{"gene":"CAB39","run_date":"2026-04-28T17:28:52","timeline":{"discoveries":[{"year":2003,"finding":"MO25α/β (CAB39) forms a trimeric complex with LKB1 and STRADα/β that functions as an upstream kinase (AMPKK) phosphorylating AMPK at Thr172; both endogenous and recombinant LKB1-STRADα/β-MO25α/β complexes activate AMPK, and catalytically active LKB1, STRAD, and MO25 are all required for full activity.","method":"Biochemical purification from rat liver, immunoprecipitation, in vitro kinase assay, genetic rescue in HeLa cells and LKB1-knockout fibroblasts","journal":"Journal of biology","confidence":"High","confidence_rationale":"Tier 1–2 — in vitro reconstitution, multiple orthogonal methods, genetic rescue, replicated across endogenous and recombinant systems","pmids":["14511394"],"is_preprint":false},{"year":2004,"finding":"MO25α has two distinct binding surfaces (on opposite faces of the armadillo-repeat scaffold) required for assembly of the MO25α-STRADα-LKB1 complex; MO25α binding markedly increases STRADα affinity for LKB1 and activates LKB1 kinase activity without requiring phosphorylation of the LKB1 T-loop; STRADα binds ATP with high affinity but this ATP-binding is not required for LKB1 activation.","method":"Site-directed mutagenesis of MO25α and LKB1 cancer mutants, co-immunoprecipitation, in vitro kinase assay","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 1–2 — mutagenesis combined with kinase assays and binding studies, multiple orthogonal methods","pmids":["15561763"],"is_preprint":false},{"year":2004,"finding":"Crystal structure of MO25α reveals a helical repeat (Armadillo-like) fold; MO25α binds directly to a conserved Trp-Glu-Phe (WEF) motif at the STRADα C-terminus via a hydrophobic pocket, markedly enhancing STRADα-LKB1 binding and LKB1 catalytic activity.","method":"X-ray crystallography (MO25α–STRADα peptide complex), mutagenesis, in vitro binding and kinase assays","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 — crystal structure with mutagenesis and functional validation","pmids":["14730349"],"is_preprint":false},{"year":2005,"finding":"Long-chain acyl-CoA esters (LCACEs, e.g., palmitoyl-CoA) inhibit phosphorylation of AMPK by the recombinant LKB1/STRAD/MO25 complex in a substrate-specific, AMP-dependent manner, without inhibiting LKB1/STRAD/MO25 activity toward a peptide substrate (LKB1tide), revealing that the MO25 complex has substrate-specific regulatory properties.","method":"In vitro kinase assay with recombinant LKB1/STRAD/MO25, liver-purified AMPKK, and defined metabolites","journal":"American journal of physiology. Endocrinology and metabolism","confidence":"Medium","confidence_rationale":"Tier 1 — in vitro reconstitution with recombinant complex, single lab","pmids":["15644453"],"is_preprint":false},{"year":2009,"finding":"Crystal structure of the core heterotrimeric LKB1-STRADα-MO25α complex reveals that STRADα adopts a closed, active-kinase-like conformation and binds LKB1 as a pseudosubstrate; MO25α stabilizes the active conformation of LKB1 by interacting with the LKB1 activation loop, providing a phosphorylation-independent allosteric activation mechanism.","method":"X-ray crystallography of the heterotrimer, mutagenesis, in vitro kinase assay","journal":"Science (New York, N.Y.)","confidence":"High","confidence_rationale":"Tier 1 — crystal structure of full complex with mutagenesis and functional validation","pmids":["19892943"],"is_preprint":false},{"year":2009,"finding":"MO25α (CAB39) directly interacts with the STE20 kinase MST4, and this interaction translocates MST4 from the Golgi to the subapical membrane compartment upon LKB1 activation; MST4 phosphorylates Ezrin at T567 as an essential step in LKB1/STRAD/MO25-induced brush border formation.","method":"Co-immunoprecipitation, live-cell imaging, kinase assay, MST4 inhibition/loss-of-function","journal":"Developmental cell","confidence":"High","confidence_rationale":"Tier 2 — reciprocal Co-IP, subcellular localization with functional consequence, kinase assay, multiple orthogonal methods","pmids":["19386264"],"is_preprint":false},{"year":2011,"finding":"MO25α/β (CAB39/CAB39L) directly binds and activates SPAK/OSR1 (~100-fold) and MST3/MST4/YSK1 (3–4-fold) STE20 kinases in a manner analogous to STRADα binding; MO25-activated SPAK/OSR1 phosphorylates ion co-transporters NKCC1, NKCC2, and NCC; siRNA knockdown of MO25 in mammalian cells reduces endogenous NKCC1 phosphorylation, rescued by MO25α re-expression.","method":"In vitro kinase assay, siRNA knockdown with phosphorylation rescue, mass spectrometry identification of new phosphorylation sites","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 — in vitro reconstitution, siRNA rescue, MS-based phosphosite identification, multiple kinase substrates tested","pmids":["21423148"],"is_preprint":false},{"year":2011,"finding":"miR-451 directly targets CAB39 3′-UTR (confirmed by luciferase reporter assay), reducing CAB39 protein levels and suppressing downstream PI3K/AKT pathway activity in glioma cells.","method":"Luciferase 3′-UTR reporter assay, Western blot, transfection of miR-451 mimics in glioma cell lines","journal":"International journal of oncology","confidence":"Medium","confidence_rationale":"Tier 2–3 — luciferase reporter confirms direct targeting; downstream pathway assessment by Western blot; single lab","pmids":["22179124"],"is_preprint":false},{"year":2012,"finding":"miR-195 and miR-451 functionally target MO25 (CAB39) in cardiac cells; overexpression of miR-195 in C2C12 cells knocks down MO25 expression and suppresses downstream AMPK signaling (reduced ACC phosphorylation and AMPK activity), phenocopying MO25 siRNA knockdown.","method":"miRNA overexpression, siRNA knockdown, AMPK activity assay, Western blot for ACC phosphorylation","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 — functional targeting validated by parallel siRNA knockdown and activity assay; single lab","pmids":["22844503"],"is_preprint":false},{"year":2013,"finding":"Crystal structure of MST4 kinase domain in complex with MO25 shows that MO25 binding rotates the αC helix of MST4 toward its catalytic core, stabilizing MST4 in an active conformation; MST4 kinase domain forms a homodimer required for trans-autophosphorylation; interface mutations disrupting MST4-MO25 interaction or MST4 homodimerization impair MST4 activation and pro-apoptotic function in HEK293T cells.","method":"X-ray crystallography, mutagenesis of interface residues, in vitro kinase assay, cell-based apoptosis assay","journal":"Structure (London, England : 1993)","confidence":"High","confidence_rationale":"Tier 1 — crystal structure with mutagenesis and functional validation in cells","pmids":["23434407"],"is_preprint":false},{"year":2013,"finding":"Crystal