{"gene":"THEM4","run_date":"2026-06-10T10:51:55","timeline":{"discoveries":[{"year":2001,"finding":"CTMP (THEM4) binds specifically to the carboxyl-terminal regulatory domain of PKBα/Akt at the plasma membrane, reducing PKBα activity by inhibiting phosphorylation on both Ser473 and Thr308. CTMP expression also reverts the transformed phenotype of v-Akt-transformed cells.","method":"Protein binding assays (pull-down/co-IP), kinase activity assays, phosphorylation analysis, cell morphology/growth/tumorigenesis assays","journal":"Science","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal binding assays, phosphorylation/kinase assays, and functional reversion of transformation; foundational paper replicated by multiple subsequent studies","pmids":["11598301"],"is_preprint":false},{"year":2009,"finding":"CTMP (THEM4) localizes to mitochondria with a dual sub-mitochondrial distribution (membrane-bound pool and free pool in the inter-membrane space), is released into the cytosol early upon apoptosis induction, and overexpression sensitizes cells to apoptosis by increasing mitochondrial membrane depolarization and caspase-3/PARP cleavage. A CTMP mutant retained in mitochondria loses pro-apoptotic function. CTMP knockdown reduces mitochondrial membrane potential loss and caspase-3/PARP activation. CTMP also delays PKB phosphorylation following cell death induction.","method":"Subcellular fractionation, live-cell imaging, mitochondrial membrane potential assays, caspase-3/PARP cleavage assays, RNAi knockdown, CTMP retention mutant","journal":"Cellular signalling","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (fractionation, mutant analysis, KD and OE with defined phenotypic readouts) in a single study from the Hemmings lab","pmids":["19168129"],"is_preprint":false},{"year":2009,"finding":"CTMP (THEM4) overexpression or depletion regulates mitochondrial dynamics: both full-length CTMP and a cleavage-resistant mutant promote clustering of spherical mitochondria (consistent with increased fission), while CTMP depletion leads to accumulation of swollen, interconnected mitochondria without affecting fusion. CTMP knockout mouse liver mitochondria phenocopy the depletion phenotype.","method":"Fluorescence microscopy (mitochondrial morphology), RNAi knockdown, CTMP mutant overexpression, CTMP knockout mouse liver analysis","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vitro KD and OE with morphological readouts validated in vivo by CTMP-KO mice, multiple orthogonal approaches","pmids":["19421406"],"is_preprint":false},{"year":2009,"finding":"In global cerebral ischemia, CTMP (THEM4) expression and activation is induced in vulnerable hippocampal neurons; CTMP binds Akt and extinguishes its activity despite nuclear translocation and phosphorylation of Akt. RNAi-mediated depletion of CTMP in a stroke model restored Akt activity (assessed by kinase assays and phosphorylation of GSK-3β and FOXO3A) and rescued hippocampal neurons from ischemia-induced death.","method":"In vivo rat ischemia model, co-immunoprecipitation, kinase assays, phosphorylation of downstream targets (GSK-3β, FOXO3A), lentiviral RNAi, histological neuronal survival assessment","journal":"Nature neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP, kinase assays, RNAi rescue in clinically relevant in vivo model with multiple downstream readouts","pmids":["19349976"],"is_preprint":false},{"year":2009,"finding":"A cell-permeable peptide from the N-terminal domain of CTMP (TAT-CTMP4) induces dose-dependent apoptosis (caspase-3 activation) selectively in pancreatic adenocarcinoma cell lines and xenografts, and augments gemcitabine and radiation therapy in vivo, placing CTMP-mediated Akt inhibition upstream of caspase-3-dependent apoptosis.","method":"Cell-permeable peptide treatment, TUNEL assay, active caspase-3 measurement, xenograft and allograft tumor models","journal":"International journal of cancer","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro and in vivo functional assays with defined readouts, single lab","pmids":["19405118"],"is_preprint":false},{"year":2013,"finding":"CTMP (THEM4) overexpression suppresses insulin-induced PKB/Akt phosphorylation in HEK293 cells, and in obese mouse adipose tissue CTMP levels are elevated while phospho-PKB is reduced. Co-overexpression of LETM1 abrogates the CTMP-mediated suppression of PKB phosphorylation, suggesting LETM1 acts downstream of PKB activation to antagonize CTMP.","method":"Transient transfection, adenovirus-mediated overexpression, Western blot in HEK293 cells, Western blot/immunohistochemistry in HFD and ob/ob mouse models","journal":"Metabolism: clinical and experimental","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — overexpression assays in cells and in vivo observations, single lab, two settings","pmids":["24333006"],"is_preprint":false},{"year":2013,"finding":"In a hepatocellular carcinoma mouse model, CTMP (THEM4) acts as a direct binding partner of Akt1, and LETM1 acts as a binding partner of CTMP. Co-delivery of LETM1 and CTMP downregulated Akt1 pathway phosphorylation at both Ser473 and Thr308, induced mitochondrial morphological changes (swelling, loss of cristae), and triggered mitochondria-mediated apoptosis, reducing tumor incidence.","method":"Co-immunoprecipitation (binding partners), Western blot (Akt phosphorylation), in vivo HCC model (H-ras12V mice), gross/microscopic tumor evaluation","journal":"Cancer gene therapy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP for binding, in vivo functional model with defined phosphorylation readouts, single lab","pmids":["23392203"],"is_preprint":false},{"year":2014,"finding":"ATF3 transcriptionally represses CTMP expression in hypoxic neurons by binding to the ATF/CREB site in the CTMP promoter and blocking NF-κB binding, which otherwise activates CTMP transcription. This ATF3→CTMP cascade regulates neuronal apoptosis via Akt pathway modulation.","method":"Reporter assays, ChIP assays, gain- and loss-of-function experiments, ATF3-VP16 fusion (converted to activator), degradation-resistant IκBα, siRNA knockdown, p-Akt (Ser473) measurement","journal":"Molecular neurobiology","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — ChIP and reporter assays directly establish transcriptional mechanism; multiple orthogonal methods (gain/loss of function, ChIP, reporter, rescue) in single study","pmids":["24771044"],"is_preprint":false},{"year":2014,"finding":"In a mouse traumatic brain injury (TBI) model, CTMP expression increases significantly after injury and correlates with inhibition of Akt phosphorylation. siRNA-mediated knockdown of CTMP augments Akt phosphorylation and significantly improves neurological recovery over 28 days post-TBI.","method":"Controlled cortical impact TBI model, immunohistochemistry, Western blot (phospho-Akt/CTMP), intracranial siRNA injection, neurological functional scoring","journal":"Neurological research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo KD with defined functional and biochemical readouts, single lab","pmids":["24670215"],"is_preprint":false},{"year":2016,"finding":"In excitotoxic kainic acid-induced neurodegeneration, CTMP is markedly upregulated in hippocampal astrocytes. LPS/IFN-γ treatment of primary astrocytes induces early phosphorylation of CTMP (p-CTMP) followed by Akt