{"gene":"CTPS1","run_date":"2026-06-09T22:57:19","timeline":{"discoveries":[{"year":2021,"finding":"Cryo-EM structures reveal that CTP regulates both CTPS1 and CTPS2 by binding in two sites that clash with substrates (product feedback inhibition). CTPS1 is less sensitive to CTP feedback inhibition than CTPS2, consistent with its role in expanding CTP pools during lymphocyte proliferation. Small-molecule inhibitors that are CTPS1-selective or non-selective mimic CTP binding at one inhibitory site, with a single amino acid substitution explaining isoform selectivity. Both inhibitors bind to CTPS1 assembled into large-scale filaments, which represent a hyperactive form of the enzyme.","method":"Cryo-electron microscopy (cryo-EM) structures of inhibitor-bound and CTP-bound CTPS1 filaments; biochemical inhibition assays; site-directed mutagenesis (amino acid substitution analysis); primary T cell proliferation assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structural determination combined with biochemical assays and mutagenesis in a single rigorous study, with functional validation in primary T cells","pmids":["34583994"],"is_preprint":false},{"year":2020,"finding":"CTPS1 deficiency caused by homozygous frameshift splice mutation (c.1692-1G>C, p.T566Dfs26X) results in 80–90% reduction of CTPS protein and CTP synthetase activity in patient lymphocytes, leading to severely impaired T cell proliferation and IL-2 secretion upon TCR activation. The mutant protein (T566Dfs26X) retains normal CTPS enzymatic activity when expressed at wild-type levels; loss of function is entirely attributable to protein instability. Inactivation of CTPS1 in a T cell leukemia line fully abolished proliferation, confirming CTPS1 is required for T cell proliferative responses.","method":"CTPS enzymatic activity assay in patient cells; genetic complementation in CTPS1-deficient leukemia cells with WT and mutant constructs; immune phenotyping; T cell proliferation and IL-2 secretion assays","journal":"JCI insight","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — enzymatic activity assay combined with genetic complementation and loss-of-function in human patient cells and leukemia cell line, multiple orthogonal methods","pmids":["32161190"],"is_preprint":false},{"year":2023,"finding":"CTPS1 has higher intrinsic CTP synthetase enzymatic activity than CTPS2 and is more resistant to inhibition by the UTP analog 3-deaza-uridine. Using inactivation and complementation experiments, CTPS1 was shown to be the primary contributor to cell proliferation; CTPS2 contributes modestly when CTPS1 is present but is essential in its absence.","method":"CTPS1 and/or CTPS2 inactivation by KO; complementation experiments; in vitro enzymatic activity assays; 3-deaza-uridine inhibition assays; analysis of >1,000 cancer cell line datasets","journal":"Life science alliance","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — enzymatic activity assays plus KO/complementation with multiple orthogonal methods and large-scale dataset validation","pmids":["37348953"],"is_preprint":false},{"year":2022,"finding":"Inhibition of CTPS1 (but not CTPS2) selectively induces DNA replication stress in MYC-overexpressing cancer cells. MYC-driven rRNA synthesis causes selective replication stress upon CTPS1 inhibition. Combined inhibition of CTPS1 and ATR is synthetically lethal in MYC-overexpressing cells in vitro and decreases tumor growth in vivo.","method":"CTPS1/CTPS2-selective knockdown/inhibition; replication stress assays; cell viability assays; in vivo xenograft tumor models; ATR inhibitor combination studies","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 2 / Strong — isoform-specific KD with defined molecular phenotype (replication stress), epistatic combination with ATR inhibitor, both in vitro and in vivo validation","pmids":["35022212"],"is_preprint":false},{"year":2025,"finding":"CTPS1 and CTPS2 directly interact with each other independently of polymerization and cytoophidium formation, forming heterocomplexes. When associated with CTPS2, CTPS1 enzymatic activity is decreased and becomes more sensitive to CTP product feedback inhibition, demonstrating that CTPS2 modulates CTPS1 activity through heterocomplex formation. CTPS2-containing filaments (cytoophidia) are dependent on CTPS1 expression. CTPS1H355A and CTPS2H355A mutants unable to form cytoophidia can still sustain normal cell proliferation, showing cytoophidia are not required for proliferation.","method":"Co-immunoprecipitation; enzymatic activity assays with purified complexes; CTPS1/CTPS2 KO and complementation; H355A cytoophidium-deficient mutant analysis; co-localization imaging","journal":"Life science alliance","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — direct interaction confirmed by Co-IP, enzymatic activity measured for complexes, multiple mutant analyses and orthogonal methods in one study","pmids":["40957650"],"is_preprint":false},{"year":2025,"finding":"CTP (the enzymatic product of CTPS1) acts as a key regulator driving hCTPS1 filamentation. Cryo-EM structures of CTP-bound hCTPS1 filaments reveal the molecular details of CTP binding in filament assembly. CTP generated from the enzymatic reaction does not trigger filament disassembly. Two distinct CTP-binding pockets mediate this filamentation, and the mechanism is evolutionarily conserved across eukaryotic CTPS.","method":"Cryo-electron microscopy (cryo-EM) of CTP-bound hCTPS1 filaments; biochemical filamentation assays; analysis of CTP-binding pocket mutants","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — cryo-EM structure with biochemical validation, single lab, preprint not yet peer-reviewed","pmids":["bio_10.1101_2025.02.22.639624"],"is_preprint":true},{"year":2024,"finding":"RASD2 promotes SUMOylation of CTPS1 and inhibits its ubiquitination, thereby increasing CTPS1 protein stability. This was demonstrated by co-immunoprecipitation and immunoprecipitation-mass spectrometry in endometriosis cells, where histone lactylation upregulates RASD2, which in turn stabilizes CTPS1 via the SUMOylation/ubiquitination balance.","method":"Co-immunoprecipitation (Co-IP); immunoprecipitation-mass spectrometry (IP-MS); Western blot for ubiquitination and SUMOylation; ChIP-qPCR; in vitro and in vivo endometriosis models","journal":"American journal of physiology. Cell physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP and IP-MS for interaction, SUMOylation/ubiquitination