structure of MST3 catalytic domain in complex with MO25β reveals that MO25β stabilizes the MST3 kinase domain in a closed, active conformation (active αC helix and A-loop) via an interaction mode similar to MO25α–STRADα; key interface residues Tyr223 of MO25β and Glu58/Ile71 of MST3, when mutated, prevent MO25β-mediated MST3 activation.","method":"X-ray crystallography, mutagenesis, in vitro kinase assay","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 1 — crystal structure with mutagenesis and in vitro functional validation","pmids":["23296203"],"is_preprint":false},{"year":2014,"finding":"CAB39 (MO25) acts as a binding partner that differentially interacts with WNK4 and SPAK/OSR1, enabling WNK4 to activate NKCC1 in a SPAK/OSR1-independent manner; a PF2-like domain in WNK4 mediates direct interaction with the N-terminal domain of NKCC1, demonstrated by yeast two-hybrid, molecular modeling, and functional cotransporter assays.","method":"Yeast two-hybrid, co-immunoprecipitation, functional NKCC1 activation assay in Xenopus oocytes, molecular modeling","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2–3 — multiple complementary methods but interaction dependency on MO25/CAB39 partially inferred; single lab","pmids":["24811174"],"is_preprint":false},{"year":2014,"finding":"Structural analysis of MO25α complexes with STK25 and MST3 reveals a unified mechanism: MO25 directly activates GCK-family kinases (MST3, MST4, STK25, OSR1, SPAK) by stabilizing an active αC helix and A-loop conformation, whereas LKB1 activation requires an additional layer using MO25 to activate the pseudokinase STRAD, which in turn activates LKB1.","method":"X-ray crystallography (MO25α–STK25 and MO25α–MST3 structures), comparative structural analysis","journal":"Journal of structural biology","confidence":"High","confidence_rationale":"Tier 1 — crystal structures providing mechanistic insight, consistent with prior structural data","pmids":["24746913"],"is_preprint":false},{"year":2018,"finding":"In Drosophila Malpighian tubules, Mo25 (CAB39 ortholog) enhances the kinase activity of Fray (SPAK homolog) in vitro and is required for stimulated (but not basal) transepithelial ion flux; Mo25 cooperates with intracellular chloride to regulate WNK signaling and cotransporter-mediated ion transport.","method":"In vitro kinase assay (Drosophila Mo25 + Fray), transgenic Drosophila knockdown, ion flux measurements, chloride sensor imaging","journal":"Journal of the American Society of Nephrology : JASN","confidence":"Medium","confidence_rationale":"Tier 2 — in vitro kinase assay and in vivo genetic knockdown with functional ion transport readout","pmids":["29602832"],"is_preprint":false},{"year":2018,"finding":"C-terminal serine phosphorylation within the WEWS motif of SPAK and OSR1 (catalyzed by WNK kinases in vitro and in cells) enhances their binding to MO25 (CAB39); mutagenesis of key MO25 residues abrogates MO25-mediated activation of SPAK/OSR1.","method":"In vitro kinase assay, phosphomimetic mutagenesis, cell-based co-immunoprecipitation, MO25 interface mutagenesis","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 — mutagenesis plus in vitro and cell-based assays; single lab","pmids":["30060950"],"is_preprint":false},{"year":2022,"finding":"Intracellular Midkine (MDK) interacts with LKB1 and STRAD to disrupt the LKB1-STRAD-MO25 (CAB39) complex, reducing LKB1 kinase activity and dampening basal and stress-induced AMPK activation.","method":"Co-immunoprecipitation, in vitro kinase assay, 2-DG/glucose starvation stress assays, cancer cell proliferation assays","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2–3 — Co-IP and kinase assay show complex disruption; functional AMPK readout; single lab","pmids":["35487917"],"is_preprint":false},{"year":2024,"finding":"Global/inducible double knockout of Cab39 and Cab39l in mouse distal convoluted tubule abolishes NCC phosphorylation (Gitelman-like phenotype); Cab39 proteins are required to localize SPAK at the apical membrane with NCC — in their absence, phosphorylated SPAK is confined to intracellular puncta, preventing NCC activation.","method":"Conditional and global mouse knockout, Western blot, immunofluorescence, urine/blood electrolyte measurements","journal":"Hypertension (Dallas, Tex. : 1979)","confidence":"High","confidence_rationale":"Tier 2 — clean double KO mouse with defined molecular phenotype (loss of NCC phosphorylation) and subcellular localization data","pmids":["38258567"],"is_preprint":false},{"year":2023,"finding":"CAB39 promotes cisplatin resistance in bladder cancer through the CAB39-LKB1-AMPK-LC3 autophagy pathway; CAB39 knockdown sensitizes cisplatin-resistant cells to cisplatin and disrupts mitophagy-mediated ROS reduction; LKB1 knockdown downstream of CAB39 phenocopies CAB39 knockdown in resistance.","method":"Proteomic analysis of resistant vs. parental cells, gene knockdown (siRNA/shRNA), overexpression, autophagy flux assays, in vivo xenograft","journal":"Free radical biology & medicine","confidence":"Medium","confidence_rationale":"Tier 2–3 — epistasis by sequential knockdown places CAB39 upstream of LKB1-AMPK-LC3; single lab","pmids":["37726090"],"is_preprint":false},{"year":2026,"finding":"In Cab39/Cab39l double-knockout mouse distal convoluted tubule, phosphorylated SPAK accumulates in cytoplasmic puncta (distinct from canonical WNK bodies) independently of KS-WNK1; these condensates require active upstream phosphorylation and contain WNK4 and L-WNK1 but are absent under high-K+ conditions; Cab39 is thus required for SPAK translocation from condensates to the apical membrane to phosphorylate NCC.","method":"Triple knockout mice (Cab39-DKO × KS-WNK1 KO), immunofluorescence, immunoblotting, dietary K+ manipulation","journal":"American journal of physiology. Renal physiology","confidence":"Medium","confidence_rationale":"Tier 2 — clean genetic KO with subcellular localization and phosphorylation readouts; single lab, single study","pmids":["41903110"],"is_preprint":false},{"year":2024,"finding":"STRAD-binding small molecules can activate LKB1 kinase activity in a target-dependent manner in cancer cell lines, demonstrating that pharmacological engagement of the STRAD subunit of the LKB1-STRAD-MO25 complex is sufficient to stimulate the complex.","method":"Small-molecule screen, LKB1 kinase activity assay, cancer cell line target-dependent proliferation assay","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 — preprint, single lab, limited mechanistic detail on MO25 role specifically","pmids":["bio_10.1101_2024.12.17.628051"],"is_preprint":true}],"current_model":"CAB39 (MO25) is an armadillo-repeat scaffold protein that allosterically activates LKB1 by stabilizing the trimeric LKB1-STRADα-MO25 complex (with STRADα acting as a pseudosubstrate and MO25 contacting the LKB1 activation loop), and independently activates multiple STE20-family kinases (SPAK, OSR1, MST3, MST4, STK25) by rotating their αC helix into an active conformation; through SPAK/OSR1 activation, CAB39 is required for apical-membrane localization of SPAK and consequent phosphorylation of ion co-transporters (NKCC1, NCC), and through the LKB1-AMPK axis it regulates cellular energy sensing, autophagy, and polarity."