass and trophectoderm cells.","date":"2005","source":"Stem cells (Dayton, Ohio)","url":"https://pubmed.ncbi.nlm.nih.gov/16081659","citation_count":179,"is_preprint":false},{"pmid":"18502893","id":"PMC_18502893","title":"Generation of multipotent cell lines from a distinct population of male germ line stem cells.","date":"2008","source":"Reproduction (Cambridge, England)","url":"https://pubmed.ncbi.nlm.nih.gov/18502893","citation_count":94,"is_preprint":false},{"pmid":"16479162","id":"PMC_16479162","title":"Diverse epigenetic profile of novel human embryonic stem cell lines.","date":"2006","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/16479162","citation_count":75,"is_preprint":false},{"pmid":"24882002","id":"PMC_24882002","title":"Dppa5 improves hematopoietic stem cell activity by reducing endoplasmic reticulum stress.","date":"2014","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/24882002","citation_count":69,"is_preprint":false},{"pmid":"17913455","id":"PMC_17913455","title":"Atypical structure and phylogenomic evolution of the new eutherian oocyte- and embryo-expressed KHDC1/DPPA5/ECAT1/OOEP gene family.","date":"2007","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/17913455","citation_count":66,"is_preprint":false},{"pmid":"20146561","id":"PMC_20146561","title":"Generation of induced pluripotent stem cells from human adipose-derived stem cells without c-MYC.","date":"2010","source":"Tissue engineering. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/20146561","citation_count":65,"is_preprint":false},{"pmid":"22989381","id":"PMC_22989381","title":"Derivation and characterization of sleeping beauty transposon-mediated porcine induced pluripotent stem cells.","date":"2012","source":"Stem cells and development","url":"https://pubmed.ncbi.nlm.nih.gov/22989381","citation_count":62,"is_preprint":false},{"pmid":"27173828","id":"PMC_27173828","title":"Derivation of Porcine Embryonic Stem-Like Cells from In Vitro-Produced Blastocyst-Stage Embryos.","date":"2016","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/27173828","citation_count":49,"is_preprint":false},{"pmid":"23016947","id":"PMC_23016947","title":"Growth requirements and chromosomal instability of induced pluripotent stem cells generated from adult canine fibroblasts.","date":"2012","source":"Stem cells and development","url":"https://pubmed.ncbi.nlm.nih.gov/23016947","citation_count":48,"is_preprint":false},{"pmid":"19221495","id":"PMC_19221495","title":"The role of promoter CpG methylation in the epigenetic control of stem cell related genes during differentiation.","date":"2009","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/19221495","citation_count":47,"is_preprint":false},{"pmid":"25338022","id":"PMC_25338022","title":"High-throughput screening of dipeptide utilization mediated by the ABC transporter DppBCDF and its substrate-binding proteins DppA1-A5 in Pseudomonas aeruginosa.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/25338022","citation_count":46,"is_preprint":false},{"pmid":"16920069","id":"PMC_16920069","title":"Analysis of nuclear reprogramming in cloned miniature pig embryos by expression of Oct-4 and Oct-4 related genes.","date":"2006","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/16920069","citation_count":45,"is_preprint":false},{"pmid":"26575221","id":"PMC_26575221","title":"Brain-specific epigenetic markers of schizophrenia.","date":"2015","source":"Translational psychiatry","url":"https://pubmed.ncbi.nlm.nih.gov/26575221","citation_count":43,"is_preprint":false},{"pmid":"24465482","id":"PMC_24465482","title":"Efficient reprogramming of naïve-like induced pluripotent stem cells from porcine adipose-derived stem cells with a feeder-independent and serum-free system.