assays, single lab with two orthogonal biochemical methods","pmids":["39672102"],"is_preprint":false},{"year":2021,"finding":"CTPS interacts with ATP synthase (ATPS) and maintains ATP content during early decidualization (Day 3). At Day 6 of decidualization, CTPS instead associates with mitochondrial stress protein STRESS-70, correlating with reduced ATP concentration. CTPS localization shifts from cytoplasm (Day 3) to both cytoplasm and nucleus (Day 6), suggesting subcellular redistribution is linked to metabolic function during the decidualization process.","method":"Co-immunoprecipitation coupled with mass spectrometry; subcellular fractionation/immunofluorescence localization; CTPS knockdown by siRNA; in vitro decidualization model","journal":"Journal of cellular physiology","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP/MS identified CTPS-ATPS and CTPS-STRESS-70 interactions, functional ATP measurements, subcellular localization with functional consequences, single lab","pmids":["33576499"],"is_preprint":false},{"year":2024,"finding":"CTPS1 upregulates expression of choline/ethanolamine phosphotransferase 1 (CEPT1) by increasing CTP availability, thereby reprogramming glycerophospholipid metabolism. Glycerophospholipids synthesized by CEPT1 maintain mitochondrial homeostasis and promote BNIP3-mediated mitophagy, driving DLBCL progression.","method":"CTPS1 knockout/knockdown; single-cell RNA sequencing; Western blot and gene expression analysis; selective CTPS1 inhibitor (R80); functional mitophagy assays","journal":"Redox biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KO with defined molecular pathway (CEPT1/phospholipid/BNIP3), pharmacological inhibitor corroboration, single lab","pmids":["41865720"],"is_preprint":false},{"year":2025,"finding":"INHBA interacts with CTPS1 and competitively inhibits SMURF1 (SMAD Specific E3 Ubiquitin Protein Ligase 1)-mediated ubiquitination of CTPS1, thereby enhancing CTPS1 protein stability and promoting pyrimidine metabolism and gemcitabine resistance in pancreatic cancer cells.","method":"Immunoprecipitation mass spectrometry to identify CTPS1 as INHBA binding partner; Co-IP; in vitro ubiquitination assay; in vivo xenograft model","journal":"Cancer cell international","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — IP-MS plus Co-IP for interaction, ubiquitination assay, single lab with two orthogonal methods","pmids":["41239468"],"is_preprint":false},{"year":2022,"finding":"YBX1 (Y-box binding protein 1) directly binds to the CTPS1 promoter and activates its transcription, as demonstrated by dual luciferase reporter assays and chromatin immunoprecipitation (ChIP). Rescue experiments confirmed that enhanced cell proliferation and invasion driven by YBX1 overexpression could be reversed by CTPS1 knockdown, placing YBX1 upstream of CTPS1 in TNBC.","method":"Dual luciferase reporter assay; chromatin immunoprecipitation (ChIP); CTPS1 knockdown; YBX1 overexpression rescue experiment","journal":"Journal of translational medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP and luciferase reporter establish direct transcriptional regulation, epistasis via rescue experiment, single lab","pmids":["34991621"],"is_preprint":false},{"year":2023,"finding":"CTPS1 pharmacological inhibition by STP-B in mantle cell lymphoma causes rapid cell cycle arrest in early S-phase, inhibition of translation including anti-apoptotic MCL1 protein, and synergistic cell death in combination with the BCL2 inhibitor venetoclax, both in vitro and in vivo.","method":"Selective CTPS1 inhibitor STP-B; flow cytometry cell cycle analysis; Western blot for MCL1; in vitro and in vivo MCL models; BCL2 inhibitor combination assay","journal":"Haematologica","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological inhibition with defined molecular consequence (MCL1 suppression, S-phase arrest), in vivo validation, single lab","pmids":["38385294"],"is_preprint":false},{"year":2023,"finding":"CTPS1 knockout in multiple myeloma cells induces apoptosis and S-phase arrest with DNA double-strand breaks. Pharmacological inhibition of CTPS1 by STP-B activates DNA damage response (DDR) pathways. Combined inhibition of CTPS1 with ATR, CHEK1, or WEE1 inhibitors results in synergistic growth inhibition and early apoptosis.","method":"CTPS1 knockout; selective CTPS1 inhibitor STP-B; flow cytometry; DNA damage marker assays (γH2AX); DDR pathway inhibitor combinations; apoptosis assays","journal":"Leukemia","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KO and pharmacological inhibition with defined molecular phenotype (DDR activation, DSBs), DDR pathway epistasis, single lab","pmids":["37898670"],"is_preprint":false},{"year":2024,"finding":"Single-molecule fluorescence imaging demonstrated that CTPS self-assembles with tetramers as the minimum structural unit driving cytoophidium formation. CTPS acts as the nucleation core to induce subsequent growth of P5CS (Δ1-pyrroline-5-carboxylate synthase) filaments, constituting a coassembly (coordinated assembly) process in vitro.","method":"Single-molecule fluorescence imaging; single-molecule photobleach counting for stoichiometry; oligomer state distribution analysis under different conditions; in vitro reconstitution of CTPS and P5CS coassembly","journal":"The journal of physical chemistry. B","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with single-molecule imaging and stoichiometry, single lab but multiple orthogonal approaches","pmids":["38236746"],"is_preprint":false},{"year":2025,"finding":"HSPD1 (HSP60) interacts with CTPS protein; interference with Hspd1 gene expression inhibits CTPS cytoophidium formation even when CTPS is overexpressed, and this inhibits C2C12 skeletal muscle cell proliferation. The cytoophidium-deficient H355A CTPS mutant similarly inhibits proliferation.","method":"Co-immunoprecipitation (Co-IP) of HSPD1-CTPS; Hspd1 knockdown; CTPS overexpression; H355A mutation; EdU incorporation and viability assays","journal":"Experimental cell research","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP for interaction, loss-of-function with phenotype but limited mechanistic depth, single lab","pmids":["39971178"],"is_preprint":false}],"current_model":"CTPS1 (CTP synthetase 1) is the primary rate-limiting enzyme for de novo CTP biosynthesis, catalyzing conversion of UTP to CTP; it is less sensitive to CTP product feedback inhibition than its paralog CTPS2 (as shown by cryo-EM and biochemical assays), forms hyperactive higher-order filaments whose assembly is driven by CTP binding, directly heterodimerizes with CTPS2 which attenuates its activity, is transcriptionally activated by YBX1 and post-translationally stabilized by RASD2-mediated SUMOylation (inhibiting ubiquitin-proteasomal degradation), and is specifically required for T cell and lymphocyte proliferation upon activation—with CTPS1 inhibition inducing DNA replication stress (particularly in MYC-overexpressing cells), S-phase arrest, and synthetic lethality when combined with ATR or CHK1 inhibitors."