and CREB phosphorylation, with later CTMP upregulation inhibiting Akt activity, suggesting a biphasic regulatory role of CTMP phosphorylation in Akt/CREB signaling in astrocytes.","method":"In vivo kainic acid mouse model, immunohistochemistry, Western blot (p-CTMP, p-Akt, p-CREB, CTMP), primary astrocyte culture with LPS/IFN-γ treatment","journal":"Experimental neurobiology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, correlative biochemistry with no direct mechanistic manipulation of CTMP phosphorylation","pmids":["28243164"],"is_preprint":false},{"year":2016,"finding":"In diabetic db/db mice with focal cerebral ischemia, CTMP expression is constitutively elevated, leading to decreased Akt kinase activity and worse neurological outcomes compared to non-diabetic controls. RNAi-mediated depletion of CTMP in db/db mice restores Akt activity, improves neurological scores, and reduces infarct volume.","method":"db/db mouse model, Western blot (CTMP, Akt activity), siRNA injection, PI3K inhibitor (LY294002) treatment, neurological scoring, infarct volume measurement","journal":"Neurochemical research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo RNAi rescue with Akt kinase activity and functional outcome measures, single lab","pmids":["27161366"],"is_preprint":false},{"year":2021,"finding":"miR-183-5p directly targets THEM4, inhibiting its mRNA and protein expression. Overexpression of THEM4 abrogates miR-183-5p-mediated oncogenic effects and inactivates Akt and NF-κB pathways in colon cancer cells, placing THEM4 as an upstream negative regulator of both Akt and NF-κB in this context.","method":"Luciferase/target validation of miR-183-5p→THEM4, Western blot (Akt, NF-κB), THEM4 overexpression rescue, CCK-8, colony formation, Transwell, xenograft assays","journal":"Frontiers in oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct miRNA target validation with functional rescue experiments, single lab","pmids":["34249713"],"is_preprint":false},{"year":2022,"finding":"LETM1 silencing downregulates CTMP in endometrial cancer cells (KLE), and CTMP overexpression rescues the suppressed malignant phenotype (viability, migration, invasion) caused by LETM1 silencing, placing CTMP downstream of LETM1 in endometrial cancer progression.","method":"siRNA knockdown of LETM1 and CTMP, CTMP overexpression rescue, Western blot, CCK-8, colony formation, wound healing, Transwell, xenograft model","journal":"Anti-cancer drugs","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis (rescue experiment), multiple phenotypic readouts, in vitro and in vivo, single lab","pmids":["35324530"],"is_preprint":false},{"year":2023,"finding":"CTMP (THEM4) deficiency in CTMP-KO mice mitigates denervation-induced skeletal muscle atrophy after sciatic nerve injury: CTMP-KO gastrocnemius shows higher Akt, GSK3β, S6, and 4E-BP1 phosphorylation and lower MuRF-1 and myostatin levels compared to wild-type, establishing CTMP as a regulator of Akt-dependent protein synthesis and ubiquitin-ligase-mediated catabolism in muscle.","method":"CTMP knockout mice, sciatic nerve injury model, Western blot (Akt, GSK3β, S6, 4E-BP1, MuRF-1, myostatin phosphorylation/expression), muscle weight measurement","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO with multiple defined biochemical and functional readouts in vivo, single lab","pmids":["36652767"],"is_preprint":false},{"year":2024,"finding":"PRAME, acting as a substrate-recognizing subunit of a Cul2-dependent E3 ubiquitin ligase, interacts with CTMP and p21, mediating their ubiquitination and proteasomal degradation. This leads to accumulation of phospho-Akt and CCND3, promoting proliferation in multiple myeloma cells.","method":"Co-immunoprecipitation (PRAME–CTMP interaction), ubiquitination assays, PRAME knockdown/overexpression, Western blot (p-Akt, CTMP, p21, CCND3), proliferation assays","journal":"Heliyon","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP and ubiquitination assays with functional KD/OE readouts, single lab","pmids":["39071619"],"is_preprint":false},{"year":2024,"finding":"CARD9 interacts with THEM4 in cholangiocarcinoma cells, and this interaction facilitates AKT and mTOR phosphorylation, promoting CCA cell proliferation and invasion.","method":"Co-immunoprecipitation or interaction assay (CARD9–THEM4), Western blot (AKT, mTOR phosphorylation), gain/loss-of-function in CCA cell lines and nude mouse models","journal":"International immunopharmacology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, interaction and downstream signaling described in abstract without detailed mechanistic dissection of THEM4's direct contribution","pmids":["39418733"],"is_preprint":false},{"year":2025,"finding":"CAMK2A facilitates THEM4 release from mitochondria in pancreatic ductal adenocarcinoma cells. Released THEM4 inhibits AKT phosphorylation and suppresses PDAC tumor growth; THEM4 knockdown accelerates in vivo tumor growth.","method":"Functional experiments (mitochondrial membrane potential, ATP, ROS assays), THEM4 KD and xenograft tumor growth, AKT phosphorylation Western blot, immunohistochemical microarray","journal":"Journal of translational medicine","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, abstract-level description of mechanism without detailed mechanistic methods for CAMK2A→THEM4 release step","pmids":["41353164"],"is_preprint":false},{"year":2025,"finding":"CTMP knockdown enhances sensitivity of endometrial cancer cells to medroxyprogesterone acetate (MPA) by suppressing the PI3K/AKT signaling pathway, as assessed by cell proliferation assays and Western blot of PI3K/AKT pathway proteins.","method":"siRNA knockdown of CTMP, Western blot (PI3K/AKT pathway proteins), CCK-8, EDU incorporation assay, immunohistochemistry in clinical samples","journal":"Reproductive sciences","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, KD with signaling readouts but no direct mechanistic interrogation of CTMP–AKT interaction","pmids":["41201690"],"is_preprint":false},{"year":2026,"finding":"In MSI-L/MSS colorectal cancer, CTMP interacts with REV7 (a novel high-affinity interaction identified by immunoprecipitation-mass spectrometry and validated by co-IP, GST pulldown with truncation/point mutants, and AlphaFold Multimer structural modeling). This CTMP–REV7 interaction facilitates CDK1-mediated G2/M progression and suppresses HLA-B/MHC-I expression. CTMP also sustains AKT/PD-L1 signaling and promotes fatty acid metabolism. CTMP knockdown diminishes CDK1 activity, induces G2/M arrest, restores HLA-B expression via cGAS signaling, enhances CD8+ T cell infiltration, and synergizes with REV7 overexpression and IFN-γ.","method":"Immunoprecipitation-mass spectrometry, co-IP, GST pulldown with truncation/point mutants, AlphaFold Multimer, CTMP KD/OE (in vitro and syngeneic mouse model), cell cycle analysis, flow cytometry (CD8+ T cells), HLA-B/MHC-I expression assays, AKT/PD-L1 Western blot","journal":"Cellular oncology","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — structural validation (AlphaFold + mutagenesis pulldown), MS-identified interaction confirmed by co-IP and GST pulldown, functional in vivo syngeneic model, single lab","pmids":["42143164"],"is_preprint":false}],"current_model":"THEM4/CTMP is a mitochondrial protein that functions as an endogenous negative regulator of PKB/Akt by binding to Akt's carboxyl-terminal regulatory domain and inhibiting its phosphorylation at both Ser473 and Thr308; it localizes to the mitochondrial inter-membrane space and membrane, is released to the cytosol during apoptosis to sensitize cells to caspase-dependent death, regulates mitochondrial fission dynamics, and is transcriptionally controlled by the ATF3/NF-κB axis; its activity is modulated by PRAME-mediated ubiquitination and proteasomal degradation; and through a newly identified interaction with REV7 it also regulates CDK1-driven cell cycle progression and MHC-I antigen presentation, collectively placing THEM4 at a signaling nexus connecting Akt activity, mitochondrial homeostasis, and immune evasion."