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/24465482","citation_count":42,"is_preprint":false},{"pmid":"15790765","id":"PMC_15790765","title":"Identification of developmental pluripotency associated 5 expression in human pluripotent stem cells.","date":"2005","source":"Stem cells (Dayton, Ohio)","url":"https://pubmed.ncbi.nlm.nih.gov/15790765","citation_count":37,"is_preprint":false},{"pmid":"25515008","id":"PMC_25515008","title":"Establishment of a primed pluripotent epiblast stem cell in FGF4-based conditions.","date":"2014","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/25515008","citation_count":35,"is_preprint":false},{"pmid":"18449859","id":"PMC_18449859","title":"Nuclear proteome analysis of undifferentiated mouse embryonic stem and germ cells.","date":"2008","source":"Electrophoresis","url":"https://pubmed.ncbi.nlm.nih.gov/18449859","citation_count":33,"is_preprint":false},{"pmid":"16504174","id":"PMC_16504174","title":"Identification and targeted disruption of the mouse gene encoding ESG1 (PH34/ECAT2/DPPA5).","date":"2006","source":"BMC developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/16504174","citation_count":32,"is_preprint":false},{"pmid":"19129511","id":"PMC_19129511","title":"Gene birth, death, and divergence: the different scenarios of reproduction-related gene evolution.","date":"2009","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/19129511","citation_count":29,"is_preprint":false},{"pmid":"28746394","id":"PMC_28746394","title":"Effects of oxidative and thermal stresses on stress granule formation in human induced pluripotent stem cells.","date":"2017","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/28746394","citation_count":26,"is_preprint":false},{"pmid":"26661329","id":"PMC_26661329","title":"DPPA5 Supports Pluripotency and Reprogramming by Regulating NANOG Turnover.","date":"2015","source":"Stem cells (Dayton, Ohio)","url":"https://pubmed.ncbi.nlm.nih.gov/26661329","citation_count":24,"is_preprint":false},{"pmid":"21337383","id":"PMC_21337383","title":"Co-culture of mesenchymal-like stromal cells derived from human foreskin permits long term propagation and differentiation of human embryonic stem cells.","date":"2011","source":"Journal of cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/21337383","citation_count":21,"is_preprint":false},{"pmid":"31210434","id":"PMC_31210434","title":"Pluripotency Potential of Embryonic Stem Cell-Like Cells Derived from Mouse Testis.","date":"2019","source":"Cell journal","url":"https://pubmed.ncbi.nlm.nih.gov/31210434","citation_count":17,"is_preprint":false},{"pmid":"19813265","id":"PMC_19813265","title":"Post-fusion treatment with MG132 increases transcription factor expression in somatic cell nuclear transfer embryos in pigs.","date":"2010","source":"Molecular reproduction and development","url":"https://pubmed.ncbi.nlm.nih.gov/19813265","citation_count":17,"is_preprint":false},{"pmid":"29927686","id":"PMC_29927686","title":"Loci-specific differences in blood DNA methylation in HBV-negative populations at risk for hepatocellular carcinoma development.","date":"2018","source":"Epigenetics","url":"https://pubmed.ncbi.nlm.nih.gov/29927686","citation_count":15,"is_preprint":false},{"pmid":"22775411","id":"PMC_22775411","title":"Recombinant rabbit leukemia inhibitory factor and rabbit embryonic fibroblasts support the derivation and maintenance of rabbit embryonic stem cells.","date":"2012","source":"Cellular reprogramming","url":"https://pubmed.ncbi.nlm.nih.gov/22775411","citation_count":14,"is_preprint":false},{"pmid":"26053518","id":"PMC_26053518","title":"Stem Cell-Derived Bioactive Materials Accelerate Development of Porcine In Vitro-Fertilized Embryos.","date":"2015","source":"Cellular reprogramming","url":"https://pubmed.ncbi.nlm.nih.gov/26053518","citation_count":12,"is_preprint":false},{"pmid":"32424450","id":"PMC_32424450","title":"Conversion between porcine naïve-like and primed ESCs and specific pluripotency marker identification.","date":"2020","source":"In vitro cellular & developmental biology. Animal","url":"https://pubmed.ncbi.nlm.nih.gov/32424450","citation_count":6,"is_preprint":false},{"pmid":"22532788","id":"PMC_22532788","title":"Data mining in networks of differentially expressed genes during sow pregnancy.","date":"2012","source":"International journal of biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/22532788","citation_count":5,"is_preprint":false},{"pmid":"38227647","id":"PMC_38227647","title":"Ubiquitin E3 Ligase FBXO9 Regulates Pluripotency by Targeting DPPA5 for Ubiquitylation and Degradation.","date":"2024","source":"Stem cells (Dayton, Ohio)","url":"https://pubmed.ncbi.nlm.nih.gov/38227647","citation_count":1,"is_preprint":false},{"pmid":"41436486","id":"PMC_41436486","title":"The solute-binding proteins DppA1-5 of Pseudomonas aeruginosa have distinct substrate