},"narrative":{"mechanistic_narrative":"CTPS1 is the principal rate-limiting enzyme for de novo CTP biosynthesis and the dominant isoform driving the CTP-pool expansion required for proliferation, particularly in activated T lymphocytes [PMID:32161190, PMID:37348953]. It is intrinsically more active than its paralog CTPS2 and relatively resistant to CTP product feedback inhibition; cryo-EM shows that CTP binds two sites that clash with substrates to enforce feedback inhibition, while a single amino acid substitution governs the isoform selectivity exploited by small-molecule inhibitors [PMID:34583994, PMID:37348953]. CTP binding also drives assembly of CTPS1 into large-scale filaments that represent a hyperactive enzyme form, a mechanism conserved across eukaryotic CTPS [PMID:34583994, PMID:bio_10.1101_2025.02.22.639624]. CTPS1 directly heterodimerizes with CTPS2 independently of polymerization; within these heterocomplexes CTPS1 activity is dampened and rendered more feedback-sensitive, and filament (cytoophidium) formation itself is dispensable for proliferation [PMID:40957650]. CTPS1 abundance is set post-translationally through competing ubiquitination and stabilization, including RASD2-promoted SUMOylation that blocks ubiquitin-proteasomal degradation [PMID:39672102] and INHBA-mediated antagonism of SMURF1-driven ubiquitination [PMID:41239468], and transcriptionally through direct YBX1 promoter binding [PMID:34991621]. Because of its proliferative role, CTPS1 inhibition induces DNA replication stress—pronounced in MYC-overexpressing cells—S-phase arrest, and DNA damage, creating synthetic lethality with ATR, CHK1, and WEE1 inhibitors and synergy with BCL2 inhibition [PMID:35022212, PMID:38385294, PMID:37898670]. Loss-of-function CTPS1 mutation causing protein instability produces a human immunodeficiency with impaired T cell proliferation [PMID:32161190].","teleology":[{"year":2020,"claim":"Established that CTPS1 is genetically required for human T cell proliferation by showing a destabilizing mutation, not catalytic loss, abolishes function.","evidence":"Enzymatic activity assays in patient cells plus genetic complementation with WT and mutant constructs in CTPS1-deficient leukemia cells","pmids":["32161190"],"confidence":"High","gaps":["Does not resolve why CTPS1 is uniquely required over CTPS2 in lymphocytes","Mechanism of mutant protein instability not defined"]},{"year":2021,"claim":"Defined the structural basis of CTP feedback inhibition and isoform-selective inhibition, explaining CTPS1's reduced feedback sensitivity and its hyperactive filament form.","evidence":"Cryo-EM of inhibitor- and CTP-bound CTPS1 filaments with mutagenesis and primary T cell proliferation assays","pmids":["34583994"],"confidence":"High","gaps":["Functional advantage of filamentation in cells not fully resolved","In vivo regulation of filament assembly not addressed"]},{"year":2021,"claim":"Linked CTPS interaction partners and subcellular redistribution to metabolic state, suggesting context-dependent associations beyond catalysis.","evidence":"Co-IP/MS identifying ATP synthase and STRESS-70 partners, ATP measurements, and localization imaging in a decidualization model","pmids":["33576499"],"confidence":"Medium","gaps":["Interactions not validated reciprocally or isoform-resolved","Functional consequence of nuclear redistribution unclear"]},{"year":2022,"claim":"Identified a transcriptional driver of CTPS1, placing YBX1 upstream of CTPS1-dependent proliferation.","evidence":"ChIP and dual luciferase reporter assays with CTPS1 knockdown rescue in TNBC","pmids":["34991621"],"confidence":"Medium","gaps":["Generality of YBX1 control beyond TNBC unknown","Other transcriptional regulators not surveyed"]},{"year":2022,"claim":"Connected CTPS1 inhibition to selective DNA replication stress in MYC-driven cells and defined ATR synthetic lethality.","evidence":"Isoform-specific knockdown/inhibition, replication stress assays, and ATR-inhibitor combinations in vitro and in xenografts","pmids":["35022212"],"confidence":"High","gaps":["Why MYC-driven rRNA synthesis specifically sensitizes to CTPS1 loss not fully mechanistic","CTPS2 redundancy threshold not defined"]},{"year":2023,"claim":"Quantified CTPS1 as the dominant proliferative isoform with higher intrinsic activity and inhibitor resistance relative to CTPS2.","evidence":"KO/complementation, in vitro enzymatic and 3-deaza-uridine inhibition assays, and analysis of >1,000 cancer cell line datasets","pmids":["37348953"],"confidence":"High","gaps":["Structural basis of CTPS2 essentiality in CTPS1 absence not resolved here"]},{"year":2023,"claim":"Showed CTPS1 inhibition triggers DNA damage and cell cycle arrest exploitable through DDR-pathway combinations and apoptotic synergy.","evidence":"CTPS1 KO and STP-B inhibition with γH2AX, cell cycle, and ATR/CHEK1/WEE1 and BCL2-inhibitor combination studies in myeloma and mantle cell lymphoma in vitro and in vivo","pmids":["37898670","38385294"],"confidence":"Medium","gaps":["Direct cause of double-strand breaks downstream of CTP depletion not fully defined","Translation suppression mechanism beyond MCL1 not detailed"]},{"year":2024,"claim":"Identified post-translational stabilization of CTPS1 via RASD2-promoted SUMOylation antagonizing ubiquitination.","evidence":"Co-IP, IP-MS, ubiquitination/SUMOylation Western blots, and ChIP-qPCR in endometriosis models","pmids":["39672102"],"confidence":"Medium","gaps":["SUMO acceptor sites on CTPS1 not mapped","Single-lab finding in one disease context"]},{"year":2024,"claim":"Extended CTPS1's role beyond nucleotide synthesis to glycerophospholipid reprogramming and mitophagy via CTP-dependent CEPT1 upregulation.","evidence":"CTPS1 KO/KD, scRNA-seq, selective inhibitor R80, and mitophagy assays in DLBCL","pmids":["41865720"],"confidence":"Medium","gaps":["Mechanism