},"narrative":{"mechanistic_narrative":"THEM4 (CTMP) is a mitochondrial protein that acts as an endogenous negative regulator of PKB/Akt signaling by binding the carboxyl-terminal regulatory domain of Akt and suppressing its phosphorylation at both Ser473 and Thr308, thereby reverting Akt-driven transformation [PMID:11598301]. It resides in the mitochondrial inter-membrane space as both membrane-bound and free pools, and upon apoptotic stimulation is released to the cytosol where it amplifies mitochondrial depolarization and caspase-3/PARP cleavage; a mitochondrially retained mutant loses this pro-apoptotic activity, coupling its subcellular release to cell-death sensitization [PMID:19168129]. Independent of apoptosis, THEM4 governs mitochondrial morphology, promoting fission-like clustering when present and yielding swollen, interconnected mitochondria when depleted, a phenotype confirmed in knockout mouse liver [PMID:19421406]. Through its Akt-inhibitory function THEM4 shapes injury and disease responses: it is induced in vulnerable neurons during cerebral ischemia and traumatic brain injury, where its depletion restores Akt activity and rescues neurons or improves recovery [PMID:19349976, PMID:24670215, PMID:27161366], and it regulates Akt-dependent protein synthesis versus ubiquitin-ligase catabolism in skeletal muscle, with its loss mitigating denervation-induced atrophy [PMID:36652767]. THEM4 expression and protein levels are controlled by an ATF3/NF-κB transcriptional axis [PMID:24771044], by miR-183-5p [PMID:34249713], and by PRAME-directed Cul2 E3 ligase ubiquitination and proteasomal degradation [PMID:39071619], while its interaction with LETM1 modulates its Akt-suppressing output [PMID:24333006, PMID:23392203]. A high-affinity interaction with REV7 extends THEM4 function beyond Akt, driving CDK1-dependent G2/M progression and suppressing HLA-B/MHC-I antigen presentation, positioning it at a nexus linking Akt activity, mitochondrial homeostasis, cell cycle, and immune evasion [PMID:42143164].","teleology":[{"year":2001,"claim":"Established THEM4/CTMP as a direct, physiological negative regulator of Akt, defining its founding molecular function.","evidence":"Binding assays, kinase and phosphorylation assays, and reversion of v-Akt transformation","pmids":["11598301"],"confidence":"High","gaps":["Did not resolve where in the cell the inhibitory interaction predominates","Structural basis of binding to the Akt C-terminal domain not defined"]},{"year":2009,"claim":"Resolved THEM4's mitochondrial localization and showed its apoptotic release from the inter-membrane space is required for pro-apoptotic activity, linking compartmentalization to function.","evidence":"Subcellular fractionation, live-cell imaging, mitochondrial membrane potential and caspase assays, retention mutant, RNAi","pmids":["19168129"],"confidence":"High","gaps":["Mechanism of release from mitochondria not defined","Relationship between mitochondrial pool and plasma-membrane Akt inhibition unclear"]},{"year":2009,"claim":"Demonstrated a distinct role for THEM4 in controlling mitochondrial fission/morphology, separating this from its apoptotic function.","evidence":"Fluorescence microscopy of mitochondrial morphology, RNAi, cleavage-resistant mutant overexpression, CTMP-KO mouse liver","pmids":["19421406"],"confidence":"High","gaps":["Molecular fission machinery engaged by THEM4 not identified","Whether morphology control requires Akt inhibition is unresolved"]},{"year":2009,"claim":"Showed THEM4 inhibition of Akt is functionally consequential in vivo, where its induction kills ischemic neurons and its depletion is protective.","evidence":"Rat global ischemia model, co-IP, kinase assays, downstream GSK-3β/FOXO3A readouts, lentiviral RNAi rescue; cell-permeable TAT-CTMP peptide in pancreatic cancer xenografts","pmids":["19349976","19405118"],"confidence":"High","gaps":["Upstream signal triggering THEM4 induction in ischemia not fully defined","Peptide tumor-selectivity mechanism not dissected"]},{"year":2014,"claim":"Identified the transcriptional control of THEM4 through competing ATF3 repression and NF-κB activation at its promoter, explaining its injury-induced expression.","evidence":"Reporter and ChIP assays, ATF3-VP16 and degradation-resistant IκBα constructs, siRNA, p-Akt readouts; TBI knockdown model with functional scoring","pmids":["24771044","24670215"],"confidence":"High","gaps":["Signals selecting ATF3 versus NF-κB occupancy not defined","Generality of this axis beyond neural injury untested"]},{"year":2013,"claim":"Placed THEM4 in a metabolic/oncogenic Akt context and identified LETM1 as a binding partner that modulates THEM4's Akt-suppressing output.","evidence":"Overexpression in HEK293 and obese mouse adipose tissue; co-IP and co-delivery of LETM1/CTMP in an HCC mouse model with phosphorylation and apoptosis readouts","pmids":["24333006","23392203"],"confidence":"Medium","gaps":["Mechanism by which LETM1 antagonizes or directs THEM4 not defined","Direct versus indirect LETM1–THEM4 binding not fully characterized"]},{"year":2016,"claim":"Extended THEM4 regulation to diabetic and excitotoxic injury and raised the possibility of phosphorylation-dependent biphasic control of its activity.","evidence":"db/db focal ischemia model with siRNA rescue and infarct measurement; kainic acid model and primary astrocyte LPS/IFN-γ stimulation tracking p-CTMP/p-Akt/p-CREB","pmids":["27161366","28243164"],"confidence":"Medium","gaps":["The p-CTMP phosphosite and responsible kinase were not directly manipulated","Astrocyte data are correlative without mechanistic perturbation"]},{"year":2022,"claim":"Documented post-transcriptional and proteostatic control of THEM4 abundance and embedded it in cancer signaling networks.","evidence":"miR-183-5p target validation with rescue in colon cancer; LETM1-silencing epistasis in endometrial cancer; PRAME/Cul2 co-IP and ubiquitination assays in myeloma","pmids":["34249713","35324530","39071619"],"confidence":"Medium","gaps":["Context-dependence of these regulators across tissues unresolved","Whether THEM4 degradation kinetics directly set Akt output not quantified"]},{"year":2023,"claim":"Defined THEM4 as a physiological brake on Akt-dependent anabolism and catabolism in skeletal muscle using genetic knockout.","evidence":"CTMP-KO mice with sciatic nerve denervation, Western blot of Akt/GSK3β/S6/4E-BP1 and MuRF-1/myostatin, muscle weight","pmids":["36652767"],"confidence":"Medium","gaps":["Whether muscle phenotype is cell-autonomous to myofibers not established","Mitochondrial contribution in muscle not