profiles.","date":"2025","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/41436486","citation_count":1,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":18159,"output_tokens":2053,"usd":0.042636,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9068,"output_tokens":2500,"usd":0.05392,"stage2_stop_reason":"end_turn"},"total_usd":0.096556,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2014,\n      \"finding\": \"DPPA5 reduces endoplasmic reticulum (ER) stress and apoptosis in hematopoietic stem cells (HSCs): ectopic expression of Dppa5 followed by in vitro culture increased HSC reconstitution capacity, while knockdown of Dppa5 impaired long-term reconstitution ability due to elevated ER stress levels, establishing a direct mechanistic link between Dppa5 and ER stress regulation in HSCs.\",\n      \"method\": \"Ectopic overexpression and knockdown (loss-of-function) in HSCs with functional readouts of ER stress levels, apoptosis, and in vivo bone marrow reconstitution assays\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal gain- and loss-of-function experiments with defined cellular and in vivo phenotypes, supported by chemical chaperone rescue confirming ER stress as the mechanism\",\n      \"pmids\": [\"24882002\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"DPPA5 directly interacts with NANOG protein and stabilizes it via a post-transcriptional mechanism, increasing NANOG protein levels without affecting NANOG mRNA, thereby enhancing pluripotency and reprogramming efficiency in human PSCs.\",\n      \"method\": \"Co-immunoprecipitation, protein stability assays, quantitative RT-PCR, DPPA5 overexpression in hPSCs with NANOG protein level measurement\",\n      \"journal\": \"Stem cells (Dayton, Ohio)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP plus protein stability assay plus qRT-PCR in single lab, orthogonal methods supporting post-transcriptional stabilization\",\n      \"pmids\": [\"26661329\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The ubiquitin E3 ligase FBXO9 targets DPPA5 for ubiquitylation and proteasomal degradation; FBXO9 silencing decreases proteasomal degradation of DPPA5, thereby facilitating induction of pluripotency, identifying FBXO9 as a writer of ubiquitin on DPPA5.\",\n      \"method\": \"RNAi screen during cellular reprogramming; FBXO9 silencing with measurement of DPPA5 protein levels and pluripotency induction efficiency\",\n      \"journal\": \"Stem cells (Dayton, Ohio)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — single lab, RNAi-based loss-of-function with proteasomal degradation readout, no in vitro ubiquitylation reconstitution described in abstract\",\n      \"pmids\": [\"38227647\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"DPPA5 encodes a protein with an atypical KH RNA-binding domain, shared with related family members (KHDC1a, KHDC1b, ECAT1, OOEP), establishing that DPPA5 belongs to a eutherian-specific gene family of KH-domain RNA-binding proteins expressed in oocytes and/or embryonic stem cells.\",\n      \"method\": \"Sequence/domain analysis combined with expression profiling (structural characterization of protein family)\",\n      \"journal\": \"Genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Strong — domain architecture established by multiple sequence analyses replicated across species; functional RNA-binding activity not directly tested in this study\",\n      \"pmids\": [\"17913455\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"DPPA5 protein is recruited to stress granules (SGs) in human induced pluripotent stem cells upon sodium arsenite or heat shock treatment, identifying DPPA5 as a novel component of SGs in hiPSCs.\",\n      \"method\": \"Immunofluorescence/protein localization analysis in hiPSCs under stress conditions (sodium arsenite, heat shock, hydrogen peroxide)\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, single method (co-localization with known SG markers), no functional consequence of SG recruitment tested\",\n      \"pmids\": [\"28746394\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"DPPA5 (ESG1) protein was identified in two isoforms (with and without N-terminal acetylation) in the nuclear proteome of mouse embryonic stem cells and embryonic germ cells, establishing its nuclear localization in pluripotent cells.