by which CTP availability raises CEPT1 expression unclear","Single-lab, single tumor type"]},{"year":2025,"claim":"Demonstrated direct CTPS1-CTPS2 heterocomplex formation that attenuates CTPS1 activity and showed cytoophidia are dispensable for proliferation.","evidence":"Co-IP, enzymatic assays of purified complexes, KO/complementation, and H355A cytoophidium-deficient mutant analysis","pmids":["40957650"],"confidence":"High","gaps":["Stoichiometry and regulation of hetero- vs homo-complexes in cells not resolved","Physiological trigger for heterocomplex assembly unknown"]},{"year":2025,"claim":"Resolved that the enzymatic product CTP drives CTPS1 filamentation through two distinct binding pockets in an evolutionarily conserved mechanism.","evidence":"Cryo-EM of CTP-bound hCTPS1 filaments with biochemical filamentation and pocket-mutant assays (preprint)","pmids":["bio_10.1101_2025.02.22.639624"],"confidence":"Medium","gaps":["Preprint not yet peer-reviewed","Cellular consequence of product-driven assembly versus disassembly not established"]},{"year":2025,"claim":"Identified INHBA as a stabilizer of CTPS1 by competing with SMURF1-mediated ubiquitination, linking CTPS1 abundance to chemoresistance.","evidence":"IP-MS, Co-IP, in vitro ubiquitination assays, and xenografts in pancreatic cancer","pmids":["41239468"],"confidence":"Medium","gaps":["SMURF1 ubiquitination sites on CTPS1 not mapped","Single-lab finding"]},{"year":2025,"claim":"Probed chaperone control of CTPS assembly, implicating HSPD1 in cytoophidium formation and proliferation.","evidence":"Co-IP of HSPD1-CTPS, Hspd1 knockdown, CTPS overexpression, and H355A mutant with proliferation assays in C2C12 cells","pmids":["39971178"],"confidence":"Low","gaps":["Single Co-IP without reciprocal validation","Direct role of HSPD1 in assembly versus folding not separated","Conflicts with finding that cytoophidia are dispensable for proliferation"]},{"year":null,"claim":"How CTPS1 abundance, heterocomplex composition, and filament state are coordinately regulated in vivo to set proliferative CTP supply remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified in vivo model integrating transcriptional, ubiquitin/SUMO, and assembly-state control","Physiological cues governing CTPS1-CTPS2 heterocomplex formation unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[0,1,2]},{"term_id":"GO:0140098","term_label":"catalytic activity, acting on RNA","supporting_discovery_ids":[0,1,2]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[0,1,2]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[7]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[7]}],"pathway":[{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[0,1,2]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[11,12]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[1]}],"complexes":["CTPS1-CTPS2 heterocomplex","CTPS cytoophidium (filament)"],"partners":["CTPS2","RASD2","INHBA","SMURF1","YBX1","HSPD1","CEPT1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P17812","full_name":"CTP synthase 1","aliases":["CTP synthetase 1","Protein-asparagine deamidase CTPS1","UTP--ammonia ligase 1"],"length_aa":591,"mass_kda":66.7,"function":"CTP synthase involved in the de novo synthesis of CTP, a precursor of DNA, RNA and phospholipids (PubMed:16179339, PubMed:17189248, PubMed:17463002, PubMed:24870241, PubMed:28459447, PubMed:34583994). Catalyzes the ATP-dependent amination of UTP to CTP with either L-glutamine or ammonia as a source of nitrogen (PubMed:16179339, PubMed:24870241, PubMed:28459447, PubMed:34583994). CTPS1 CTP synthase activity plays a crucial role in the proliferation of activated lymphocytes and immunity; additional CTP being required to meet increased demand for DNA, RNA and lipid membrane biosynthesis in proliferating lymphocytes (PubMed:24870241, PubMed:8530356). In addition to CTP synthase activity, also acts as a protein deamidase that catalyzes the side chain deamidation of specific asparagine residues of proteins to aspartate (PubMed:40240600). Acts as a negative regulator of innate immunity by mediating deamidation of 'Asn-85' of IRF3, preventing IRF3 from binding DNA (By similarity). Facilitates chromatin relaxation in response to DNA damage by mediating deamidation of 'Asn-76' and 'Asn-77' of histone H1, thereby promoting subsequent acetylation of histone H1 at 'Lys-75' (H1K75ac), increasing chromatin accessibility to facilitate the recruitment of DNA repair proteins (PubMed:40240600)","subcellular_location":"Cytoplasm, cytosol; Nucleus; Chromosome","url":"https://www.uniprot.org/uniprotkb/P17812/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/CTPS1","classification":"Common Essential","n_dependent_lines":781,"n_total_lines":1208,"dependency_fraction":0.6465231788079471},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000171793","cell_line_id":"CID001768","localizations":[{"compartment":"cytoplasmic","grade":3},{"compartment":"vesicles","grade":2}],"interactors":[{"gene":"CTPS2","stoichiometry":10.0},{"gene":"ATP6V1E1","stoichiometry":0.2},{"gene":"GLUL","stoichiometry":0.2},{"gene":"GSPT1","stoichiometry":0.2},{"gene":"MRPL20","stoichiometry":0.2},{"gene":"C21ORF59","stoichiometry":0.2},{"gene":"SUGP1","stoichiometry":0.2},{"gene":"METAP2","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001768","total_profiled":1310},"omim":[{"mim_id":"618534","title":"IMMUNODEFICIENCY 64 WITH LYMPHOPROLIFERATION; IMD64","url":"https://www.omim.org/entry/618534"},{"mim_id":"615897","title":"IMMUNODEFICIENCY 24; IMD24","url":"https://www.omim.org/entry/615897"},{"mim_id":"603962","title":"RAS GUANYL NUCLEOTIDE-RELEASING PROTEIN 1; RASGRP1","url":"https://www.omim.org/entry/603962"},{"mim_id":"123860","title":"CYTIDINE 5-PRIME TRIPHOSPHATE SYNTHETASE 1; CTPS1","url":"https://www.omim.org/entry/123860"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Cytosol","reliability":"Approved"},{"location":"Actin filaments","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/CTPS1"},"hgnc":{"alias_symbol":["GATD5A"],"prev_symbol":["CTPS"]},"alphafold":{"accession":"P17812","domains":[{"cath_id":"3.40.50.300","chopping":"1-289","consensus_level":"high","plddt":94.4962,"start":1,"end":289},{"cath_id":"3.40.50.880","chopping":"295-556","consensus_level":"high","plddt":94.6708,"start":295,"end":556}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P17812","model_url":"https://alphafold.ebi.ac.uk/files/AF-P17812-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P17812-F1-predicted_aligned_error_v6.png","plddt_mean":91.