{"gene":"STYXL1","run_date":"2026-06-10T10:51:54","timeline":{"discoveries":[{"year":2010,"finding":"MK-STYX (STYXL1) is catalytically inactive due to replacement of the nucleophilic cysteine and adjacent histidine in the dual-specificity phosphatase signature motif with serine and phenylalanine; reintroducing histidine and cysteine via mutagenesis restored phosphatase activity. MK-STYX binds G3BP1 (identified by MS), and this interaction is dramatically reduced in the catalytically active mutant. Expression of wild-type MK-STYX inhibits stress granule formation induced by arsenite or G3BP1 overexpression, whereas the active mutant is impaired in this inhibition.","method":"Active-site mutagenesis, MS-based binding partner identification, stress granule imaging in transfected cells","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — active-site mutagenesis establishing catalytic mechanism, MS identification of binding partner, and functional cell-based assay; multiple orthogonal methods in one study","pmids":["20180778"],"is_preprint":false},{"year":2011,"finding":"RNAi-mediated knockdown of MK-STYX inhibits mitochondrion-dependent apoptosis: cells depleted of MK-STYX cannot release cytochrome c in response to pro-apoptotic BCL-2 family members (Bax, Bid, Bim), placing the block at mitochondrial outer membrane permeabilization (MOMP). MK-STYX localizes to mitochondria but is not released upon apoptotic stress. MK-STYX knockdown does not significantly alter MAPK signaling in response to growth factors or apoptotic stimuli.","method":"RNAi knockdown, cytochrome c release assay, subcellular fractionation/localization, apoptosis assays","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — genome-wide RNAi screen followed by targeted knockdown with multiple orthogonal functional readouts (cytochrome c release, MOMP, apoptosis, localization) in single rigorous study","pmids":["21262771"],"is_preprint":false},{"year":2012,"finding":"MK-STYX inhibits stress granule assembly independently of G3BP-1 phosphorylation at Ser149. The active-site mutant MK-STYX(active) (with histidine and cysteine introduced) not only induces stress granules but also dephosphorylates G3BP-1, demonstrating that the pseudo-catalytic nature of wild-type MK-STYX is essential for its inhibitory function on stress granules.","method":"Phosphomimetic and non-phosphorylatable G3BP-1 mutants, MK-STYX active-site point mutants, stress granule imaging","journal":"The FEBS journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — active-site mutagenesis combined with phosphomutant constructs and functional stress granule assays; single lab, two orthogonal approaches","pmids":["23163895"],"is_preprint":false},{"year":2014,"finding":"MK-STYX physically interacts with the mitochondrial phosphatase PTPMT1 (identified by unbiased proteomics) and suppresses PTPMT1 catalytic activity. Knockdown of PTPMT1 resensitizes MK-STYX-knockdown cells to chemotherapeutics and restores cytochrome c release, supporting a model in which MK-STYX controls mitochondrial apoptosis by negatively regulating PTPMT1.","method":"Unbiased proteomics (interaction screen), co-immunoprecipitation, PTPMT1 activity assay, RNAi epistasis (double knockdown), cytochrome c release assay","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Moderate — proteomics-identified interaction confirmed by co-IP, enzymatic activity assay, and genetic epistasis (double knockdown) with defined functional readout; multiple orthogonal methods","pmids":["24709986"],"is_preprint":false},{"year":2014,"finding":"MK-STYX, but not its catalytically active mutant, induces neurite-like outgrowths in PC12 cells and enhances NGF-induced neurite extensions. This occurs independently of the Ras-ERK/MAPK pathway (outgrowths persist with MEK inhibitor). MK-STYX decreases RhoA activation; RhoA activity increases when MK-STYX is downregulated. MK-STYX also affects the downstream RhoA effector cofilin: it decreases cofilin phosphorylation in unstimulated cells but increases it in NGF-stimulated cells.","method":"Overexpression and knockdown in PC12 cells, MEK inhibitor treatment, RhoA activity assay, cofilin phosphorylation immunoblot","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — epistasis via pharmacological inhibitor, RhoA activity assay, and phospho-cofilin immunoblot in single lab with multiple orthogonal methods","pmids":["25479605"],"is_preprint":false},{"year":2017,"finding":"MK-STYX-induced outgrowths in PC12 cells contain microtubules and microfilaments, form synapses (confirmed by TEM), and express both axonal (Tau-1) and dendritic (MAP2) markers with dopaminergic character. MK-STYX modulates actin expression (immunoblot) and increases growth cone formation in response to NGF. In rat hippocampal primary neurons, MK-STYX overexpression increases the number of primary neurites.","method":"Immunofluorescence, transmission electron microscopy, immunoblotting, primary neuron culture with overexpression","journal":"Frontiers in molecular biosciences","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — TEM for synapse confirmation is Tier 1; neurite marker and actin immunoblot are Tier 3; multiple orthogonal