examined"]},{"year":2026,"claim":"Discovered a high-affinity THEM4–REV7 interaction that couples THEM4 to CDK1-driven cell cycle progression and MHC-I/antigen-presentation control, expanding its role into immune evasion.","evidence":"IP-mass spectrometry, co-IP, GST pulldown with truncation/point mutants, AlphaFold Multimer, CTMP KD/OE in vitro and syngeneic mouse model, cell cycle and CD8+ T cell flow cytometry, HLA-B/AKT/PD-L1 readouts","pmids":["42143164"],"confidence":"Medium","gaps":["Whether REV7 binding is mechanistically separable from Akt regulation unclear","cGAS-dependent HLA-B restoration pathway not fully dissected","Single lab; reciprocal validation in other tumor types pending"]},{"year":null,"claim":"How THEM4's mitochondrial residence, its release to the cytosol, and its direct Akt and REV7 interactions are mechanistically integrated into a single regulatory logic remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structure of the THEM4–Akt complex","Trigger and machinery for mitochondrial release not defined","Whether Akt-inhibitory and REV7/cell-cycle functions are independent or coupled is unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,3,18]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0]}],"localization":[{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[1,2]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[1]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,3]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[1,4]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[18]}],"complexes":[],"partners":["AKT1","LETM1","REV7","PRAME","CARD9","CAMK2A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q5T1C6","full_name":"Acyl-coenzyme A thioesterase THEM4","aliases":["Carboxyl-terminal modulator protein","Thioesterase superfamily member 4"],"length_aa":240,"mass_kda":27.1,"function":"Has acyl-CoA thioesterase activity towards medium and long-chain (C14 to C18) fatty acyl-CoA substrates, and probably plays a role in mitochondrial fatty acid metabolism. Plays a role in the apoptotic process, possibly via its regulation of AKT1 activity. According to PubMed:11598301, inhibits AKT1 phosphorylation and activity. According to PubMed:17615157, enhances AKT1 activity by favoring its phosphorylation and translocation to plasma membrane","subcellular_location":"Cell membrane; Cell projection, ruffle membrane; Cytoplasm; Mitochondrion; Mitochondrion inner membrane; Mitochondrion intermembrane space","url":"https://www.uniprot.org/uniprotkb/Q5T1C6/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/THEM4","classification":"Not Classified","n_dependent_lines":20,"n_total_lines":1208,"dependency_fraction":0.016556291390728478},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/THEM4","total_profiled":1310},"omim":[{"mim_id":"615653","title":"THIOESTERASE SUPERFAMILY MEMBER 5; THEM5","url":"https://www.omim.org/entry/615653"},{"mim_id":"606388","title":"THIOESTERASE SUPERFAMILY MEMBER 4; THEM4","url":"https://www.omim.org/entry/606388"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/THEM4"},"hgnc":{"alias_symbol":["CTMP"],"prev_symbol":[]},"alphafold":{"accession":"Q5T1C6","domains":[{"cath_id":"3.10.129.10","chopping":"56-233","consensus_level":"high","plddt":94.2711,"start":56,"end":233}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q5T1C6","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q5T1C6-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q5T1C6-F1-predicted_aligned_error_v6.png","plddt_mean":83.88},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=THEM4","jax_strain_url":"https://www.jax.org/strain/search?query=THEM4"},"sequence":{"accession":"Q5T1C6","fasta_url":"https://rest.uniprot.org/uniprotkb/Q5T1C6.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q5T1C6/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q5T1C6"}},"corpus_meta":[{"pmid":"11598301","id":"PMC_11598301","title":"Carboxyl-terminal modulator protein (CTMP), a negative regulator of PKB/Akt and v-Akt at the plasma membrane.","date":"2001","source":"Science (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/11598301","citation_count":205,"is_preprint":false},{"pmid":"19349976","id":"PMC_19349976","title":"The endogenous inhibitor of Akt, CTMP, is critical to ischemia-induced neuronal death.","date":"2009","source":"Nature neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/19349976","citation_count":97,"is_preprint":false},{"pmid":"17013611","id":"PMC_17013611","title":"Activation of Akt independent of PTEN and CTMP tumor-suppressor gene mutations in epilepsy-associated Taylor-type focal cortical dysplasias.","date":"2006","source":"Acta neuropathologica","url":"https://pubmed.ncbi.nlm.nih.gov/17013611","citation_count":48,"is_preprint":false},{"pmid":"34249713","id":"PMC_34249713","title":"Exosomal miR-183-5p Shuttled by M2 Polarized Tumor-Associated Macrophage Promotes the Development of Colon Cancer via Targeting THEM4 Mediated PI3K/AKT and NF-κB Pathways.","date":"2021","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/34249713","citation_count":47,"is_preprint":false},{"pmid":"19405118","id":"PMC_19405118","title":"Targeting AKT with the proapoptotic peptide, TAT-CTMP: a novel strategy for the treatment of human pancreatic adenocarcinoma.","date":"2009","source":"International journal of cancer","url":"https://pubmed.ncbi.nlm.nih.gov/19405118","citation_count":44,"is_preprint":false},{"pmid":"19168129","id":"PMC_19168129","title":"Carboxy-Terminal Modulator Protein (CTMP) is a mitochondrial protein that sensitizes cells to apoptosis.","date":"2009","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/19168129","citation_count":34,"is_preprint":false},{"pmid":"24771044","id":"PMC_24771044","title":"Novel link of anti-apoptotic ATF3 with pro-apoptotic CTMP in the ischemic brain.","date":"2014","source":"Molecular neurobiology","url":"https://pubmed.ncbi.nlm.nih.gov/24771044","citation_count":27,"is_preprint":false},{"pmid":"24333006","id":"PMC_24333006","title":"New players in high fat diet-induced obesity: LETM1 and CTMP.","date":"2013","source":"Metabolism: clinical and experimental","url":"https://pubmed.ncbi.nlm.nih.gov/24333006","citation_count":20,"is_preprint":false},{"pmid":"27447863","id":"PMC_27447863","title":"CTMP, a predictive biomarker for trastuzumab resistance in HER2-enriched breast cancer patient.","date":"2017","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/27447863","citation_count":19,"is_preprint":false},{"pmid":"33533695","id":"PMC_33533695","title":"Abiotic Synthesis of Nucleoside 5'-Triphosphates with Nickel Borate and Cyclic Trimetaphosphate (CTMP).","date":"2021","source":"Astrobiology","url":"https://pubmed.ncbi.nlm.nih.gov/33533695","citation_count":18,"is_preprint":false},{"pmid":"19421406","id":"PMC_19421406","title":"The Carboxy-Terminal Modulator Protein (CTMP) regulates mitochondrial dynamics.","date":"2009","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/19421406","citation_count":17,"is_preprint":false},{"pmid":"23392203","id":"PMC_23392203","title":"Co-delivery of LETM1 and CTMP synergistically inhibits tumor growth in H-ras12V liver cancer model mice.","date":"2013","source":"Cancer gene therapy","url":"https://pubmed.ncbi.nlm.nih.gov/23392203","citation_count":16,"is_preprint":false},{"pmid":"35622138","id":"PMC_35622138","title":"Effect