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CTPS1","jax_strain_url":"https://www.jax.org/strain/search?query=CTPS1"},"sequence":{"accession":"P17812","fasta_url":"https://rest.uniprot.org/uniprotkb/P17812.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P17812/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P17812"}},"corpus_meta":[{"pmid":"1959918","id":"PMC_1959918","title":"Chromosome mapping of the human cytidine-5'-triphosphate synthetase (CTPS) gene to band 1p34.1-p34.3 by fluorescence in situ hybridization.","date":"1991","source":"Human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/1959918","citation_count":124,"is_preprint":false},{"pmid":"30085408","id":"PMC_30085408","title":"Interfilament interaction between IMPDH and CTPS cytoophidia.","date":"2018","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/30085408","citation_count":54,"is_preprint":false},{"pmid":"34991621","id":"PMC_34991621","title":"CTPS1 promotes malignant progression of triple-negative breast cancer with transcriptional activation by YBX1.","date":"2022","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/34991621","citation_count":47,"is_preprint":false},{"pmid":"35022212","id":"PMC_35022212","title":"Combined Inactivation of CTPS1 and ATR Is Synthetically Lethal to MYC-Overexpressing Cancer Cells.","date":"2022","source":"Cancer 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America","url":"https://pubmed.ncbi.nlm.nih.gov/34583994","citation_count":33,"is_preprint":false},{"pmid":"34022203","id":"PMC_34022203","title":"CTPS and IMPDH form cytoophidia in developmental thymocytes.","date":"2021","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/34022203","citation_count":22,"is_preprint":false},{"pmid":"37898670","id":"PMC_37898670","title":"CTPS1 is a novel therapeutic target in multiple myeloma which synergizes with inhibition of CHEK1, ATR or WEE1.","date":"2023","source":"Leukemia","url":"https://pubmed.ncbi.nlm.nih.gov/37898670","citation_count":20,"is_preprint":false},{"pmid":"34129847","id":"PMC_34129847","title":"CTPS forms the cytoophidium in zebrafish.","date":"2021","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/34129847","citation_count":20,"is_preprint":false},{"pmid":"37348953","id":"PMC_37348953","title":"Differential roles of CTP synthetases CTPS1 and CTPS2 in cell 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CTP regulates both CTPS1 and CTPS2 by binding in two sites that clash with substrates (product feedback inhibition). CTPS1 is less sensitive to CTP feedback inhibition than CTPS2, consistent with its role in expanding CTP pools during lymphocyte proliferation. Small-molecule inhibitors that are CTPS1-selective or non-selective mimic CTP binding at one inhibitory site, with a single amino acid substitution explaining isoform selectivity. Both inhibitors bind to CTPS1 assembled into large-scale filaments, which represent a hyperactive form of the enzyme.\",\n      \"method\": \"Cryo-electron microscopy (cryo-EM) structures of inhibitor-bound and CTP-bound CTPS1 filaments; biochemical inhibition assays; site-directed mutagenesis (amino acid substitution analysis); primary T cell proliferation assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structural determination combined with biochemical assays and mutagenesis in a single rigorous study, with functional validation in primary T cells\",\n      \"pmids\": [\"34583994\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"CTPS1 deficiency caused by homozygous frameshift splice mutation (c.1692-1G>C, p.T566Dfs26X) results in 80–90% reduction of CTPS protein and CTP synthetase activity in patient lymphocytes, leading to severely impaired T cell proliferation and IL-2 secretion upon TCR activation. The mutant protein (T566Dfs26X) retains normal CTPS enzymatic activity when expressed at wild-type levels; loss of function is entirely attributable to protein instability. Inactivation of CTPS1 in a T cell leukemia line fully abolished proliferation, confirming CTPS1 is required for T cell proliferative responses.\",\n      \"method\": \"CTPS enzymatic activity assay in patient cells; genetic complementation in CTPS1-deficient leukemia cells with WT and mutant constructs; immune phenotyping; T cell proliferation and IL-2 secretion assays\",\n      \"journal\": \"JCI insight\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — enzymatic activity assay combined with genetic complementation and loss-of-function in human patient cells and leukemia cell line, multiple orthogonal methods\",\n      \"pmids\": [\"32161190\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CTPS1 has higher intrinsic CTP synthetase enzymatic activity than CTPS2 and is more resistant to inhibition by the UTP analog 3-deaza-uridine. Using inactivation and complementation experiments, CTPS1 was shown to be the primary contributor to cell proliferation; CTPS2 contributes modestly when CTPS1 is present but is essential in its absence.\",\n      \"method\": \"CTPS1 and/or CTPS2 inactivation by KO; complementation experiments; in vitro enzymatic activity assays; 3-deaza-uridine inhibition assays; analysis of >1,000 cancer cell line datasets\",\n      \"journal\": \"Life science alliance\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — enzymatic activity assays plus KO/complementation with multiple orthogonal methods and large-scale dataset validation\",\n      \"pmids\": [\"37348953\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Inhibition of CTPS1 (but not CTPS2) selectively induces DNA replication stress in MYC-overexpressing cancer cells. MYC-driven rRNA synthesis causes selective replication stress upon CTPS1 inhibition. Combined inhibition of CTPS1 and ATR is synthetically lethal in MYC-overexpressing cells in vitro and decreases tumor growth in vivo.\",\n      \"method\": \"CTPS1/CTPS2-selective knockdown/inhibition; replication stress assays; cell viability assays; in vivo xenograft tumor models; ATR inhibitor combination studies\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — isoform-specific KD with defined molecular phenotype (replication stress), epistatic combination with ATR inhibitor, both in vitro and in vivo validation\",\n      \"pmids\": [\"35022212\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CTPS1 and CTPS2 directly interact with each other independently of polymerization and cytoophidium formation, forming heterocomplexes. When associated with CTPS2, CTPS1 enzymatic activity is decreased and becomes more sensitive to CTP product feedback inhibition, demonstrating that CTPS2 modulates CTPS1 activity through heterocomplex formation. CTPS2-containing filaments (cytoophidia) are dependent on CTPS1 expression. CTPS1H355A and CTPS2H355A mutants unable to form cytoophidia can still sustain normal cell proliferation, showing cytoophidia are not required for proliferation.