methods in single lab","pmids":["29250526"],"is_preprint":false},{"year":2019,"finding":"MK-STYX alters HDAC6 subcellular localization: in control cells HDAC6 is cytosolic, but in MK-STYX-overexpressing cells HDAC6 is both nuclear and cytosolic. MK-STYX decreases HDAC6 serine phosphorylation, protein tyrosine phosphorylation, and lysine acetylation. MK-STYX increases tubulin acetylation and detyrosination without affecting microtubule organization.","method":"Overexpression in HEK293 cells, immunofluorescence localization, immunoblotting for post-translational modifications","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — localization and PTM changes measured by immunofluorescence and immunoblot; multiple readouts but single lab without mechanistic reconstitution","pmids":["30909412"],"is_preprint":false},{"year":2023,"finding":"The DUSP domain of MK-STYX (not the CH2 domain) is sufficient to interact with G3BP1 (confirmed by co-IP with both overexpressed and endogenous G3BP1) and to decrease stress granule formation. The DUSP domain decreases G3BP1 tyrosine phosphorylation (required for stress granule formation), while the CH2 domain increases stress granules and G3BP1 tyrosine phosphorylation.","method":"Co-immunoprecipitation of truncated domain constructs (DUSP, CH2) with G3BP1, stress granule imaging in HEK293 and HeLa cells, tyrosine phosphorylation immunoblot","journal":"Archives of biochemistry and biophysics","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — reciprocal domain-mapping Co-IP plus functional stress granule assay and PTM readout; single lab, two orthogonal methods","pmids":["37516290"],"is_preprint":false},{"year":2023,"finding":"STYXL1 knockdown enhances trafficking of β-glucocerebrosidase (β-GC) to lysosomes and increases lysosomal β-GC activity in HeLa cells. STYXL1 depletion causes nuclear translocation of UPR and lysosomal biogenesis transcription factors and increases ER–lysosome contacts; the enhanced β-GC activity is ER-stress-dependent (reversed by 4-PBA) but independent of TFEB/TFE3 nuclear localization.","method":"RNAi knockdown, lysosomal enzyme activity assay, nuclear localization imaging of transcription factors, 4-PBA pharmacological treatment, ER-lysosome contact imaging","journal":"Traffic (Copenhagen, Denmark)","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — functional lysosomal assay combined with pharmacological epistasis (4-PBA) and localization imaging; single lab, multiple orthogonal methods","pmids":["37198709"],"is_preprint":false},{"year":2024,"finding":"STYXL1 acts as a germ cell-specific co-factor of the CCT chaperonin complex. The N-terminal rhodanese-like domain of STYXL1 mediates interactions with CCT complex subunits CCT1, CCT6, and CCT7. Deletion of Styxl1 in mice causes male infertility with microtubule defects in sperm flagella, defects in CCT complex assembly, and reduced tubulin polymerization.","method":"Gene knockout in mice, proteomics of Styxl1−/− sperm, co-immunoprecipitation of STYXL1 with CCT subunits, domain mapping (N-terminal rhodanese-like domain), tubulin polymerization assay","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vivo knockout with defined male infertility phenotype, proteomic profiling, co-IP domain mapping, and biochemical tubulin polymerization assay; multiple orthogonal methods in single rigorous study","pmids":["38168070"],"is_preprint":false}],"current_model":"STYXL1/MK-STYX is a catalytically inactive dual-specificity pseudophosphatase whose DUSP domain binds G3BP1 and suppresses stress granule assembly by reducing G3BP1 tyrosine phosphorylation; it localizes to mitochondria where it negatively regulates the phosphatase PTPMT1 to control mitochondrial outer membrane permeabilization and cytochrome c release during apoptosis; it suppresses RhoA activity to promote neurite outgrowth and cytoskeletal remodeling; it alters HDAC6 localization and tubulin post-translational modifications; and in germ cells its N-terminal rhodanese-like domain associates with CCT chaperonin subunits to facilitate tubulin folding and sperm flagella formation."},"narrative":{"mechanistic_narrative":"STYXL1 (MK-STYX) is a catalytically inactive dual-specificity pseudophosphatase that acts as a protein-interaction adaptor controlling stress responses, mitochondrial apoptosis, cytoskeletal remodeling, and germ cell development [PMID:20180778, PMID:21262771, PMID:38168070]. Its catalytic dead state—the nucleophilic cysteine and adjacent histidine of the DUSP signature motif are replaced by serine and phenylalanine—is essential to its function: restoring these residues creates an active phosphatase that loses STYXL1's biological activities [PMID:20180778, PMID:23163895]. Through its DUSP domain it binds G3BP1 and suppresses stress granule assembly by lowering G3BP1 tyrosine phosphorylation, while the CH2 domain has the opposite effect of promoting stress granules and G3BP1 tyrosine phosphorylation [PMID:20180778, PMID:37516290]. At the mitochondrion STYXL1 is required for cytochrome c release and outer membrane permeabilization during BCL-2-family-driven apoptosis, acting by binding and suppressing the mitochondrial phosphatase PTPMT1, such