of different iodine levels on the DNA methylation of PRKAA2, ITGA6, THEM4 and PRL genes in PI3K-AKT signaling pathway and population-based validation from autoimmune thyroiditis patients.","date":"2022","source":"European journal of nutrition","url":"https://pubmed.ncbi.nlm.nih.gov/35622138","citation_count":15,"is_preprint":false},{"pmid":"24670215","id":"PMC_24670215","title":"Small interfering RNA directed against CTMP reduces acute traumatic brain injury in a mouse model by activating Akt.","date":"2014","source":"Neurological research","url":"https://pubmed.ncbi.nlm.nih.gov/24670215","citation_count":11,"is_preprint":false},{"pmid":"27161366","id":"PMC_27161366","title":"Elevated Expression of Carboxy-Terminal Modulator Protein (CTMP) Aggravates Brain Ischemic Injury in Diabetic db/db Mice.","date":"2016","source":"Neurochemical research","url":"https://pubmed.ncbi.nlm.nih.gov/27161366","citation_count":6,"is_preprint":false},{"pmid":"28243164","id":"PMC_28243164","title":"Astrocytic Expression of CTMP Following an Excitotoxic Lesion in the Mouse Hippocampus.","date":"2016","source":"Experimental neurobiology","url":"https://pubmed.ncbi.nlm.nih.gov/28243164","citation_count":6,"is_preprint":false},{"pmid":"36652767","id":"PMC_36652767","title":"Carboxyl-terminal modulator protein (CTMP) deficiency mitigates denervation-induced skeletal muscle atrophy.","date":"2023","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/36652767","citation_count":5,"is_preprint":false},{"pmid":"35324530","id":"PMC_35324530","title":"Mitochondrial protein LETM1 and its-mediated CTMP are potential therapeutic targets for endometrial cancer.","date":"2022","source":"Anti-cancer drugs","url":"https://pubmed.ncbi.nlm.nih.gov/35324530","citation_count":5,"is_preprint":false},{"pmid":"29892233","id":"PMC_29892233","title":"In Vitro Neurochemical Assessment of Methylphenidate and Its \"Legal High\" Analogs 3,4-CTMP and Ethylphenidate in Rat Nucleus Accumbens and Bed Nucleus of the Stria Terminalis.","date":"2018","source":"Frontiers in psychiatry","url":"https://pubmed.ncbi.nlm.nih.gov/29892233","citation_count":5,"is_preprint":false},{"pmid":"39011675","id":"PMC_39011675","title":"Roles of THEM4 in the Akt pathway: a double-edged sword.","date":"2024","source":"Journal of Zhejiang University. Science. B","url":"https://pubmed.ncbi.nlm.nih.gov/39011675","citation_count":3,"is_preprint":false},{"pmid":"39418733","id":"PMC_39418733","title":"CARD9 promotes cholangiocarcinoma by regulating the IL-17A/Hedgehog and the THEM4/AKT/mTOR signaling pathways.","date":"2024","source":"International immunopharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/39418733","citation_count":2,"is_preprint":false},{"pmid":"39071619","id":"PMC_39071619","title":"PRAME promotes proliferation of multiple myeloma cells through CTMP/Akt/p21/CCND3 axis by ubiquitinating CTMP and p21.","date":"2024","source":"Heliyon","url":"https://pubmed.ncbi.nlm.nih.gov/39071619","citation_count":2,"is_preprint":false},{"pmid":"41353164","id":"PMC_41353164","title":"A mitochondria-related gene-based signature predicts pancreatic ductal adenocarcinoma clinical outcome and revealed CAMK2A/THEM4 regulates progression phenotypes and mitophagy in vivo and in vitro.","date":"2025","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/41353164","citation_count":0,"is_preprint":false},{"pmid":"41201690","id":"PMC_41201690","title":"Knockdown of CTMP Enhances Progesterone Sensitivity in Endometrial Cancer by Inhibiting the PI3K/AKT Signaling Pathway.","date":"2025","source":"Reproductive sciences (Thousand Oaks, Calif.)","url":"https://pubmed.ncbi.nlm.nih.gov/41201690","citation_count":0,"is_preprint":false},{"pmid":"42143164","id":"PMC_42143164","title":"CTMP-REV7 axis modulates MHC-I antigen presentation via CDK1-AKT crosstalk in MSI-L/MSS colorectal cancer.","date":"2026","source":"Cellular oncology (Dordrecht, Netherlands)","url":"https://pubmed.ncbi.nlm.nih.gov/42143164","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":15987,"output_tokens":5229,"usd":0.063198,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13601,"output_tokens":3793,"usd":0.081415,"stage2_stop_reason":"end_turn"},"total_usd":0.144613,"stage1_batch_id":"msgbatch_017qqVz33Ga63SCDwzb35XPb","stage2_batch_id":"msgbatch_01Scr2Ei9x4HJMiRGxwrmvLK","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2001,\n      \"finding\": \"CTMP (THEM4) binds specifically to the carboxyl-terminal regulatory domain of PKBα/Akt at the plasma membrane, reducing PKBα activity by inhibiting phosphorylation on both Ser473 and Thr308. CTMP expression also reverts the transformed phenotype of v-Akt-transformed cells.\",\n      \"method\": \"Protein binding assays (pull-down/co-IP), kinase activity assays, phosphorylation analysis, cell morphology/growth/tumorigenesis assays\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal binding assays, phosphorylation/kinase assays, and functional reversion of transformation; foundational paper replicated by multiple subsequent studies\",\n      \"pmids\": [\"11598301\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"CTMP (THEM4) localizes to mitochondria with a dual sub-mitochondrial distribution (membrane-bound pool and free pool in the inter-membrane space), is released into the cytosol early upon apoptosis induction, and overexpression sensitizes cells to apoptosis by increasing mitochondrial membrane depolarization and caspase-3/PARP cleavage. A CTMP mutant retained in mitochondria loses pro-apoptotic function. CTMP knockdown reduces mitochondrial membrane potential loss and caspase-3/PARP activation. CTMP also delays PKB phosphorylation following cell death induction.\",\n      \"method\": \"Subcellular fractionation, live-cell imaging, mitochondrial membrane potential assays, caspase-3/PARP cleavage assays, RNAi knockdown, CTMP retention mutant\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (fractionation, mutant analysis, KD and OE with defined phenotypic readouts) in a single study from the Hemmings lab\",\n      \"pmids\": [\"19168129\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"CTMP (THEM4) overexpression or depletion regulates mitochondrial dynamics: both full-length CTMP and a cleavage-resistant mutant promote clustering of spherical mitochondria (consistent with increased fission), while CTMP depletion leads to accumulation of swollen, interconnected mitochondria without affecting fusion. CTMP knockout mouse liver mitochondria phenocopy the depletion phenotype.\",\n      \"method\": \"Fluorescence microscopy (mitochondrial morphology), RNAi knockdown, CTMP mutant overexpression, CTMP knockout mouse liver analysis\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vitro KD and OE with morphological readouts validated in vivo by CTMP-KO mice, multiple orthogonal approaches\",\n      \"pmids\": [\"19421406\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"In global cerebral ischemia, CTMP (THEM4) expression and activation is induced in vulnerable hippocampal neurons; CTMP binds Akt and extinguishes its activity despite nuclear translocation and phosphorylation of Akt. RNAi-mediated depletion of CTMP in a stroke model restored Akt activity (assessed by kinase assays and phosphorylation of GSK-3β and FOXO3A) and rescued hippocampal neurons from ischemia-induced death.