\",\n      \"method\": \"Co-immunoprecipitation; enzymatic activity assays with purified complexes; CTPS1/CTPS2 KO and complementation; H355A cytoophidium-deficient mutant analysis; co-localization imaging\",\n      \"journal\": \"Life science alliance\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — direct interaction confirmed by Co-IP, enzymatic activity measured for complexes, multiple mutant analyses and orthogonal methods in one study\",\n      \"pmids\": [\"40957650\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CTP (the enzymatic product of CTPS1) acts as a key regulator driving hCTPS1 filamentation. Cryo-EM structures of CTP-bound hCTPS1 filaments reveal the molecular details of CTP binding in filament assembly. CTP generated from the enzymatic reaction does not trigger filament disassembly. Two distinct CTP-binding pockets mediate this filamentation, and the mechanism is evolutionarily conserved across eukaryotic CTPS.\",\n      \"method\": \"Cryo-electron microscopy (cryo-EM) of CTP-bound hCTPS1 filaments; biochemical filamentation assays; analysis of CTP-binding pocket mutants\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — cryo-EM structure with biochemical validation, single lab, preprint not yet peer-reviewed\",\n      \"pmids\": [\"bio_10.1101_2025.02.22.639624\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"RASD2 promotes SUMOylation of CTPS1 and inhibits its ubiquitination, thereby increasing CTPS1 protein stability. This was demonstrated by co-immunoprecipitation and immunoprecipitation-mass spectrometry in endometriosis cells, where histone lactylation upregulates RASD2, which in turn stabilizes CTPS1 via the SUMOylation/ubiquitination balance.\",\n      \"method\": \"Co-immunoprecipitation (Co-IP); immunoprecipitation-mass spectrometry (IP-MS); Western blot for ubiquitination and SUMOylation; ChIP-qPCR; in vitro and in vivo endometriosis models\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP and IP-MS for interaction, SUMOylation/ubiquitination assays, single lab with two orthogonal biochemical methods\",\n      \"pmids\": [\"39672102\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CTPS interacts with ATP synthase (ATPS) and maintains ATP content during early decidualization (Day 3). At Day 6 of decidualization, CTPS instead associates with mitochondrial stress protein STRESS-70, correlating with reduced ATP concentration. CTPS localization shifts from cytoplasm (Day 3) to both cytoplasm and nucleus (Day 6), suggesting subcellular redistribution is linked to metabolic function during the decidualization process.\",\n      \"method\": \"Co-immunoprecipitation coupled with mass spectrometry; subcellular fractionation/immunofluorescence localization; CTPS knockdown by siRNA; in vitro decidualization model\",\n      \"journal\": \"Journal of cellular physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP/MS identified CTPS-ATPS and CTPS-STRESS-70 interactions, functional ATP measurements, subcellular localization with functional consequences, single lab\",\n      \"pmids\": [\"33576499\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"CTPS1 upregulates expression of choline/ethanolamine phosphotransferase 1 (CEPT1) by increasing CTP availability, thereby reprogramming glycerophospholipid metabolism. Glycerophospholipids synthesized by CEPT1 maintain mitochondrial homeostasis and promote BNIP3-mediated mitophagy, driving DLBCL progression.\",\n      \"method\": \"CTPS1 knockout/knockdown; single-cell RNA sequencing; Western blot and gene expression analysis; selective CTPS1 inhibitor (R80); functional mitophagy assays\",\n      \"journal\": \"Redox biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO with defined molecular pathway (CEPT1/phospholipid/BNIP3), pharmacological inhibitor corroboration, single lab\",\n      \"pmids\": [\"41865720\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"INHBA interacts with CTPS1 and competitively inhibits SMURF1 (SMAD Specific E3 Ubiquitin Protein Ligase 1)-mediated ubiquitination of CTPS1, thereby enhancing CTPS1 protein stability and promoting pyrimidine metabolism and gemcitabine resistance in pancreatic cancer cells.\",\n      \"method\": \"Immunoprecipitation mass spectrometry to identify CTPS1 as INHBA binding partner; Co-IP; in vitro ubiquitination assay; in vivo xenograft model\",\n      \"journal\": \"Cancer cell international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — IP-MS plus Co-IP for interaction, ubiquitination assay, single lab with two orthogonal methods\",\n      \"pmids\": [\"41239468\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"YBX1 (Y-box binding protein 1) directly binds to the CTPS1 promoter and activates its transcription, as demonstrated by dual luciferase reporter assays and chromatin immunoprecipitation (ChIP). Rescue experiments confirmed that enhanced cell proliferation and invasion driven by YBX1 overexpression could be reversed by CTPS1 knockdown, placing YBX1 upstream of CTPS1 in TNBC.\",\n      \"method\": \"Dual luciferase reporter assay; chromatin immunoprecipitation (ChIP); CTPS1 knockdown; YBX1 overexpression rescue experiment\",\n      \"journal\": \"Journal of translational medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP and luciferase reporter establish direct transcriptional regulation, epistasis via rescue experiment, single lab\",\n      \"pmids\": [\"34991621\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CTPS1 pharmacological inhibition by STP-B in mantle cell lymphoma causes rapid cell cycle arrest in early S-phase, inhibition of translation including anti-apoptotic MCL1 protein, and synergistic cell death in combination with the BCL2 inhibitor venetoclax, both in vitro and in vivo.