that PTPMT1 depletion restores apoptotic sensitivity to STYXL1-depleted cells [PMID:21262771, PMID:24709986]. STYXL1 also drives neurite outgrowth in PC12 and primary neurons independently of Ras-ERK signaling by reducing RhoA activation and modulating the downstream effector cofilin [PMID:25479605, PMID:29250526], and it shifts HDAC6 localization and increases tubulin acetylation and detyrosination [PMID:30909412]. In germ cells, its N-terminal rhodanese-like domain associates with CCT chaperonin subunits CCT1, CCT6, and CCT7 to support tubulin folding; Styxl1 knockout mice are male infertile with sperm flagellar microtubule defects, impaired CCT assembly, and reduced tubulin polymerization [PMID:38168070].","teleology":[{"year":2010,"claim":"Established that STYXL1 is a genuine pseudophosphatase whose catalytic inactivity is functionally required, and identified G3BP1 as a partner through which it suppresses stress granules.","evidence":"Active-site mutagenesis restoring catalytic residues, MS-based partner identification, and stress granule imaging in transfected cells","pmids":["20180778"],"confidence":"High","gaps":["Did not define which STYXL1 domain mediates G3BP1 binding","Mechanism linking G3BP1 binding to stress granule suppression undefined"]},{"year":2011,"claim":"Placed STYXL1 as a positive regulator of mitochondrial apoptosis, with its requirement mapped to the outer membrane permeabilization step.","evidence":"RNAi knockdown with cytochrome c release, MOMP, and apoptosis readouts plus subcellular fractionation","pmids":["21262771"],"confidence":"High","gaps":["Molecular target at the mitochondrion not yet identified","How a pseudophosphatase controls MOMP unresolved"]},{"year":2012,"claim":"Refined the stress granule mechanism by showing suppression is independent of G3BP1 Ser149 phosphorylation and depends on STYXL1's pseudo-catalytic state.","evidence":"Phosphomimetic/non-phosphorylatable G3BP1 mutants and STYXL1 active-site mutants with stress granule imaging","pmids":["23163895"],"confidence":"Medium","gaps":["The relevant phosphosite STYXL1 affects on G3BP1 not pinpointed here","Single-lab evidence"]},{"year":2014,"claim":"Identified PTPMT1 as the mitochondrial effector through which STYXL1 controls apoptosis, completing the MOMP mechanism.","evidence":"Unbiased proteomics, co-IP, PTPMT1 activity assay, and RNAi epistasis with cytochrome c readout","pmids":["24709986"],"confidence":"High","gaps":["How catalytically dead STYXL1 suppresses PTPMT1 activity mechanistically unknown","Structural basis of the interaction undetermined"]},{"year":2014,"claim":"Connected STYXL1 to cytoskeletal control in neurons, showing it promotes neurite outgrowth via RhoA suppression rather than MAPK signaling.","evidence":"Overexpression/knockdown in PC12 cells, MEK inhibitor epistasis, RhoA activity assay, phospho-cofilin immunoblot","pmids":["25479605"],"confidence":"Medium","gaps":["Direct link between STYXL1 and RhoA regulation not established","Biphasic cofilin phosphorylation effect unexplained"]},{"year":2017,"claim":"Characterized STYXL1-induced outgrowths as bona fide neuritic structures forming synapses, extending the neuronal differentiation role to primary neurons.","evidence":"Immunofluorescence, TEM synapse confirmation, neurite marker immunoblots, primary hippocampal neuron culture","pmids":["29250526"],"confidence":"Medium","gaps":["Molecular pathway driving synapse formation not defined","In vivo neuronal relevance untested"]},{"year":2019,"claim":"Linked STYXL1 to tubulin post-translational modification and HDAC6 regulation, suggesting a route to cytoskeletal control.","evidence":"Overexpression in HEK293, immunofluorescence localization, PTM immunoblots","pmids":["30909412"],"confidence":"Medium","gaps":["Whether STYXL1 acts directly on HDAC6 unknown","Functional consequence of altered tubulin modification untested"]},{"year":2023,"claim":"Mapped opposing stress granule functions to STYXL1's DUSP versus CH2 domains and identified G3BP1 tyrosine phosphorylation as the relevant modification.","evidence":"Reciprocal domain-mapping co-IP with endogenous and overexpressed G3BP1, stress granule imaging, tyrosine phosphorylation immunoblot","pmids":["37516290"],"confidence":"Medium","gaps":["The tyrosine kinase/phosphatase STYXL1 modulates not identified","Single-lab evidence"]},{"year":2023,"claim":"Revealed a role for STYXL1 in lysosomal trafficking and ER stress, linking its depletion to enhanced β-glucocerebrosidase activity.","evidence":"RNAi knockdown, lysosomal enzyme assay, transcription factor localization imaging, 4-PBA epistasis, ER-lysosome contact imaging","pmids":["37198709"],"confidence":"Medium","gaps":["Direct molecular substrate or partner in this pathway unknown","Relationship to STYXL1's mitochondrial/stress granule roles unclear"]},{"year":2024,"claim":"Defined an in vivo physiological role: STYXL1 is a germ-cell CCT chaperonin co-factor required for tubulin folding and sperm flagellar assembly.","evidence":"Mouse knockout with male infertility phenotype, sperm proteomics, co-IP domain mapping to N-terminal rhodanese-like domain, tubulin polymerization assay","pmids":["38168070"],"confidence":"High","gaps":["Whether STYXL1 modulates CCT activity in non-germ tissues unknown","How rhodanese-like domain engagement of CCT promotes folding mechanistically undefined"]},{"year":null,"claim":"It remains unresolved how a single catalytically dead pseudophosphatase integrates its distinct roles in stress granules, mitochondrial apoptosis, cytoskeletal/neuronal remodeling, lysosomal trafficking, and CCT-dependent tubulin folding.