},"narrative":{"teleology":[{"year":2003,"claim":"Identification of CAB39 (MO25) as an obligate component of the LKB1–STRAD complex that constitutes the long-sought AMPK kinase (AMPKK) established that LKB1 kinase activity toward AMPK requires both STRAD and MO25 cofactors.","evidence":"Biochemical purification from rat liver, reconstitution with recombinant proteins, kinase assays, and genetic rescue in LKB1-null cells","pmids":["14511394"],"confidence":"High","gaps":["How MO25 contacts LKB1 and STRAD at the structural level was unknown","Whether MO25 has activating roles independent of LKB1 was not addressed"]},{"year":2004,"claim":"Structural and mutagenesis studies revealed that MO25α is an armadillo-repeat protein with two distinct binding surfaces: one engages the STRAD C-terminal WEF motif via a hydrophobic pocket and the other faces LKB1, and MO25 binding dramatically increases STRADα–LKB1 affinity without requiring LKB1 T-loop phosphorylation, establishing MO25 as an allosteric activator.","evidence":"X-ray crystallography of MO25α–STRADα peptide complex, site-directed mutagenesis, in vitro kinase and binding assays","pmids":["14730349","15561763"],"confidence":"High","gaps":["Full heterotrimeric structure was not yet resolved","How MO25 engages the LKB1 activation loop remained speculative"]},{"year":2009,"claim":"The crystal structure of the complete LKB1–STRADα–MO25α heterotrimer showed that STRADα acts as a pseudosubstrate locked in an active-kinase-like conformation and that MO25α directly contacts the LKB1 activation loop, providing the structural basis for phosphorylation-independent allosteric activation of LKB1.","evidence":"X-ray crystallography of the full heterotrimer with mutagenesis and kinase assays","pmids":["19892943"],"confidence":"High","gaps":["Regulation of the complex by metabolites or post-translational modifications of MO25 itself was not resolved","In vivo tissue-specific requirements of MO25 were not tested"]},{"year":2009,"claim":"Discovery that MO25 directly binds and relocates the STE20 kinase MST4 from the Golgi to the subapical membrane — where MST4 phosphorylates Ezrin to initiate brush border formation — revealed the first LKB1-independent function of MO25 as an activator of GCK-family kinases in epithelial polarity.","evidence":"Co-immunoprecipitation, live-cell imaging, kinase assays, MST4 loss-of-function in polarizing epithelial cells","pmids":["19386264"],"confidence":"High","gaps":["Whether MO25 activates MST4 catalytically or only controls its localization was ambiguous","Generality to other GCK kinases was not established"]},{"year":2011,"claim":"MO25 was shown to directly and potently activate SPAK/OSR1 (~100-fold) and MST3/MST4/YSK1 (3–4-fold), with SPAK/OSR1 activation driving phosphorylation of ion co-transporters NKCC1/NKCC2/NCC; siRNA knockdown of MO25 reduced endogenous NKCC1 phosphorylation, establishing MO25 as a master activator of multiple STE20 kinases beyond LKB1.","evidence":"In vitro kinase assays with recombinant proteins, siRNA knockdown and rescue, mass spectrometry phosphosite mapping","pmids":["21423148"],"confidence":"High","gaps":["How MO25 structurally engages SPAK/OSR1 versus STRAD was not resolved","In vivo physiological significance for renal ion transport was not tested"]},{"year":2013,"claim":"Crystal structures of MO25 in complex with MST4, MST3, and STK25 revealed a unified activation mechanism: MO25 rotates the αC helix of GCK-family kinases into an active conformation, analogous to but distinct from the two-step mechanism used for LKB1 (where MO25 first activates pseudokinase STRAD, which then activates LKB1).","evidence":"X-ray crystallography of MO25α–MST4, MO25β–MST3, and MO25α–STK25 complexes with mutagenesis and kinase assays","pmids":["23434407","23296203","24746913"],"confidence":"High","gaps":["Whether αC-helix rotation is sufficient or also requires dimerization-dependent trans-autophosphorylation in vivo was not fully resolved","No structural data for MO25–SPAK/OSR1 complexes"]},{"year":2018,"claim":"WNK-catalyzed phosphorylation of the C-terminal WEWS motif of SPAK/OSR1 was found to enhance their binding to MO25, providing a mechanism by which upstream WNK signaling primes SPAK/OSR1 for MO25-mediated activation; Drosophila studies confirmed that Mo25 is required for stimulated (but not basal) transepithelial ion flux via the WNK–SPAK axis.","evidence":"Phosphomimetic mutagenesis, in vitro kinase assays, co-immunoprecipitation, Drosophila Mo25 knockdown with ion flux measurements","pmids":["30060950","29602832"],"confidence":"Medium","gaps":["Relative contributions of WNK phosphorylation versus chloride sensing in regulating MO25–SPAK interaction in vivo were not dissected","No mammalian in vivo validation at this stage"]},{"year":2024,"claim":"Double knockout of Cab39/Cab39l in mouse distal convoluted tubule abolished NCC phosphorylation and produced a Gitelman-like phenotype, directly demonstrating that MO25 proteins are essential for SPAK apical membrane localization and NCC activation in vivo; without MO25, phosphorylated SPAK accumulates in cytoplasmic condensates that contain WNK4 and L-WNK1 but cannot reach the apical membrane.","evidence":"Conditional and global double-knockout mice, immunofluorescence, immunoblotting, electrolyte measurements, triple-KO genetic epistasis with KS-WNK1","pmids":["38258567","41903110"],"confidence":"High","gaps":["Identity and biophysical nature of the SPAK-containing cytoplasmic condensates are not fully characterized","Whether MO25 loss affects other renal transport pathways beyond NCC/NKCC remains open"]},{"year":null,"claim":"Key open questions include: (1) how MO25 selectively partitions between the LKB1–STRAD complex and free GCK kinases in different tissues; (2) whether post-translational modifications of MO25 itself regulate its activity or localization; (3) the structural basis of the MO25–SPAK/OSR1 interaction; and (4) whether therapeutic targeting of the MO25 interface can selectively modulate AMPK versus ion-transport signaling.","evidence":"","pmids":[],"confidence":"Low","gaps":["No structure of MO25–SPAK/OSR1 complex exists","No post-translational modification map for MO25 has been reported","Tissue-specific stoichiometric