\",\n      \"method\": \"Selective CTPS1 inhibitor STP-B; flow cytometry cell cycle analysis; Western blot for MCL1; in vitro and in vivo MCL models; BCL2 inhibitor combination assay\",\n      \"journal\": \"Haematologica\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological inhibition with defined molecular consequence (MCL1 suppression, S-phase arrest), in vivo validation, single lab\",\n      \"pmids\": [\"38385294\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CTPS1 knockout in multiple myeloma cells induces apoptosis and S-phase arrest with DNA double-strand breaks. Pharmacological inhibition of CTPS1 by STP-B activates DNA damage response (DDR) pathways. Combined inhibition of CTPS1 with ATR, CHEK1, or WEE1 inhibitors results in synergistic growth inhibition and early apoptosis.\",\n      \"method\": \"CTPS1 knockout; selective CTPS1 inhibitor STP-B; flow cytometry; DNA damage marker assays (γH2AX); DDR pathway inhibitor combinations; apoptosis assays\",\n      \"journal\": \"Leukemia\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO and pharmacological inhibition with defined molecular phenotype (DDR activation, DSBs), DDR pathway epistasis, single lab\",\n      \"pmids\": [\"37898670\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Single-molecule fluorescence imaging demonstrated that CTPS self-assembles with tetramers as the minimum structural unit driving cytoophidium formation. CTPS acts as the nucleation core to induce subsequent growth of P5CS (Δ1-pyrroline-5-carboxylate synthase) filaments, constituting a coassembly (coordinated assembly) process in vitro.\",\n      \"method\": \"Single-molecule fluorescence imaging; single-molecule photobleach counting for stoichiometry; oligomer state distribution analysis under different conditions; in vitro reconstitution of CTPS and P5CS coassembly\",\n      \"journal\": \"The journal of physical chemistry. B\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with single-molecule imaging and stoichiometry, single lab but multiple orthogonal approaches\",\n      \"pmids\": [\"38236746\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"HSPD1 (HSP60) interacts with CTPS protein; interference with Hspd1 gene expression inhibits CTPS cytoophidium formation even when CTPS is overexpressed, and this inhibits C2C12 skeletal muscle cell proliferation. The cytoophidium-deficient H355A CTPS mutant similarly inhibits proliferation.\",\n      \"method\": \"Co-immunoprecipitation (Co-IP) of HSPD1-CTPS; Hspd1 knockdown; CTPS overexpression; H355A mutation; EdU incorporation and viability assays\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP for interaction, loss-of-function with phenotype but limited mechanistic depth, single lab\",\n      \"pmids\": [\"39971178\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"CTPS1 (CTP synthetase 1) is the primary rate-limiting enzyme for de novo CTP biosynthesis, catalyzing conversion of UTP to CTP; it is less sensitive to CTP product feedback inhibition than its paralog CTPS2 (as shown by cryo-EM and biochemical assays), forms hyperactive higher-order filaments whose assembly is driven by CTP binding, directly heterodimerizes with CTPS2 which attenuates its activity, is transcriptionally activated by YBX1 and post-translationally stabilized by RASD2-mediated SUMOylation (inhibiting ubiquitin-proteasomal degradation), and is specifically required for T cell and lymphocyte proliferation upon activation—with CTPS1 inhibition inducing DNA replication stress (particularly in MYC-overexpressing cells), S-phase arrest, and synthetic lethality when combined with ATR or CHK1 inhibitors.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CTPS1 is the principal rate-limiting enzyme for de novo CTP biosynthesis and the dominant isoform driving the CTP-pool expansion required for proliferation, particularly in activated T lymphocytes [#1, #2]. It is intrinsically more active than its paralog CTPS2 and relatively resistant to CTP product feedback inhibition; cryo-EM shows that CTP binds two sites that clash with substrates to enforce feedback inhibition, while a single amino acid substitution governs the isoform selectivity exploited by small-molecule inhibitors [#0, #2]. CTP binding also drives assembly of CTPS1 into large-scale filaments that represent a hyperactive enzyme form, a mechanism conserved across eukaryotic CTPS [#0, #5]. CTPS1 directly heterodimerizes with CTPS2 independently of polymerization; within these heterocomplexes CTPS1 activity is dampened and rendered more feedback-sensitive, and filament (cytoophidium) formation itself is dispensable for proliferation [#4]. CTPS1 abundance is set post-translationally through competing ubiquitination and stabilization, including RASD2-promoted SUMOylation that blocks ubiquitin-proteasomal degradation [#6] and INHBA-mediated antagonism of SMURF1-driven ubiquitination [#9], and transcriptionally through direct YBX1 promoter binding [#10]. Because of its proliferative role, CTPS1 inhibition induces DNA replication stress—pronounced in MYC-overexpressing cells—S-phase arrest, and DNA damage, creating synthetic lethality with ATR, CHK1, and WEE1 inhibitors and synergy with BCL2 inhibition [#3, #11, #12]. Loss-of-function CTPS1 mutation causing protein instability produces a human immunodeficiency with impaired T cell proliferation [#1].\",\n  \"teleology\": [\n    {\n      \"year\": 2020,\n      \"claim\": \"Established that CTPS1 is genetically required for human T cell proliferation by showing a destabilizing mutation, not catalytic loss, abolishes function.\",\n      \"evidence\": \"Enzymatic activity assays in patient cells plus genetic complementation with WT and mutant constructs in CTPS1-deficient leukemia cells\",\n      \"pmids\": [\"32161190\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not resolve why CTPS1 is uniquely required over CTPS2 in lymphocytes\", \"Mechanism of mutant protein instability not defined\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Defined the structural basis of CTP feedback inhibition and isoform-selective inhibition, explaining CTPS1's reduced feedback sensitivity and its hyperactive filament form.\",\n      \"evidence\": \"Cryo-EM of inhibitor- and CTP-bound CTPS1 filaments with mutagenesis and primary T cell proliferation assays\",\n      \"pmids\": [\"34583994\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional advantage of filamentation in cells not fully resolved\", \"In vivo regulation of filament assembly not addressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Linked CTPS interaction partners and subcellular redistribution to metabolic state, suggesting context-dependent associations beyond catalysis.