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unifying structural or regulatory model across the reported functions","Tissue- and context-specificity of each function not delineated"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,3,7]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,9]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[6,9]}],"localization":[{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[1,3]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[0,7]}],"pathway":[{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[1,3]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,7]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[4,5,9]}],"complexes":["CCT/TRiC chaperonin"],"partners":["G3BP1","PTPMT1","HDAC6","CCT1","CCT6","CCT7"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y6J8","full_name":"Serine/threonine/tyrosine-interacting-like protein 1","aliases":["Dual specificity phosphatase inhibitor MK-STYX","Dual specificity protein phosphatase 24","Inactive dual specificity protein phosphatase MK-STYX","Map kinase phosphatase-like protein MK-STYX"],"length_aa":313,"mass_kda":35.8,"function":"Catalytically inactive phosphatase (PubMed:20180778, PubMed:23163895). By binding to G3BP1, inhibits the formation of G3BP1-induced stress granules (PubMed:20180778, PubMed:23163895). Does not act by protecting the dephosphorylation of G3BP1 at 'Ser-149' (PubMed:23163895). Inhibits PTPMT1 phosphatase activity (PubMed:24709986). By inhibiting PTPMT1, positively regulates intrinsic apoptosis (PubMed:21262771). May play a role in the formation of neurites during neuronal development (PubMed:29250526)","subcellular_location":"Mitochondrion matrix","url":"https://www.uniprot.org/uniprotkb/Q9Y6J8/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/STYXL1","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/STYXL1","total_profiled":1310},"omim":[{"mim_id":"616695","title":"SERINE/THREONINE/TYROSINE-INTERACTING PROTEIN-LIKE 1; STYXL1","url":"https://www.omim.org/entry/616695"}],"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/STYXL1"},"hgnc":{"alias_symbol":["MK-STYX"],"prev_symbol":["DUSP24"]},"alphafold":{"accession":"Q9Y6J8","domains":[{"cath_id":"3.40.250.10","chopping":"5-88_106-156","consensus_level":"high","plddt":90.9727,"start":5,"end":156},{"cath_id":"3.90.190.10","chopping":"162-302","consensus_level":"high","plddt":97.2473,"start":162,"end":302}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y6J8","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y6J8-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y6J8-F1-predicted_aligned_error_v6.png","plddt_mean":90.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=STYXL1","jax_strain_url":"https://www.jax.org/strain/search?query=STYXL1"},"sequence":{"accession":"Q9Y6J8","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y6J8.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y6J8/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y6J8"}},"corpus_meta":[{"pmid":"20180778","id":"PMC_20180778","title":"The pseudophosphatase MK-STYX interacts with G3BP and decreases stress granule formation.","date":"2010","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/20180778","citation_count":40,"is_preprint":false},{"pmid":"21262771","id":"PMC_21262771","title":"MK-STYX, a catalytically inactive phosphatase regulating mitochondrially dependent apoptosis.","date":"2011","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/21262771","citation_count":32,"is_preprint":false},{"pmid":"23163895","id":"PMC_23163895","title":"The pseudophosphatase MK-STYX inhibits stress granule assembly independently of Ser149 phosphorylation of G3BP-1.","date":"2012","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/23163895","citation_count":26,"is_preprint":false},{"pmid":"24709986","id":"PMC_24709986","title":"The pseudophosphatase MK-STYX physically and genetically interacts with the mitochondrial phosphatase PTPMT1.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/24709986","citation_count":24,"is_preprint":false},{"pmid":"38168070","id":"PMC_38168070","title":"STYXL1 regulates CCT complex assembly and flagellar tubulin folding in sperm formation.","date":"2024","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/38168070","citation_count":18,"is_preprint":false},{"pmid":"32271415","id":"PMC_32271415","title":"STYXL1 promotes malignant progression of hepatocellular carcinoma via downregulating CELF2 through the PI3K/Akt pathway.","date":"2020","source":"European review for medical and pharmacological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/32271415","citation_count":18,"is_preprint":false},{"pmid":"25479605","id":"PMC_25479605","title":"The pseudophosphatase MK-STYX induces neurite-like outgrowths in PC12 cells.