competition between LKB1 and GCK kinases for MO25 has not been measured"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,1,2,4,6,9,10,12]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,4,6,16]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[0,6,18]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[5,16]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,6,8,13,14,17]},{"term_id":"R-HSA-382551","term_label":"Transport of small molecules","supporting_discovery_ids":[6,13,16,18]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[17]}],"complexes":["LKB1–STRADα–MO25α heterotrimer"],"partners":["LKB1","STRADΑ","SPAK","OSR1","MST3","MST4","STK25","WNK4"],"other_free_text":[]},"mechanistic_narrative":"CAB39 (MO25) is an armadillo-repeat scaffold protein that allosterically activates two major kinase systems: the LKB1–STRADα tumor-suppressor complex and multiple STE20/GCK-family kinases (SPAK, OSR1, MST3, MST4, STK25). In the LKB1 pathway, CAB39 stabilizes the heterotrimeric LKB1–STRADα–MO25 complex by engaging the STRADα C-terminal WEF motif and directly contacting the LKB1 activation loop, thereby activating LKB1 in a phosphorylation-independent manner to drive downstream AMPK signaling, energy sensing, polarity, and autophagy [PMID:14511394, PMID:19892943, PMID:14730349]. CAB39 independently binds and potently activates SPAK/OSR1 (~100-fold) and MST3/MST4/STK25 (3–4-fold) by rotating the αC helix of these kinases into an active conformation; through SPAK/OSR1, CAB39 is required for apical membrane targeting of SPAK and consequent phosphorylation of the ion co-transporters NKCC1 and NCC, and loss of CAB39 in mouse distal convoluted tubule abolishes NCC phosphorylation, producing a Gitelman-like salt-wasting phenotype [PMID:21423148, PMID:23434407, PMID:24746913, PMID:38258567]. CAB39 expression is post-transcriptionally regulated by miR-451 and miR-195, linking its abundance to AMPK pathway output in cardiac and glioma contexts [PMID:22179124, PMID:22844503]."},"prefetch_data":{"uniprot":{"accession":"Q9Y376","full_name":"Calcium-binding protein 39","aliases":["MO25alpha","Protein Mo25"],"length_aa":341,"mass_kda":39.9,"function":"Component of a complex that binds and activates STK11/LKB1. In the complex, required to stabilize the interaction between CAB39/MO25 (CAB39/MO25alpha or CAB39L/MO25beta) and STK11/LKB1","subcellular_location":"Cytoplasm","url":"https://www.uniprot.org/uniprotkb/Q9Y376/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CAB39","classification":"Not Classified","n_dependent_lines":303,"n_total_lines":1208,"dependency_fraction":0.2508278145695364},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"STK11","stoichiometry":4.0}],"url":"https://opencell.sf.czbiohub.org/search/CAB39","total_profiled":1310},"omim":[{"mim_id":"612760","title":"SNF-RELATED KINASE; SNRK","url":"https://www.omim.org/entry/612760"},{"mim_id":"612175","title":"CALCIUM-BINDING PROTEIN 39-LIKE PROTEIN; CAB39L","url":"https://www.omim.org/entry/612175"},{"mim_id":"612174","title":"CALCIUM-BINDING PROTEIN 39; CAB39","url":"https://www.omim.org/entry/612174"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Vesicles","reliability":"Approved"},{"location":"Endoplasmic reticulum","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"skeletal muscle","ntpm":168.0}],"url":"https://www.proteinatlas.org/search/CAB39"},"hgnc":{"alias_symbol":["CGI-66","MO25"],"prev_symbol":[]},"alphafold":{"accession":"Q9Y376","domains":[],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y376","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y376-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y376-F1-predicted_aligned_error_v6.png","plddt_mean":94.69},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CAB39","jax_strain_url":"https://www.jax.org/strain/search?query=CAB39"},"sequence":{"accession":"Q9Y376","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y376.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y376/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y376"}},"corpus_meta":[{"pmid":"14511394","id":"PMC_14511394","title":"Complexes between the LKB1 tumor suppressor, STRAD alpha/beta and MO25 alpha/beta are upstream kinases in the AMP-activated protein kinase cascade.","date":"2003","source":"Journal of biology","url":"https://pubmed.ncbi.nlm.nih.gov/14511394","citation_count":1335,"is_preprint":false},{"pmid":"19892943","id":"PMC_19892943","title":"Structure of the LKB1-STRAD-MO25 complex reveals an allosteric mechanism of kinase activation.","date":"2009","source":"Science (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/19892943","citation_count":284,"is_preprint":false},{"pmid":"19386264","id":"PMC_19386264","title":"Mst4 and Ezrin induce brush borders downstream of the Lkb1/Strad/Mo25 polarization complex.","date":"2009","source":"Developmental cell","url":"https://pubmed.ncbi.nlm.nih.gov/19386264","citation_count":131,"is_preprint":false},{"pmid":"15561763","id":"PMC_15561763","title":"Analysis of the LKB1-STRAD-MO25 complex.","date":"2004","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/15561763","citation_count":129,"is_preprint":false},{"pmid":"22179124","id":"PMC_22179124","title":"MicroRNA miR-451 downregulates the PI3K/AKT pathway through CAB39 in human glioma.","date":"2011","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/22179124","citation_count":127,"is_preprint":false},{"pmid":"21423148","id":"PMC_21423148","title":"MO25 is a master regulator of SPAK/OSR1 and MST3/MST4/YSK1 protein kinases.","date":"2011","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/21423148","citation_count":121,"is_preprint":false},{"pmid":"14730349","id":"PMC_14730349","title":"Crystal structure of MO25 alpha in complex with the C terminus of the pseudo kinase STE20-related adaptor.","date":"2004","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/14730349","citation_count":63,"is_preprint":false},{"pmid":"16096637","id":"PMC_16096637","title":"Fission yeast MO25 protein is localized at SPB and septum and is essential for cell morphogenesis.","date":"2005","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/16096637","citation_count":61,"is_preprint":false},{"pmid":"22844503","id":"PMC_22844503","title":"Micro-RNA-195 and -451 regulate the LKB1/AMPK signaling axis by targeting MO25.","date":"2012","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/22844503","citation_count":60,"is_preprint":false},{"pmid":"30180756","id":"PMC_30180756","title":"MiRNA-451 Inhibits Glioma Cell Proliferation and Invasion Through the mTOR/HIF-1α/VEGF Signaling Pathway by Targeting CAB39.","date":"2018","source":"Human gene therapy. Clinical development","url":"https://pubmed.ncbi.nlm.nih.gov/30180756","citation_count":53,"is_preprint":false},{"pmid":"15644453","id":"PMC_15644453","title":"Long-chain