\",\n      \"evidence\": \"Co-IP/MS identifying ATP synthase and STRESS-70 partners, ATP measurements, and localization imaging in a decidualization model\",\n      \"pmids\": [\"33576499\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Interactions not validated reciprocally or isoform-resolved\", \"Functional consequence of nuclear redistribution unclear\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified a transcriptional driver of CTPS1, placing YBX1 upstream of CTPS1-dependent proliferation.\",\n      \"evidence\": \"ChIP and dual luciferase reporter assays with CTPS1 knockdown rescue in TNBC\",\n      \"pmids\": [\"34991621\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Generality of YBX1 control beyond TNBC unknown\", \"Other transcriptional regulators not surveyed\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Connected CTPS1 inhibition to selective DNA replication stress in MYC-driven cells and defined ATR synthetic lethality.\",\n      \"evidence\": \"Isoform-specific knockdown/inhibition, replication stress assays, and ATR-inhibitor combinations in vitro and in xenografts\",\n      \"pmids\": [\"35022212\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Why MYC-driven rRNA synthesis specifically sensitizes to CTPS1 loss not fully mechanistic\", \"CTPS2 redundancy threshold not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Quantified CTPS1 as the dominant proliferative isoform with higher intrinsic activity and inhibitor resistance relative to CTPS2.\",\n      \"evidence\": \"KO/complementation, in vitro enzymatic and 3-deaza-uridine inhibition assays, and analysis of >1,000 cancer cell line datasets\",\n      \"pmids\": [\"37348953\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of CTPS2 essentiality in CTPS1 absence not resolved here\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Showed CTPS1 inhibition triggers DNA damage and cell cycle arrest exploitable through DDR-pathway combinations and apoptotic synergy.\",\n      \"evidence\": \"CTPS1 KO and STP-B inhibition with γH2AX, cell cycle, and ATR/CHEK1/WEE1 and BCL2-inhibitor combination studies in myeloma and mantle cell lymphoma in vitro and in vivo\",\n      \"pmids\": [\"37898670\", \"38385294\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct cause of double-strand breaks downstream of CTP depletion not fully defined\", \"Translation suppression mechanism beyond MCL1 not detailed\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified post-translational stabilization of CTPS1 via RASD2-promoted SUMOylation antagonizing ubiquitination.\",\n      \"evidence\": \"Co-IP, IP-MS, ubiquitination/SUMOylation Western blots, and ChIP-qPCR in endometriosis models\",\n      \"pmids\": [\"39672102\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"SUMO acceptor sites on CTPS1 not mapped\", \"Single-lab finding in one disease context\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Extended CTPS1's role beyond nucleotide synthesis to glycerophospholipid reprogramming and mitophagy via CTP-dependent CEPT1 upregulation.\",\n      \"evidence\": \"CTPS1 KO/KD, scRNA-seq, selective inhibitor R80, and mitophagy assays in DLBCL\",\n      \"pmids\": [\"41865720\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which CTP availability raises CEPT1 expression unclear\", \"Single-lab, single tumor type\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Demonstrated direct CTPS1-CTPS2 heterocomplex formation that attenuates CTPS1 activity and showed cytoophidia are dispensable for proliferation.\",\n      \"evidence\": \"Co-IP, enzymatic assays of purified complexes, KO/complementation, and H355A cytoophidium-deficient mutant analysis\",\n      \"pmids\": [\"40957650\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry and regulation of hetero- vs homo-complexes in cells not resolved\", \"Physiological trigger for heterocomplex assembly unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Resolved that the enzymatic product CTP drives CTPS1 filamentation through two distinct binding pockets in an evolutionarily conserved mechanism.\",\n      \"evidence\": \"Cryo-EM of CTP-bound hCTPS1 filaments with biochemical filamentation and pocket-mutant assays (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.02.22.639624\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Preprint not yet peer-reviewed\", \"Cellular consequence of product-driven assembly versus disassembly not established\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified INHBA as a stabilizer of CTPS1 by competing with SMURF1-mediated ubiquitination, linking CTPS1 abundance to chemoresistance.\",\n      \"evidence\": \"IP-MS, Co-IP, in vitro ubiquitination assays, and xenografts in pancreatic cancer\",\n      \"pmids\": [\"41239468\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"SMURF1 ubiquitination sites on CTPS1 not mapped\", \"Single-lab finding\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Probed chaperone control of CTPS assembly, implicating HSPD1 in cytoophidium formation and proliferation.\",\n      \"evidence\": \"Co-IP of HSPD1-CTPS, Hspd1 knockdown, CTPS overexpression, and H355A mutant with proliferation assays in C2C12 cells\",\n      \"pmids\": [\"39971178\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single Co-IP without reciprocal validation\", \"Direct role of HSPD1 in assembly versus folding not separated\", \"Conflicts with finding that cytoophidia are dispensable for proliferation\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How CTPS1 abundance, heterocomplex composition, and filament state are coordinately regulated in vivo to set proliferative CTP supply remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified in vivo model integrating transcriptional, ubiquitin/SUMO, and assembly-state control\", \"Physiological cues governing CTPS1-CTPS2 heterocomplex formation unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"GO:0140098\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [0, 1, 2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [7]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [11, 12]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"complexes\": [\"CTPS1-CTPS2 heterocomplex\", \"CTPS cytoophidium (filament)\"],\n    \"partners\": [\"CTPS2\", \"RASD2\", \"INHBA\", \"SMURF1\", \"YBX1\", \"HSPD1\", \"CEPT1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}