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/25479605","citation_count":17,"is_preprint":false},{"pmid":"29250526","id":"PMC_29250526","title":"MK-STYX Alters the Morphology of Primary Neurons, and Outgrowths in MK-STYX Overexpressing PC-12 Cells Develop a Neuronal Phenotype.","date":"2017","source":"Frontiers in molecular biosciences","url":"https://pubmed.ncbi.nlm.nih.gov/29250526","citation_count":15,"is_preprint":false},{"pmid":"32472731","id":"PMC_32472731","title":"Pseudophosphatase MK-STYX: the atypical member of the MAP kinase phosphatases.","date":"2020","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/32472731","citation_count":12,"is_preprint":false},{"pmid":"25724587","id":"PMC_25724587","title":"Homozygous missense mutation in STYXL1 associated with moderate intellectual disability, epilepsy and behavioural complexities.","date":"2015","source":"European journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/25724587","citation_count":12,"is_preprint":false},{"pmid":"31146910","id":"PMC_31146910","title":"Serine/threonine/tyrosine-interacting-like protein 1 (STYXL1), a pseudo phosphatase, promotes oncogenesis in glioma.","date":"2019","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/31146910","citation_count":11,"is_preprint":false},{"pmid":"30909412","id":"PMC_30909412","title":"Pseudophosphatase MK-STYX Alters Histone Deacetylase 6 Cytoplasmic Localization, Decreases Its Phosphorylation, and Increases Detyrosination of Tubulin.","date":"2019","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/30909412","citation_count":10,"is_preprint":false},{"pmid":"35264672","id":"PMC_35264672","title":"Evolutionary genomic relationships and coupling in MK-STYX and STYX pseudophosphatases.","date":"2022","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/35264672","citation_count":3,"is_preprint":false},{"pmid":"37198709","id":"PMC_37198709","title":"Pseudophosphatase STYXL1 depletion enhances glucocerebrosidase trafficking to lysosomes via ER stress.","date":"2023","source":"Traffic (Copenhagen, Denmark)","url":"https://pubmed.ncbi.nlm.nih.gov/37198709","citation_count":3,"is_preprint":false},{"pmid":"30472486","id":"PMC_30472486","title":"A previously identified missense mutation in STYXL1 is likely benign.","date":"2018","source":"European journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/30472486","citation_count":2,"is_preprint":false},{"pmid":"37516290","id":"PMC_37516290","title":"The DUSP domain of pseudophosphatase MK-STYX interacts with G3BP1 to decrease stress granules.","date":"2023","source":"Archives of biochemistry and biophysics","url":"https://pubmed.ncbi.nlm.nih.gov/37516290","citation_count":1,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":10820,"output_tokens":3084,"usd":0.03936,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10426,"output_tokens":3456,"usd":0.069265,"stage2_stop_reason":"end_turn"},"total_usd":0.108625,"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\": 2010,\n      \"finding\": \"MK-STYX (STYXL1) is catalytically inactive due to replacement of the nucleophilic cysteine and adjacent histidine in the dual-specificity phosphatase signature motif with serine and phenylalanine; reintroducing histidine and cysteine via mutagenesis restored phosphatase activity. MK-STYX binds G3BP1 (identified by MS), and this interaction is dramatically reduced in the catalytically active mutant. Expression of wild-type MK-STYX inhibits stress granule formation induced by arsenite or G3BP1 overexpression, whereas the active mutant is impaired in this inhibition.\",\n      \"method\": \"Active-site mutagenesis, MS-based binding partner identification, stress granule imaging in transfected cells\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — active-site mutagenesis establishing catalytic mechanism, MS identification of binding partner, and functional cell-based assay; multiple orthogonal methods in one study\",\n      \"pmids\": [\"20180778\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"RNAi-mediated knockdown of MK-STYX inhibits mitochondrion-dependent apoptosis: cells depleted of MK-STYX cannot release cytochrome c in response to pro-apoptotic BCL-2 family members (Bax, Bid, Bim), placing the block at mitochondrial outer membrane permeabilization (MOMP). MK-STYX localizes to mitochondria but is not released upon apoptotic stress. MK-STYX knockdown does not significantly alter MAPK signaling in response to growth factors or apoptotic stimuli.\",\n      \"method\": \"RNAi knockdown, cytochrome c release assay, subcellular fractionation/localization, apoptosis assays\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genome-wide RNAi screen followed by targeted knockdown with multiple orthogonal functional readouts (cytochrome c release, MOMP, apoptosis, localization) in single rigorous study\",\n      \"pmids\": [\"21262771\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"MK-STYX inhibits stress granule assembly independently of G3BP-1 phosphorylation at Ser149. The active-site mutant MK-STYX(active) (with histidine and cysteine introduced) not only induces stress granules but also dephosphorylates G3BP-1, demonstrating that the pseudo-catalytic nature of wild-type MK-STYX is essential for its inhibitory function on stress granules.