\",\n      \"method\": \"Subcellular fractionation followed by 2D gel electrophoresis and MALDI-TOF-MS/nano-LC-MS/MS identification\",\n      \"journal\": \"Electrophoresis\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab proteomics identification; nuclear localization established but no functional consequence directly tested\",\n      \"pmids\": [\"18449859\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Targeted disruption of the mouse ESG1 (Dppa5) gene showed that ESG1-knockout mice develop normally and are fertile, and ESG1-/- ES cells show normal morphology, proliferation, and differentiation, demonstrating that ESG1/Dppa5 is dispensable for ES cell self-renewal and germ cell establishment despite its specific expression in these cells.\",\n      \"method\": \"Homologous recombination gene knockout in mouse ES cells and generation of knockout mice; Northern and Western blot confirmation of absence of ESG1; phenotypic characterization\",\n      \"journal\": \"BMC developmental biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean genetic knockout with multiple confirmatory methods (two independent homologous recombination events, blastocyst-derived KO ES cells), confirmed absence of protein, assessed multiple phenotypic readouts\",\n      \"pmids\": [\"16504174\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"The DPPA5 promoter CpG island is densely methylated and the gene is silenced in normal somatic tissues, but is unmethylated and expressed in testis and sperm; treatment with a DNA demethylating agent or loss of DNMT1 and/or DNMT3b reactivated DPPA5 expression, establishing DNA methylation as the primary mechanism of tissue-specific silencing of DPPA5.\",\n      \"method\": \"Methylated CpG island amplification/microarray, bisulfite cloning and sequencing, treatment with DNA demethylating agent, analysis of DNMT1/DNMT3b-deficient cell lines\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (bisulfite sequencing, demethylating agent, DNMT-deficient cells) in single study establishing epigenetic silencing mechanism\",\n      \"pmids\": [\"17967063\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"DPPA5 is a eutherian-specific KH-domain RNA-binding protein expressed in pluripotent stem cells and germ cells whose expression is controlled by promoter CpG methylation; it reduces endoplasmic reticulum stress to support hematopoietic stem cell reconstitution, stabilizes NANOG protein post-transcriptionally to promote pluripotency and reprogramming, is itself targeted for ubiquitylation and proteasomal degradation by the E3 ligase FBXO9, localizes to the nucleus and to stress granules under cellular stress, and is dispensable for mouse ES cell self-renewal and germ cell development under normal conditions.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"DPPA5 is a eutherian-specific KH-domain RNA-binding protein restricted to pluripotent stem cells and germ cells, where it acts to buffer cellular stress and reinforce the pluripotency network [#3, #0]. In hematopoietic stem cells it lowers endoplasmic reticulum stress and apoptosis, with gain of function enhancing reconstitution capacity and knockdown impairing long-term reconstitution through elevated ER stress [#0]. In pluripotent cells it physically binds NANOG and stabilizes the protein post-transcriptionally—raising NANOG protein without changing its mRNA—thereby promoting pluripotency and reprogramming efficiency [#1]. DPPA5 abundance is itself controlled by the E3 ubiquitin ligase FBXO9, which targets it for proteasomal degradation; relieving this degradation facilitates induction of pluripotency [#2]. Its tissue-restricted expression is enforced by promoter CpG island methylation, with silencing in somatic tissues reversed by demethylation or loss of DNMT1/DNMT3b [#7]. Despite this expression pattern, genetic ablation in mouse shows DPPA5 is dispensable for ES cell self-renewal and germ cell development under normal conditions [#6].\",\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Established whether DPPA5, despite its highly specific expression, is genetically required for the pluripotent and germ cell compartments where it is found.\",\n      \"evidence\": \"Homologous recombination knockout of mouse Esg1/Dppa5 in ES cells and mice with phenotypic characterization\",\n      \"pmids\": [\"16504174\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not test redundancy with paralogous KH-domain family members\", \"No challenge or stress condition tested where loss might manifest a phenotype\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Defined how DPPA5's restriction to germline/pluripotent tissues is enforced, identifying promoter CpG methylation as the silencing mechanism in soma.