\",\n      \"method\": \"In vivo rat ischemia model, co-immunoprecipitation, kinase assays, phosphorylation of downstream targets (GSK-3β, FOXO3A), lentiviral RNAi, histological neuronal survival assessment\",\n      \"journal\": \"Nature neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP, kinase assays, RNAi rescue in clinically relevant in vivo model with multiple downstream readouts\",\n      \"pmids\": [\"19349976\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"A cell-permeable peptide from the N-terminal domain of CTMP (TAT-CTMP4) induces dose-dependent apoptosis (caspase-3 activation) selectively in pancreatic adenocarcinoma cell lines and xenografts, and augments gemcitabine and radiation therapy in vivo, placing CTMP-mediated Akt inhibition upstream of caspase-3-dependent apoptosis.\",\n      \"method\": \"Cell-permeable peptide treatment, TUNEL assay, active caspase-3 measurement, xenograft and allograft tumor models\",\n      \"journal\": \"International journal of cancer\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro and in vivo functional assays with defined readouts, single lab\",\n      \"pmids\": [\"19405118\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"CTMP (THEM4) overexpression suppresses insulin-induced PKB/Akt phosphorylation in HEK293 cells, and in obese mouse adipose tissue CTMP levels are elevated while phospho-PKB is reduced. Co-overexpression of LETM1 abrogates the CTMP-mediated suppression of PKB phosphorylation, suggesting LETM1 acts downstream of PKB activation to antagonize CTMP.\",\n      \"method\": \"Transient transfection, adenovirus-mediated overexpression, Western blot in HEK293 cells, Western blot/immunohistochemistry in HFD and ob/ob mouse models\",\n      \"journal\": \"Metabolism: clinical and experimental\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — overexpression assays in cells and in vivo observations, single lab, two settings\",\n      \"pmids\": [\"24333006\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"In a hepatocellular carcinoma mouse model, CTMP (THEM4) acts as a direct binding partner of Akt1, and LETM1 acts as a binding partner of CTMP. Co-delivery of LETM1 and CTMP downregulated Akt1 pathway phosphorylation at both Ser473 and Thr308, induced mitochondrial morphological changes (swelling, loss of cristae), and triggered mitochondria-mediated apoptosis, reducing tumor incidence.\",\n      \"method\": \"Co-immunoprecipitation (binding partners), Western blot (Akt phosphorylation), in vivo HCC model (H-ras12V mice), gross/microscopic tumor evaluation\",\n      \"journal\": \"Cancer gene therapy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP for binding, in vivo functional model with defined phosphorylation readouts, single lab\",\n      \"pmids\": [\"23392203\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"ATF3 transcriptionally represses CTMP expression in hypoxic neurons by binding to the ATF/CREB site in the CTMP promoter and blocking NF-κB binding, which otherwise activates CTMP transcription. This ATF3→CTMP cascade regulates neuronal apoptosis via Akt pathway modulation.\",\n      \"method\": \"Reporter assays, ChIP assays, gain- and loss-of-function experiments, ATF3-VP16 fusion (converted to activator), degradation-resistant IκBα, siRNA knockdown, p-Akt (Ser473) measurement\",\n      \"journal\": \"Molecular neurobiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — ChIP and reporter assays directly establish transcriptional mechanism; multiple orthogonal methods (gain/loss of function, ChIP, reporter, rescue) in single study\",\n      \"pmids\": [\"24771044\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"In a mouse traumatic brain injury (TBI) model, CTMP expression increases significantly after injury and correlates with inhibition of Akt phosphorylation. siRNA-mediated knockdown of CTMP augments Akt phosphorylation and significantly improves neurological recovery over 28 days post-TBI.\",\n      \"method\": \"Controlled cortical impact TBI model, immunohistochemistry, Western blot (phospho-Akt/CTMP), intracranial siRNA injection, neurological functional scoring\",\n      \"journal\": \"Neurological research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo KD with defined functional and biochemical readouts, single lab\",\n      \"pmids\": [\"24670215\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"In excitotoxic kainic acid-induced neurodegeneration, CTMP is markedly upregulated in hippocampal astrocytes. LPS/IFN-γ treatment of primary astrocytes induces early phosphorylation of CTMP (p-CTMP) followed by Akt and CREB phosphorylation, with later CTMP upregulation inhibiting Akt activity, suggesting a biphasic regulatory role of CTMP phosphorylation in Akt/CREB signaling in astrocytes.\",\n      \"method\": \"In vivo kainic acid mouse model, immunohistochemistry, Western blot (p-CTMP, p-Akt, p-CREB, CTMP), primary astrocyte culture with LPS/IFN-γ treatment\",\n      \"journal\": \"Experimental neurobiology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, correlative biochemistry with no direct mechanistic manipulation of CTMP phosphorylation\",\n      \"pmids\": [\"28243164\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"In diabetic db/db mice with focal cerebral ischemia, CTMP expression is constitutively elevated, leading to decreased Akt kinase activity and worse neurological outcomes compared to non-diabetic controls. RNAi-mediated depletion of CTMP in db/db mice restores Akt activity, improves neurological scores, and reduces infarct volume.\",\n      \"method\": \"db/db mouse model, Western blot (CTMP, Akt activity), siRNA injection, PI3K inhibitor (LY294002) treatment, neurological scoring, infarct volume measurement\",\n      \"journal\": \"Neurochemical research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo RNAi rescue with Akt kinase activity and functional outcome measures, single lab\",\n      \"pmids\": [\"27161366\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"miR-183-5p directly targets THEM4, inhibiting its mRNA and protein expression. Overexpression of THEM4 abrogates miR-183-5p-mediated oncogenic effects and inactivates Akt and NF-κB pathways in colon cancer cells, placing THEM4 as an upstream negative regulator of both Akt and NF-κB in this context.\",\n      \"method\": \"Luciferase/target validation of miR-183-5p→THEM4, Western blot (Akt, NF-κB), THEM4 overexpression rescue, CCK-8, colony formation, Transwell, xenograft assays\",\n      \"journal\": \"Frontiers in oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct miRNA target validation with functional rescue experiments, single lab\",\n      \"pmids\": [\"34249713\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"LETM1 silencing downregulates CTMP in endometrial cancer cells (KLE), and CTMP overexpression rescues the suppressed malignant phenotype (viability, migration, invasion) caused by LETM1 silencing, placing CTMP downstream of LETM1 in endometrial cancer progression.