\",\n      \"method\": \"Phosphomimetic and non-phosphorylatable G3BP-1 mutants, MK-STYX active-site point mutants, stress granule imaging\",\n      \"journal\": \"The FEBS journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — active-site mutagenesis combined with phosphomutant constructs and functional stress granule assays; single lab, two orthogonal approaches\",\n      \"pmids\": [\"23163895\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"MK-STYX physically interacts with the mitochondrial phosphatase PTPMT1 (identified by unbiased proteomics) and suppresses PTPMT1 catalytic activity. Knockdown of PTPMT1 resensitizes MK-STYX-knockdown cells to chemotherapeutics and restores cytochrome c release, supporting a model in which MK-STYX controls mitochondrial apoptosis by negatively regulating PTPMT1.\",\n      \"method\": \"Unbiased proteomics (interaction screen), co-immunoprecipitation, PTPMT1 activity assay, RNAi epistasis (double knockdown), cytochrome c release assay\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proteomics-identified interaction confirmed by co-IP, enzymatic activity assay, and genetic epistasis (double knockdown) with defined functional readout; multiple orthogonal methods\",\n      \"pmids\": [\"24709986\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"MK-STYX, but not its catalytically active mutant, induces neurite-like outgrowths in PC12 cells and enhances NGF-induced neurite extensions. This occurs independently of the Ras-ERK/MAPK pathway (outgrowths persist with MEK inhibitor). MK-STYX decreases RhoA activation; RhoA activity increases when MK-STYX is downregulated. MK-STYX also affects the downstream RhoA effector cofilin: it decreases cofilin phosphorylation in unstimulated cells but increases it in NGF-stimulated cells.\",\n      \"method\": \"Overexpression and knockdown in PC12 cells, MEK inhibitor treatment, RhoA activity assay, cofilin phosphorylation immunoblot\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis via pharmacological inhibitor, RhoA activity assay, and phospho-cofilin immunoblot in single lab with multiple orthogonal methods\",\n      \"pmids\": [\"25479605\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"MK-STYX-induced outgrowths in PC12 cells contain microtubules and microfilaments, form synapses (confirmed by TEM), and express both axonal (Tau-1) and dendritic (MAP2) markers with dopaminergic character. MK-STYX modulates actin expression (immunoblot) and increases growth cone formation in response to NGF. In rat hippocampal primary neurons, MK-STYX overexpression increases the number of primary neurites.\",\n      \"method\": \"Immunofluorescence, transmission electron microscopy, immunoblotting, primary neuron culture with overexpression\",\n      \"journal\": \"Frontiers in molecular biosciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — TEM for synapse confirmation is Tier 1; neurite marker and actin immunoblot are Tier 3; multiple orthogonal methods in single lab\",\n      \"pmids\": [\"29250526\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"MK-STYX alters HDAC6 subcellular localization: in control cells HDAC6 is cytosolic, but in MK-STYX-overexpressing cells HDAC6 is both nuclear and cytosolic. MK-STYX decreases HDAC6 serine phosphorylation, protein tyrosine phosphorylation, and lysine acetylation. MK-STYX increases tubulin acetylation and detyrosination without affecting microtubule organization.\",\n      \"method\": \"Overexpression in HEK293 cells, immunofluorescence localization, immunoblotting for post-translational modifications\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — localization and PTM changes measured by immunofluorescence and immunoblot; multiple readouts but single lab without mechanistic reconstitution\",\n      \"pmids\": [\"30909412\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"The DUSP domain of MK-STYX (not the CH2 domain) is sufficient to interact with G3BP1 (confirmed by co-IP with both overexpressed and endogenous G3BP1) and to decrease stress granule formation. The DUSP domain decreases G3BP1 tyrosine phosphorylation (required for stress granule formation), while the CH2 domain increases stress granules and G3BP1 tyrosine phosphorylation.\",\n      \"method\": \"Co-immunoprecipitation of truncated domain constructs (DUSP, CH2) with G3BP1, stress granule imaging in HEK293 and HeLa cells, tyrosine phosphorylation immunoblot\",\n      \"journal\": \"Archives of biochemistry and biophysics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — reciprocal domain-mapping Co-IP plus functional stress granule assay and PTM readout; single lab, two orthogonal methods\",\n      \"pmids\": [\"37516290\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"STYXL1 knockdown enhances trafficking of β-glucocerebrosidase (β-GC) to lysosomes and increases lysosomal β-GC activity in HeLa cells. STYXL1 depletion causes nuclear translocation of UPR and lysosomal biogenesis transcription factors and increases ER–lysosome contacts; the enhanced β-GC activity is ER-stress-dependent (reversed by 4-PBA) but independent of TFEB/TFE3 nuclear localization.