acyl-CoA esters inhibit phosphorylation of AMP-activated protein kinase at threonine-172 by LKB1/STRAD/MO25.","date":"2005","source":"American journal of physiology. Endocrinology and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/15644453","citation_count":52,"is_preprint":false},{"pmid":"23028357","id":"PMC_23028357","title":"The NDR kinase scaffold HYM1/MO25 is essential for MAK2 map kinase signaling in Neurospora crassa.","date":"2012","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/23028357","citation_count":49,"is_preprint":false},{"pmid":"33505470","id":"PMC_33505470","title":"Metformin-Induced MicroRNA-34a-3p Downregulation Alleviates Senescence in Human Dental Pulp Stem Cells by Targeting CAB39 through the AMPK/mTOR Signaling Pathway.","date":"2021","source":"Stem cells international","url":"https://pubmed.ncbi.nlm.nih.gov/33505470","citation_count":45,"is_preprint":false},{"pmid":"18669938","id":"PMC_18669938","title":"Thyroid hormone effects on LKB1, MO25, phospho-AMPK, phospho-CREB, and PGC-1alpha in rat muscle.","date":"2008","source":"Journal of applied physiology (Bethesda, Md. : 1985)","url":"https://pubmed.ncbi.nlm.nih.gov/18669938","citation_count":42,"is_preprint":false},{"pmid":"29602832","id":"PMC_29602832","title":"Intracellular Chloride and Scaffold Protein Mo25 Cooperatively Regulate Transepithelial Ion Transport through WNK Signaling in the Malpighian Tubule.","date":"2018","source":"Journal of the American Society of Nephrology : JASN","url":"https://pubmed.ncbi.nlm.nih.gov/29602832","citation_count":40,"is_preprint":false},{"pmid":"28197410","id":"PMC_28197410","title":"MiR-451 Promotes Cell Proliferation and Metastasis in Pancreatic Cancer through Targeting CAB39.","date":"2017","source":"BioMed research international","url":"https://pubmed.ncbi.nlm.nih.gov/28197410","citation_count":39,"is_preprint":false},{"pmid":"23434407","id":"PMC_23434407","title":"Structure of the MST4 in complex with MO25 provides insights into its activation mechanism.","date":"2013","source":"Structure (London, England : 1993)","url":"https://pubmed.ncbi.nlm.nih.gov/23434407","citation_count":39,"is_preprint":false},{"pmid":"33101593","id":"PMC_33101593","title":"MicroRNA-31-5p Exacerbates Lipopolysaccharide-Induced Acute Lung Injury via Inactivating Cab39/AMPKα Pathway.","date":"2020","source":"Oxidative medicine and cellular longevity","url":"https://pubmed.ncbi.nlm.nih.gov/33101593","citation_count":35,"is_preprint":false},{"pmid":"24811174","id":"PMC_24811174","title":"A novel Ste20-related proline/alanine-rich kinase (SPAK)-independent pathway involving calcium-binding protein 39 (Cab39) and serine threonine kinase with no lysine member 4 (WNK4) in the activation of Na-K-Cl cotransporters.","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/24811174","citation_count":35,"is_preprint":false},{"pmid":"16325501","id":"PMC_16325501","title":"The fission yeast MO25 protein functions in polar growth and cell separation.","date":"2005","source":"European journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/16325501","citation_count":35,"is_preprint":false},{"pmid":"35661494","id":"PMC_35661494","title":"Eucalyptol relieves imidacloprid-induced autophagy through the miR-451/Cab39/AMPK axis in Ctenopharyngodon idellus kidney cells†.","date":"2022","source":"Aquatic toxicology (Amsterdam, Netherlands)","url":"https://pubmed.ncbi.nlm.nih.gov/35661494","citation_count":34,"is_preprint":false},{"pmid":"8672247","id":"PMC_8672247","title":"Molecular characterization of the Drosophila Mo25 gene, which is conserved among Drosophila, mouse, and yeast.","date":"1996","source":"DNA and cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/8672247","citation_count":26,"is_preprint":false},{"pmid":"27798271","id":"PMC_27798271","title":"WNK-Cab39-NKCC1 signaling increases the susceptibility to ischemic brain damage in hypertensive rats.","date":"2016","source":"Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/27798271","citation_count":26,"is_preprint":false},{"pmid":"32527986","id":"PMC_32527986","title":"miR-107 inhibition upregulates CAB39 and activates AMPK-Nrf2 signaling to protect osteoblasts from dexamethasone-induced oxidative injury and cytotoxicity.","date":"2020","source":"Aging","url":"https://pubmed.ncbi.nlm.nih.gov/32527986","citation_count":25,"is_preprint":false},{"pmid":"15292028","id":"PMC_15292028","title":"Endurance training increases LKB1 and MO25 protein but not AMP-activated protein kinase kinase activity in skeletal muscle.","date":"2004","source":"American journal of physiology. Endocrinology and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/15292028","citation_count":25,"is_preprint":false},{"pmid":"35487917","id":"PMC_35487917","title":"Midkine noncanonically suppresses AMPK activation through disrupting the LKB1-STRAD-Mo25 complex.","date":"2022","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/35487917","citation_count":23,"is_preprint":false},{"pmid":"23296203","id":"PMC_23296203","title":"Structural insights into the activation of MST3 by MO25.","date":"2013","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/23296203","citation_count":22,"is_preprint":false},{"pmid":"16278246","id":"PMC_16278246","title":"Evidence against regulation of AMP-activated protein kinase and LKB1/STRAD/MO25 activity by creatine phosphate.","date":"2005","source":"American journal of physiology. Endocrinology and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/16278246","citation_count":21,"is_preprint":false},{"pmid":"32994913","id":"PMC_32994913","title":"HPV16 E6/E7 promote the translocation and glucose uptake of GLUT1 by PI3K/AKT pathway via relieving miR-451 inhibitory effect on CAB39 in lung cancer cells.","date":"2020","source":"Therapeutic advances in chronic disease","url":"https://pubmed.ncbi.nlm.nih.gov/32994913","citation_count":19,"is_preprint":false},{"pmid":"31494170","id":"PMC_31494170","title":"Antagonism of miR-429 ameliorates anoxia/reoxygenation injury in cardiomyocytes by enhancing MO25/LKB1/AMPK mediated autophagy.","date":"2019","source":"Life sciences","url":"https://pubmed.ncbi.nlm.nih.gov/31494170","citation_count":18,"is_preprint":false},{"pmid":"24746913","id":"PMC_24746913","title":"Structural insights into regulatory mechanisms of MO25-mediated kinase activation.","date":"2014","source":"Journal of structural biology","url":"https://pubmed.ncbi.nlm.nih.gov/24746913","citation_count":17,"is_preprint":false},{"pmid":"35190902","id":"PMC_35190902","title":"MicroRNA-22 promoted osteogenic differentiation of valvular interstitial cells by inhibiting CAB39 expression during aortic valve calcification.","date":"2022","source":"Cellular