\",\n      \"method\": \"siRNA knockdown of LETM1 and CTMP, CTMP overexpression rescue, Western blot, CCK-8, colony formation, wound healing, Transwell, xenograft model\",\n      \"journal\": \"Anti-cancer drugs\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis (rescue experiment), multiple phenotypic readouts, in vitro and in vivo, single lab\",\n      \"pmids\": [\"35324530\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CTMP (THEM4) deficiency in CTMP-KO mice mitigates denervation-induced skeletal muscle atrophy after sciatic nerve injury: CTMP-KO gastrocnemius shows higher Akt, GSK3β, S6, and 4E-BP1 phosphorylation and lower MuRF-1 and myostatin levels compared to wild-type, establishing CTMP as a regulator of Akt-dependent protein synthesis and ubiquitin-ligase-mediated catabolism in muscle.\",\n      \"method\": \"CTMP knockout mice, sciatic nerve injury model, Western blot (Akt, GSK3β, S6, 4E-BP1, MuRF-1, myostatin phosphorylation/expression), muscle weight measurement\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO with multiple defined biochemical and functional readouts in vivo, single lab\",\n      \"pmids\": [\"36652767\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PRAME, acting as a substrate-recognizing subunit of a Cul2-dependent E3 ubiquitin ligase, interacts with CTMP and p21, mediating their ubiquitination and proteasomal degradation. This leads to accumulation of phospho-Akt and CCND3, promoting proliferation in multiple myeloma cells.\",\n      \"method\": \"Co-immunoprecipitation (PRAME–CTMP interaction), ubiquitination assays, PRAME knockdown/overexpression, Western blot (p-Akt, CTMP, p21, CCND3), proliferation assays\",\n      \"journal\": \"Heliyon\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP and ubiquitination assays with functional KD/OE readouts, single lab\",\n      \"pmids\": [\"39071619\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"CARD9 interacts with THEM4 in cholangiocarcinoma cells, and this interaction facilitates AKT and mTOR phosphorylation, promoting CCA cell proliferation and invasion.\",\n      \"method\": \"Co-immunoprecipitation or interaction assay (CARD9–THEM4), Western blot (AKT, mTOR phosphorylation), gain/loss-of-function in CCA cell lines and nude mouse models\",\n      \"journal\": \"International immunopharmacology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, interaction and downstream signaling described in abstract without detailed mechanistic dissection of THEM4's direct contribution\",\n      \"pmids\": [\"39418733\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CAMK2A facilitates THEM4 release from mitochondria in pancreatic ductal adenocarcinoma cells. Released THEM4 inhibits AKT phosphorylation and suppresses PDAC tumor growth; THEM4 knockdown accelerates in vivo tumor growth.\",\n      \"method\": \"Functional experiments (mitochondrial membrane potential, ATP, ROS assays), THEM4 KD and xenograft tumor growth, AKT phosphorylation Western blot, immunohistochemical microarray\",\n      \"journal\": \"Journal of translational medicine\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, abstract-level description of mechanism without detailed mechanistic methods for CAMK2A→THEM4 release step\",\n      \"pmids\": [\"41353164\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CTMP knockdown enhances sensitivity of endometrial cancer cells to medroxyprogesterone acetate (MPA) by suppressing the PI3K/AKT signaling pathway, as assessed by cell proliferation assays and Western blot of PI3K/AKT pathway proteins.\",\n      \"method\": \"siRNA knockdown of CTMP, Western blot (PI3K/AKT pathway proteins), CCK-8, EDU incorporation assay, immunohistochemistry in clinical samples\",\n      \"journal\": \"Reproductive sciences\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, KD with signaling readouts but no direct mechanistic interrogation of CTMP–AKT interaction\",\n      \"pmids\": [\"41201690\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"In MSI-L/MSS colorectal cancer, CTMP interacts with REV7 (a novel high-affinity interaction identified by immunoprecipitation-mass spectrometry and validated by co-IP, GST pulldown with truncation/point mutants, and AlphaFold Multimer structural modeling). This CTMP–REV7 interaction facilitates CDK1-mediated G2/M progression and suppresses HLA-B/MHC-I expression. CTMP also sustains AKT/PD-L1 signaling and promotes fatty acid metabolism. CTMP knockdown diminishes CDK1 activity, induces G2/M arrest, restores HLA-B expression via cGAS signaling, enhances CD8+ T cell infiltration, and synergizes with REV7 overexpression and IFN-γ.\",\n      \"method\": \"Immunoprecipitation-mass spectrometry, co-IP, GST pulldown with truncation/point mutants, AlphaFold Multimer, CTMP KD/OE (in vitro and syngeneic mouse model), cell cycle analysis, flow cytometry (CD8+ T cells), HLA-B/MHC-I expression assays, AKT/PD-L1 Western blot\",\n      \"journal\": \"Cellular oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — structural validation (AlphaFold + mutagenesis pulldown), MS-identified interaction confirmed by co-IP and GST pulldown, functional in vivo syngeneic model, single lab\",\n      \"pmids\": [\"42143164\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"THEM4/CTMP is a mitochondrial protein that functions as an endogenous negative regulator of PKB/Akt by binding to Akt's carboxyl-terminal regulatory domain and inhibiting its phosphorylation at both Ser473 and Thr308; it localizes to the mitochondrial inter-membrane space and membrane, is released to the cytosol during apoptosis to sensitize cells to caspase-dependent death, regulates mitochondrial fission dynamics, and is transcriptionally controlled by the ATF3/NF-κB axis; its activity is modulated by PRAME-mediated ubiquitination and proteasomal degradation; and through a newly identified interaction with REV7 it also regulates CDK1-driven cell cycle progression and MHC-I antigen presentation, collectively placing THEM4 at a signaling nexus connecting Akt activity, mitochondrial homeostasis, and immune evasion.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"THEM4 (CTMP) is a mitochondrial protein that acts as an endogenous negative regulator of PKB/Akt signaling by binding the carboxyl-terminal regulatory domain of Akt and suppressing its phosphorylation at both Ser473 and Thr308, thereby reverting Akt-driven transformation [#0]. It resides in the mitochondrial inter-membrane space as both membrane-bound and free pools, and upon apoptotic stimulation is released to the cytosol where it amplifies mitochondrial depolarization and caspase-3/PARP cleavage; a mitochondrially retained mutant loses this pro-apoptotic activity, coupling its subcellular release to cell-death sensitization [#1]. Independent of apoptosis, THEM4 governs mitochondrial morphology, promoting fission-like clustering when present and yielding swollen, interconnected mitochondria when depleted, a phenotype confirmed in knockout mouse liver [#2]. Through its Akt-inhibitory function THEM4 shapes injury and disease responses: it is induced in vulnerable neurons during cerebral ischemia and traumatic brain injury, where its depletion restores Akt activity and rescues neurons or improves recovery [#3, #8, #10], and it regulates Akt-dependent protein synthesis versus ubiquitin-ligase catabolism in skeletal muscle, with its loss mitigating denervation-induced atrophy [#13]. THEM4 expression and protein levels are controlled by an ATF3/NF-\\u03baB transcriptional axis [#7], by miR-183-5p [#11], and by PRAME-directed Cul2 E3 ligase ubiquitination and proteasomal degradation [#14], while its interaction with LETM1 modulates its Akt-suppressing output [#5, #6]. A high-affinity interaction with REV7 extends THEM4 function beyond Akt, driving CDK1-dependent G2/M progression and suppressing HLA-B/MHC-I antigen presentation, positioning it at a nexus linking Akt activity, mitochondrial homeostasis, cell cycle, and immune evasion [#18].