\",\n      \"method\": \"RNAi knockdown, lysosomal enzyme activity assay, nuclear localization imaging of transcription factors, 4-PBA pharmacological treatment, ER-lysosome contact imaging\",\n      \"journal\": \"Traffic (Copenhagen, Denmark)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — functional lysosomal assay combined with pharmacological epistasis (4-PBA) and localization imaging; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"37198709\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"STYXL1 acts as a germ cell-specific co-factor of the CCT chaperonin complex. The N-terminal rhodanese-like domain of STYXL1 mediates interactions with CCT complex subunits CCT1, CCT6, and CCT7. Deletion of Styxl1 in mice causes male infertility with microtubule defects in sperm flagella, defects in CCT complex assembly, and reduced tubulin polymerization.\",\n      \"method\": \"Gene knockout in mice, proteomics of Styxl1−/− sperm, co-immunoprecipitation of STYXL1 with CCT subunits, domain mapping (N-terminal rhodanese-like domain), tubulin polymerization assay\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vivo knockout with defined male infertility phenotype, proteomic profiling, co-IP domain mapping, and biochemical tubulin polymerization assay; multiple orthogonal methods in single rigorous study\",\n      \"pmids\": [\"38168070\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"STYXL1/MK-STYX is a catalytically inactive dual-specificity pseudophosphatase whose DUSP domain binds G3BP1 and suppresses stress granule assembly by reducing G3BP1 tyrosine phosphorylation; it localizes to mitochondria where it negatively regulates the phosphatase PTPMT1 to control mitochondrial outer membrane permeabilization and cytochrome c release during apoptosis; it suppresses RhoA activity to promote neurite outgrowth and cytoskeletal remodeling; it alters HDAC6 localization and tubulin post-translational modifications; and in germ cells its N-terminal rhodanese-like domain associates with CCT chaperonin subunits to facilitate tubulin folding and sperm flagella formation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"STYXL1 (MK-STYX) is a catalytically inactive dual-specificity pseudophosphatase that acts as a protein-interaction adaptor controlling stress responses, mitochondrial apoptosis, cytoskeletal remodeling, and germ cell development [#0, #1, #9]. Its catalytic dead state—the nucleophilic cysteine and adjacent histidine of the DUSP signature motif are replaced by serine and phenylalanine—is essential to its function: restoring these residues creates an active phosphatase that loses STYXL1's biological activities [#0, #2]. Through its DUSP domain it binds G3BP1 and suppresses stress granule assembly by lowering G3BP1 tyrosine phosphorylation, while the CH2 domain has the opposite effect of promoting stress granules and G3BP1 tyrosine phosphorylation [#0, #7]. At the mitochondrion STYXL1 is required for cytochrome c release and outer membrane permeabilization during BCL-2-family-driven apoptosis, acting by binding and suppressing the mitochondrial phosphatase PTPMT1, such that PTPMT1 depletion restores apoptotic sensitivity to STYXL1-depleted cells [#1, #3]. STYXL1 also drives neurite outgrowth in PC12 and primary neurons independently of Ras-ERK signaling by reducing RhoA activation and modulating the downstream effector cofilin [#4, #5], and it shifts HDAC6 localization and increases tubulin acetylation and detyrosination [#6]. In germ cells, its N-terminal rhodanese-like domain associates with CCT chaperonin subunits CCT1, CCT6, and CCT7 to support tubulin folding; Styxl1 knockout mice are male infertile with sperm flagellar microtubule defects, impaired CCT assembly, and reduced tubulin polymerization [#9].\",\n  \"teleology\": [\n    {\n      \"year\": 2010,\n      \"claim\": \"Established that STYXL1 is a genuine pseudophosphatase whose catalytic inactivity is functionally required, and identified G3BP1 as a partner through which it suppresses stress granules.\",\n      \"evidence\": \"Active-site mutagenesis restoring catalytic residues, MS-based partner identification, and stress granule imaging in transfected cells\",\n      \"pmids\": [\"20180778\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define which STYXL1 domain mediates G3BP1 binding\", \"Mechanism linking G3BP1 binding to stress granule suppression undefined\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Placed STYXL1 as a positive regulator of mitochondrial apoptosis, with its requirement mapped to the outer membrane permeabilization step.\",\n      \"evidence\": \"RNAi knockdown with cytochrome c release, MOMP, and apoptosis readouts plus subcellular fractionation\",\n      \"pmids\": [\"21262771\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular target at the mitochondrion not yet identified\", \"How a pseudophosphatase controls MOMP unresolved\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Refined the stress granule mechanism by showing suppression is independent of G3BP1 Ser149 phosphorylation and depends on STYXL1's pseudo-catalytic state.