and molecular life sciences : CMLS","url":"https://pubmed.ncbi.nlm.nih.gov/35190902","citation_count":17,"is_preprint":false},{"pmid":"37726090","id":"PMC_37726090","title":"CAB39 promotes cisplatin resistance in bladder cancer via the LKB1-AMPK-LC3 pathway.","date":"2023","source":"Free radical biology & medicine","url":"https://pubmed.ncbi.nlm.nih.gov/37726090","citation_count":16,"is_preprint":false},{"pmid":"18054329","id":"PMC_18054329","title":"The GC kinase Fray and Mo25 regulate Drosophila asymmetric divisions.","date":"2007","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/18054329","citation_count":16,"is_preprint":false},{"pmid":"16985256","id":"PMC_16985256","title":"Effects of 3-phosphoglycerate and other metabolites on the activation of AMP-activated protein kinase by LKB1-STRAD-MO25.","date":"2006","source":"American journal of physiology. Endocrinology and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/16985256","citation_count":16,"is_preprint":false},{"pmid":"33262952","id":"PMC_33262952","title":"Circular RNA circGSK3B Promotes Cell Proliferation, Migration, and Invasion by Sponging miR-1265 and Regulating CAB39 Expression in Hepatocellular Carcinoma.","date":"2020","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/33262952","citation_count":15,"is_preprint":false},{"pmid":"24205201","id":"PMC_24205201","title":"Cell cycle regulated interaction of a yeast Hippo kinase and its activator MO25/Hym1.","date":"2013","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/24205201","citation_count":15,"is_preprint":false},{"pmid":"25809261","id":"PMC_25809261","title":"LKB1/Mo25/STRAD uniquely impacts sarcomeric contractile function and posttranslational modification.","date":"2015","source":"Biophysical journal","url":"https://pubmed.ncbi.nlm.nih.gov/25809261","citation_count":13,"is_preprint":false},{"pmid":"38258567","id":"PMC_38258567","title":"Kinase Scaffold Cab39 Is Necessary for Phospho-Activation of the Thiazide-Sensitive NCC.","date":"2024","source":"Hypertension (Dallas, Tex. : 1979)","url":"https://pubmed.ncbi.nlm.nih.gov/38258567","citation_count":11,"is_preprint":false},{"pmid":"28004876","id":"PMC_28004876","title":"Towards the Development of Small-Molecule MO25 Binders as Potential Indirect SPAK/OSR1 Kinase Inhibitors.","date":"2017","source":"Chembiochem : a European journal of chemical biology","url":"https://pubmed.ncbi.nlm.nih.gov/28004876","citation_count":9,"is_preprint":false},{"pmid":"30060950","id":"PMC_30060950","title":"C-terminal phosphorylation of SPAK and OSR1 kinases promotes their binding and activation by the scaffolding protein MO25.","date":"2018","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/30060950","citation_count":9,"is_preprint":false},{"pmid":"30208334","id":"PMC_30208334","title":"Evolutionary history of Mo25 gene in plants, a component of RAM/MOR signaling network.","date":"2018","source":"Mechanisms of development","url":"https://pubmed.ncbi.nlm.nih.gov/30208334","citation_count":7,"is_preprint":false},{"pmid":"37379707","id":"PMC_37379707","title":"MicroRNA-32-3p facilitates cerebral ischemia/reperfusion injury through inhibiting Cab39/AMPK.","date":"2023","source":"International immunopharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/37379707","citation_count":5,"is_preprint":false},{"pmid":"35794445","id":"PMC_35794445","title":"Long noncoding RNA TRG-AS1 protects against glucocorticoid-induced osteoporosis in a rat model by regulating miR-802-mediated CAB39/AMPK/SIRT-1/NF-κB axis.","date":"2022","source":"Human cell","url":"https://pubmed.ncbi.nlm.nih.gov/35794445","citation_count":5,"is_preprint":false},{"pmid":"36867352","id":"PMC_36867352","title":"MAPKAPK5-AS1/miR-515-5p/CAB39 Axis Contributes to Non-small Cell Lung Cancer Cell Proliferation and Migration.","date":"2023","source":"Molecular biotechnology","url":"https://pubmed.ncbi.nlm.nih.gov/36867352","citation_count":5,"is_preprint":false},{"pmid":"23989145","id":"PMC_23989145","title":"Structure of zebrafish MO25.","date":"2013","source":"Acta crystallographica. Section F, Structural biology and crystallization communications","url":"https://pubmed.ncbi.nlm.nih.gov/23989145","citation_count":4,"is_preprint":false},{"pmid":"38649419","id":"PMC_38649419","title":"miR-451a was selectively sorted into exosomes and promoted the progression of esophageal squamous cell carcinoma through CAB39.","date":"2024","source":"Cancer gene therapy","url":"https://pubmed.ncbi.nlm.nih.gov/38649419","citation_count":4,"is_preprint":false},{"pmid":"33177871","id":"PMC_33177871","title":"CAB39 Promotes the Proliferation of Nasopharyngeal Carcinoma CNE-1 Cells via Up-Regulating p-JNK.","date":"2020","source":"Cancer management and research","url":"https://pubmed.ncbi.nlm.nih.gov/33177871","citation_count":4,"is_preprint":false},{"pmid":"39171884","id":"PMC_39171884","title":"CAB39 modulates epithelial-mesenchymal transition through NF-κB signaling activation, enhancing invasion, and metastasis in bladder cancer.","date":"2024","source":"Environmental toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/39171884","citation_count":2,"is_preprint":false},{"pmid":"40463288","id":"PMC_40463288","title":"Roles of the MO25 protein Pmo25 in contractile-ring stability and localization of the NDR kinase Sid2 during cytokinesis.","date":"2025","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/40463288","citation_count":2,"is_preprint":false},{"pmid":"33855067","id":"PMC_33855067","title":"Retracted: MiR-451 Promotes Cell Proliferation and Metastasis in Pancreatic Cancer through Targeting CAB39.","date":"2021","source":"BioMed research international","url":"https://pubmed.ncbi.nlm.nih.gov/33855067","citation_count":1,"is_preprint":false},{"pmid":"41903110","id":"PMC_41903110","title":"Deletion of Cab39 adaptors results in KS-WNK1 independent accumulation of SPAK in biomolecular condensates.","date":"2026","source":"American journal of physiology. Renal physiology","url":"https://pubmed.ncbi.nlm.nih.gov/41903110","citation_count":0,"is_preprint":false},{"pmid":"41550769","id":"PMC_41550769","title":"The MO25 protein Pmo25 functions in contractile ring stability and Sid2 localization during cytokinesis.","date":"2025","source":"iScience","url":"https://pubmed.ncbi.nlm.nih.gov/41550769","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2024.12.17.628051","title":"Small molecule activation of the tumor suppressor kinase LKB1","date":"2024-12-20","source":"bioRxiv","url":"https://doi.org/10.1101/2024.12.17.628051","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":28113,"output_tokens":5020,"usd":0.07982},"stage2":{"model":"claude-opus-4-6","input_tokens":8753,"output_tokens":2970,"usd":0.177022},"total_usd":0.256842,"stage1_batch_id":"msgbatch_011kfMfE81o6CTcyhVUqumuW","stage2_batch_id":"msgbatch_01DTputS6WNkH745x1nVSoBo","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2003,\n      \"finding\": \"MO25α/β (CAB39) forms a trimeric complex with LKB1 and STRADα/β that functions as an upstream kinase (AMPKK) phosphorylating AMPK at Thr172; both endogenous and recombinant LKB1-STRADα/β-MO25α/β complexes activate AMPK, and catalytically active LKB1, STRAD, and MO25 are all required for full activity.