\",\n      \"evidence\": \"Bisulfite sequencing, demethylating agent, and DNMT1/DNMT3b-deficient cell analysis\",\n      \"pmids\": [\"17967063\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not identify the transcription factors driving expression in unmethylated states\", \"Functional role of expressed protein not addressed\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Placed DPPA5 in a defined molecular class by showing it carries an atypical KH RNA-binding domain shared across a eutherian-specific oocyte/ES-cell gene family.\",\n      \"evidence\": \"Sequence and domain analysis with expression profiling across family members\",\n      \"pmids\": [\"17913455\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"RNA-binding activity not directly demonstrated\", \"No RNA targets identified\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Localized the endogenous protein, establishing nuclear residence in pluripotent cells and the existence of N-terminally acetylated and unmodified isoforms.\",\n      \"evidence\": \"Subcellular fractionation with 2D gel and mass spectrometry of mouse ES/EG cell nuclear proteome\",\n      \"pmids\": [\"18449859\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single-lab proteomics without functional follow-up\", \"Significance of acetylated isoform unknown\", \"Does not exclude cytoplasmic pools\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Assigned a cellular function by linking DPPA5 to ER stress control and stem cell maintenance beyond the pluripotent compartment.\",\n      \"evidence\": \"Reciprocal overexpression/knockdown in HSCs with ER stress, apoptosis, and in vivo reconstitution readouts plus chemical chaperone rescue\",\n      \"pmids\": [\"24882002\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism connecting DPPA5 to the unfolded protein response not resolved\", \"Direct RNA or protein effectors in the ER stress pathway not identified\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identified a direct molecular partner and mechanism, showing DPPA5 stabilizes NANOG protein to reinforce pluripotency.\",\n      \"evidence\": \"Co-immunoprecipitation, protein stability assays, and qRT-PCR in human PSCs with reprogramming readouts\",\n      \"pmids\": [\"26661329\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Domain mediating the DPPA5-NANOG interaction not mapped\", \"Mechanism of stabilization (e.g., blocking degradation) not defined\", \"Reciprocal validation in additional systems absent\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Extended DPPA5 localization to a stress-responsive compartment, identifying it as a stress granule component upon oxidative and heat stress.\",\n      \"evidence\": \"Immunofluorescence co-localization with SG markers in hiPSCs under arsenite and heat shock\",\n      \"pmids\": [\"28746394\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single co-localization method without functional consequence tested\", \"Relationship to its ER stress role unclear\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined how DPPA5 protein levels are regulated, identifying FBXO9-mediated ubiquitylation and proteasomal turnover as a control point during reprogramming.\",\n      \"evidence\": \"RNAi screen during reprogramming with FBXO9 silencing and DPPA5 protein/pluripotency readouts\",\n      \"pmids\": [\"38227647\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No in vitro ubiquitylation reconstitution to prove direct ligase activity\", \"Ubiquitylation site on DPPA5 not mapped\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Whether DPPA5's RNA-binding KH domain has bona fide RNA targets that mechanistically connect its roles in ER stress buffering, NANOG stabilization, and stress granule recruitment remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No direct RNA targets identified\", \"Unifying biochemical mechanism across phenotypes not established\", \"Whether nuclear, stress granule, and stabilizing functions reflect distinct activities is unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [0, 4]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"NANOG\", \"FBXO9\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}