\",\n      \"evidence\": \"Phosphomimetic/non-phosphorylatable G3BP1 mutants and STYXL1 active-site mutants with stress granule imaging\",\n      \"pmids\": [\"23163895\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The relevant phosphosite STYXL1 affects on G3BP1 not pinpointed here\", \"Single-lab evidence\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Identified PTPMT1 as the mitochondrial effector through which STYXL1 controls apoptosis, completing the MOMP mechanism.\",\n      \"evidence\": \"Unbiased proteomics, co-IP, PTPMT1 activity assay, and RNAi epistasis with cytochrome c readout\",\n      \"pmids\": [\"24709986\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How catalytically dead STYXL1 suppresses PTPMT1 activity mechanistically unknown\", \"Structural basis of the interaction undetermined\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Connected STYXL1 to cytoskeletal control in neurons, showing it promotes neurite outgrowth via RhoA suppression rather than MAPK signaling.\",\n      \"evidence\": \"Overexpression/knockdown in PC12 cells, MEK inhibitor epistasis, RhoA activity assay, phospho-cofilin immunoblot\",\n      \"pmids\": [\"25479605\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct link between STYXL1 and RhoA regulation not established\", \"Biphasic cofilin phosphorylation effect unexplained\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Characterized STYXL1-induced outgrowths as bona fide neuritic structures forming synapses, extending the neuronal differentiation role to primary neurons.\",\n      \"evidence\": \"Immunofluorescence, TEM synapse confirmation, neurite marker immunoblots, primary hippocampal neuron culture\",\n      \"pmids\": [\"29250526\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular pathway driving synapse formation not defined\", \"In vivo neuronal relevance untested\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Linked STYXL1 to tubulin post-translational modification and HDAC6 regulation, suggesting a route to cytoskeletal control.\",\n      \"evidence\": \"Overexpression in HEK293, immunofluorescence localization, PTM immunoblots\",\n      \"pmids\": [\"30909412\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether STYXL1 acts directly on HDAC6 unknown\", \"Functional consequence of altered tubulin modification untested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Mapped opposing stress granule functions to STYXL1's DUSP versus CH2 domains and identified G3BP1 tyrosine phosphorylation as the relevant modification.\",\n      \"evidence\": \"Reciprocal domain-mapping co-IP with endogenous and overexpressed G3BP1, stress granule imaging, tyrosine phosphorylation immunoblot\",\n      \"pmids\": [\"37516290\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The tyrosine kinase/phosphatase STYXL1 modulates not identified\", \"Single-lab evidence\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Revealed a role for STYXL1 in lysosomal trafficking and ER stress, linking its depletion to enhanced β-glucocerebrosidase activity.\",\n      \"evidence\": \"RNAi knockdown, lysosomal enzyme assay, transcription factor localization imaging, 4-PBA epistasis, ER-lysosome contact imaging\",\n      \"pmids\": [\"37198709\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct molecular substrate or partner in this pathway unknown\", \"Relationship to STYXL1's mitochondrial/stress granule roles unclear\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined an in vivo physiological role: STYXL1 is a germ-cell CCT chaperonin co-factor required for tubulin folding and sperm flagellar assembly.\",\n      \"evidence\": \"Mouse knockout with male infertility phenotype, sperm proteomics, co-IP domain mapping to N-terminal rhodanese-like domain, tubulin polymerization assay\",\n      \"pmids\": [\"38168070\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether STYXL1 modulates CCT activity in non-germ tissues unknown\", \"How rhodanese-like domain engagement of CCT promotes folding mechanistically undefined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how a single catalytically dead pseudophosphatase integrates its distinct roles in stress granules, mitochondrial apoptosis, cytoskeletal/neuronal remodeling, lysosomal trafficking, and CCT-dependent tubulin folding.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unifying structural or regulatory model across the reported functions\", \"Tissue- and context-specificity of each function not delineated\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 3, 7]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 9]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [6, 9]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [1, 3]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [0, 7]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [1, 3]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 7]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [4, 5, 9]}\n    ],\n    \"complexes\": [\"CCT/TRiC chaperonin\"],\n    \"partners\": [\"G3BP1\", \"PTPMT1\", \"HDAC6\", \"CCT1\", \"CCT6\", \"CCT7\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}