\",\n      \"method\": \"Biochemical purification from rat liver, immunoprecipitation, in vitro kinase assay, genetic rescue in HeLa cells and LKB1-knockout fibroblasts\",\n      \"journal\": \"Journal of biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — in vitro reconstitution, multiple orthogonal methods, genetic rescue, replicated across endogenous and recombinant systems\",\n      \"pmids\": [\"14511394\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"MO25α has two distinct binding surfaces (on opposite faces of the armadillo-repeat scaffold) required for assembly of the MO25α-STRADα-LKB1 complex; MO25α binding markedly increases STRADα affinity for LKB1 and activates LKB1 kinase activity without requiring phosphorylation of the LKB1 T-loop; STRADα binds ATP with high affinity but this ATP-binding is not required for LKB1 activation.\",\n      \"method\": \"Site-directed mutagenesis of MO25α and LKB1 cancer mutants, co-immunoprecipitation, in vitro kinase assay\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — mutagenesis combined with kinase assays and binding studies, multiple orthogonal methods\",\n      \"pmids\": [\"15561763\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Crystal structure of MO25α reveals a helical repeat (Armadillo-like) fold; MO25α binds directly to a conserved Trp-Glu-Phe (WEF) motif at the STRADα C-terminus via a hydrophobic pocket, markedly enhancing STRADα-LKB1 binding and LKB1 catalytic activity.\",\n      \"method\": \"X-ray crystallography (MO25α–STRADα peptide complex), mutagenesis, in vitro binding and kinase assays\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — crystal structure with mutagenesis and functional validation\",\n      \"pmids\": [\"14730349\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Long-chain acyl-CoA esters (LCACEs, e.g., palmitoyl-CoA) inhibit phosphorylation of AMPK by the recombinant LKB1/STRAD/MO25 complex in a substrate-specific, AMP-dependent manner, without inhibiting LKB1/STRAD/MO25 activity toward a peptide substrate (LKB1tide), revealing that the MO25 complex has substrate-specific regulatory properties.\",\n      \"method\": \"In vitro kinase assay with recombinant LKB1/STRAD/MO25, liver-purified AMPKK, and defined metabolites\",\n      \"journal\": \"American journal of physiology. Endocrinology and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 — in vitro reconstitution with recombinant complex, single lab\",\n      \"pmids\": [\"15644453\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Crystal structure of the core heterotrimeric LKB1-STRADα-MO25α complex reveals that STRADα adopts a closed, active-kinase-like conformation and binds LKB1 as a pseudosubstrate; MO25α stabilizes the active conformation of LKB1 by interacting with the LKB1 activation loop, providing a phosphorylation-independent allosteric activation mechanism.\",\n      \"method\": \"X-ray crystallography of the heterotrimer, mutagenesis, in vitro kinase assay\",\n      \"journal\": \"Science (New York, N.Y.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — crystal structure of full complex with mutagenesis and functional validation\",\n      \"pmids\": [\"19892943\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"MO25α (CAB39) directly interacts with the STE20 kinase MST4, and this interaction translocates MST4 from the Golgi to the subapical membrane compartment upon LKB1 activation; MST4 phosphorylates Ezrin at T567 as an essential step in LKB1/STRAD/MO25-induced brush border formation.\",\n      \"method\": \"Co-immunoprecipitation, live-cell imaging, kinase assay, MST4 inhibition/loss-of-function\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal Co-IP, subcellular localization with functional consequence, kinase assay, multiple orthogonal methods\",\n      \"pmids\": [\"19386264\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"MO25α/β (CAB39/CAB39L) directly binds and activates SPAK/OSR1 (~100-fold) and MST3/MST4/YSK1 (3–4-fold) STE20 kinases in a manner analogous to STRADα binding; MO25-activated SPAK/OSR1 phosphorylates ion co-transporters NKCC1, NKCC2, and NCC; siRNA knockdown of MO25 in mammalian cells reduces endogenous NKCC1 phosphorylation, rescued by MO25α re-expression.\",\n      \"method\": \"In vitro kinase assay, siRNA knockdown with phosphorylation rescue, mass spectrometry identification of new phosphorylation sites\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — in vitro reconstitution, siRNA rescue, MS-based phosphosite identification, multiple kinase substrates tested\",\n      \"pmids\": [\"21423148\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"miR-451 directly targets CAB39 3′-UTR (confirmed by luciferase reporter assay), reducing CAB39 protein levels and suppressing downstream PI3K/AKT pathway activity in glioma cells.\",\n      \"method\": \"Luciferase 3′-UTR reporter assay, Western blot, transfection of miR-451 mimics in glioma cell lines\",\n      \"journal\": \"International journal of oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — luciferase reporter confirms direct targeting; downstream pathway assessment by Western blot; single lab\",\n      \"pmids\": [\"22179124\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"miR-195 and miR-451 functionally target MO25 (CAB39) in cardiac cells; overexpression of miR-195 in C2C12 cells knocks down MO25 expression and suppresses downstream AMPK signaling (reduced ACC phosphorylation and AMPK activity), phenocopying MO25 siRNA knockdown.\",\n      \"method\": \"miRNA overexpression, siRNA knockdown, AMPK activity assay, Western blot for ACC phosphorylation\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — functional targeting validated by parallel siRNA knockdown and activity assay; single lab\",\n      \"pmids\": [\"22844503\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Crystal structure of MST4 kinase domain in complex with MO25 shows that MO25 binding rotates the αC helix of MST4 toward its catalytic core, stabilizing MST4 in an active conformation; MST4 kinase domain forms a homodimer required for trans-autophosphorylation; interface mutations disrupting MST4-MO25 interaction or MST4 homodimerization impair MST4 activation and pro-apoptotic function in HEK293T cells.