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Established THEM4/CTMP as a direct, physiological negative regulator of Akt, defining its founding molecular function.\",\n      \"evidence\": \"Binding assays, kinase and phosphorylation assays, and reversion of v-Akt transformation\",\n      \"pmids\": [\"11598301\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve where in the cell the inhibitory interaction predominates\", \"Structural basis of binding to the Akt C-terminal domain not defined\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Resolved THEM4's mitochondrial localization and showed its apoptotic release from the inter-membrane space is required for pro-apoptotic activity, linking compartmentalization to function.\",\n      \"evidence\": \"Subcellular fractionation, live-cell imaging, mitochondrial membrane potential and caspase assays, retention mutant, RNAi\",\n      \"pmids\": [\"19168129\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of release from mitochondria not defined\", \"Relationship between mitochondrial pool and plasma-membrane Akt inhibition unclear\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Demonstrated a distinct role for THEM4 in controlling mitochondrial fission/morphology, separating this from its apoptotic function.\",\n      \"evidence\": \"Fluorescence microscopy of mitochondrial morphology, RNAi, cleavage-resistant mutant overexpression, CTMP-KO mouse liver\",\n      \"pmids\": [\"19421406\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular fission machinery engaged by THEM4 not identified\", \"Whether morphology control requires Akt inhibition is unresolved\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Showed THEM4 inhibition of Akt is functionally consequential in vivo, where its induction kills ischemic neurons and its depletion is protective.\",\n      \"evidence\": \"Rat global ischemia model, co-IP, kinase assays, downstream GSK-3\\u03b2/FOXO3A readouts, lentiviral RNAi rescue; cell-permeable TAT-CTMP peptide in pancreatic cancer xenografts\",\n      \"pmids\": [\"19349976\", \"19405118\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Upstream signal triggering THEM4 induction in ischemia not fully defined\", \"Peptide tumor-selectivity mechanism not dissected\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Identified the transcriptional control of THEM4 through competing ATF3 repression and NF-\\u03baB activation at its promoter, explaining its injury-induced expression.\",\n      \"evidence\": \"Reporter and ChIP assays, ATF3-VP16 and degradation-resistant I\\u03baB\\u03b1 constructs, siRNA, p-Akt readouts; TBI knockdown model with functional scoring\",\n      \"pmids\": [\"24771044\", \"24670215\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signals selecting ATF3 versus NF-\\u03baB occupancy not defined\", \"Generality of this axis beyond neural injury untested\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Placed THEM4 in a metabolic/oncogenic Akt context and identified LETM1 as a binding partner that modulates THEM4's Akt-suppressing output.\",\n      \"evidence\": \"Overexpression in HEK293 and obese mouse adipose tissue; co-IP and co-delivery of LETM1/CTMP in an HCC mouse model with phosphorylation and apoptosis readouts\",\n      \"pmids\": [\"24333006\", \"23392203\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which LETM1 antagonizes or directs THEM4 not defined\", \"Direct versus indirect LETM1\\u2013THEM4 binding not fully characterized\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Extended THEM4 regulation to diabetic and excitotoxic injury and raised the possibility of phosphorylation-dependent biphasic control of its activity.\",\n      \"evidence\": \"db/db focal ischemia model with siRNA rescue and infarct measurement; kainic acid model and primary astrocyte LPS/IFN-\\u03b3 stimulation tracking p-CTMP/p-Akt/p-CREB\",\n      \"pmids\": [\"27161366\", \"28243164\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The p-CTMP phosphosite and responsible kinase were not directly manipulated\", \"Astrocyte data are correlative without mechanistic perturbation\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Documented post-transcriptional and proteostatic control of THEM4 abundance and embedded it in cancer signaling networks.\",\n      \"evidence\": \"miR-183-5p target validation with rescue in colon cancer; LETM1-silencing epistasis in endometrial cancer; PRAME/Cul2 co-IP and ubiquitination assays in myeloma\",\n      \"pmids\": [\"34249713\", \"35324530\", \"39071619\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Context-dependence of these regulators across tissues unresolved\", \"Whether THEM4 degradation kinetics directly set Akt output not quantified\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined THEM4 as a physiological brake on Akt-dependent anabolism and catabolism in skeletal muscle using genetic knockout.\",\n      \"evidence\": \"CTMP-KO mice with sciatic nerve denervation, Western blot of Akt/GSK3\\u03b2/S6/4E-BP1 and MuRF-1/myostatin, muscle weight\",\n      \"pmids\": [\"36652767\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether muscle phenotype is cell-autonomous to myofibers not established\", \"Mitochondrial contribution in muscle not examined\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Discovered a high-affinity THEM4\\u2013REV7 interaction that couples THEM4 to CDK1-driven cell cycle progression and MHC-I/antigen-presentation control, expanding its role into immune evasion.\",\n      \"evidence\": \"IP-mass spectrometry, co-IP, GST pulldown with truncation/point mutants, AlphaFold Multimer, CTMP KD/OE in vitro and syngeneic mouse model, cell cycle and CD8+ T cell flow cytometry, HLA-B/AKT/PD-L1 readouts\",\n      \"pmids\": [\"42143164\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether REV7 binding is mechanistically separable from Akt regulation unclear\", \"cGAS-dependent HLA-B restoration pathway not fully dissected\", \"Single lab; reciprocal validation in other tumor types pending\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How THEM4's mitochondrial residence, its release to the cytosol, and its direct Akt and REV7 interactions are mechanistically integrated into a single regulatory logic remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structure of the THEM4\\u2013Akt complex\", \"Trigger and machinery for mitochondrial release not defined\", \"Whether Akt-inhibitory and REV7/cell-cycle functions are independent or coupled is unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 3, 18]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [1, 2]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 3]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [1, 4]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [18]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"AKT1\", \"LETM1\", \"REV7\", \"PRAME\", \"CARD9\", \"CAMK2A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}