\",\n      \"method\": \"X-ray crystallography, mutagenesis of interface residues, in vitro kinase assay, cell-based apoptosis assay\",\n      \"journal\": \"Structure (London, England : 1993)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — crystal structure with mutagenesis and functional validation in cells\",\n      \"pmids\": [\"23434407\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Crystal structure of MST3 catalytic domain in complex with MO25β reveals that MO25β stabilizes the MST3 kinase domain in a closed, active conformation (active αC helix and A-loop) via an interaction mode similar to MO25α–STRADα; key interface residues Tyr223 of MO25β and Glu58/Ile71 of MST3, when mutated, prevent MO25β-mediated MST3 activation.\",\n      \"method\": \"X-ray crystallography, mutagenesis, in vitro kinase assay\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — crystal structure with mutagenesis and in vitro functional validation\",\n      \"pmids\": [\"23296203\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CAB39 (MO25) acts as a binding partner that differentially interacts with WNK4 and SPAK/OSR1, enabling WNK4 to activate NKCC1 in a SPAK/OSR1-independent manner; a PF2-like domain in WNK4 mediates direct interaction with the N-terminal domain of NKCC1, demonstrated by yeast two-hybrid, molecular modeling, and functional cotransporter assays.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, functional NKCC1 activation assay in Xenopus oocytes, molecular modeling\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — multiple complementary methods but interaction dependency on MO25/CAB39 partially inferred; single lab\",\n      \"pmids\": [\"24811174\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Structural analysis of MO25α complexes with STK25 and MST3 reveals a unified mechanism: MO25 directly activates GCK-family kinases (MST3, MST4, STK25, OSR1, SPAK) by stabilizing an active αC helix and A-loop conformation, whereas LKB1 activation requires an additional layer using MO25 to activate the pseudokinase STRAD, which in turn activates LKB1.\",\n      \"method\": \"X-ray crystallography (MO25α–STK25 and MO25α–MST3 structures), comparative structural analysis\",\n      \"journal\": \"Journal of structural biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — crystal structures providing mechanistic insight, consistent with prior structural data\",\n      \"pmids\": [\"24746913\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"In Drosophila Malpighian tubules, Mo25 (CAB39 ortholog) enhances the kinase activity of Fray (SPAK homolog) in vitro and is required for stimulated (but not basal) transepithelial ion flux; Mo25 cooperates with intracellular chloride to regulate WNK signaling and cotransporter-mediated ion transport.\",\n      \"method\": \"In vitro kinase assay (Drosophila Mo25 + Fray), transgenic Drosophila knockdown, ion flux measurements, chloride sensor imaging\",\n      \"journal\": \"Journal of the American Society of Nephrology : JASN\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vitro kinase assay and in vivo genetic knockdown with functional ion transport readout\",\n      \"pmids\": [\"29602832\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"C-terminal serine phosphorylation within the WEWS motif of SPAK and OSR1 (catalyzed by WNK kinases in vitro and in cells) enhances their binding to MO25 (CAB39); mutagenesis of key MO25 residues abrogates MO25-mediated activation of SPAK/OSR1.\",\n      \"method\": \"In vitro kinase assay, phosphomimetic mutagenesis, cell-based co-immunoprecipitation, MO25 interface mutagenesis\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — mutagenesis plus in vitro and cell-based assays; single lab\",\n      \"pmids\": [\"30060950\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Intracellular Midkine (MDK) interacts with LKB1 and STRAD to disrupt the LKB1-STRAD-MO25 (CAB39) complex, reducing LKB1 kinase activity and dampening basal and stress-induced AMPK activation.\",\n      \"method\": \"Co-immunoprecipitation, in vitro kinase assay, 2-DG/glucose starvation stress assays, cancer cell proliferation assays\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — Co-IP and kinase assay show complex disruption; functional AMPK readout; single lab\",\n      \"pmids\": [\"35487917\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Global/inducible double knockout of Cab39 and Cab39l in mouse distal convoluted tubule abolishes NCC phosphorylation (Gitelman-like phenotype); Cab39 proteins are required to localize SPAK at the apical membrane with NCC — in their absence, phosphorylated SPAK is confined to intracellular puncta, preventing NCC activation.\",\n      \"method\": \"Conditional and global mouse knockout, Western blot, immunofluorescence, urine/blood electrolyte measurements\",\n      \"journal\": \"Hypertension (Dallas, Tex. : 1979)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean double KO mouse with defined molecular phenotype (loss of NCC phosphorylation) and subcellular localization data\",\n      \"pmids\": [\"38258567\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CAB39 promotes cisplatin resistance in bladder cancer through the CAB39-LKB1-AMPK-LC3 autophagy pathway; CAB39 knockdown sensitizes cisplatin-resistant cells to cisplatin and disrupts mitophagy-mediated ROS reduction; LKB1 knockdown downstream of CAB39 phenocopies CAB39 knockdown in resistance.\",\n      \"method\": \"Proteomic analysis of resistant vs. parental cells, gene knockdown (siRNA/shRNA), overexpression, autophagy flux assays, in vivo xenograft\",\n      \"journal\": \"Free radical biology & medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — epistasis by sequential knockdown places CAB39 upstream of LKB1-AMPK-LC3; single lab\",\n      \"pmids\": [\"37726090\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"In Cab39/Cab39l double-knockout mouse distal convoluted tubule, phosphorylated SPAK accumulates in cytoplasmic puncta (distinct from canonical WNK bodies) independently of KS-WNK1; these condensates require active upstream phosphorylation and contain WNK4 and L-WNK1 but are absent under high-K+ conditions; Cab39 is thus required for SPAK translocation from condensates to the apical membrane to phosphorylate NCC.\",\n      \"method\": \"Triple knockout mice (Cab39-DKO × KS-WNK1 KO), immunofluorescence, immunoblotting, dietary K+ manipulation\",\n      \"journal\": \"American journal of physiology. Renal physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — clean genetic KO with subcellular localization and phosphorylation readouts; single lab, single study\",\n      \"pmids\": [\"41903110\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"STRAD-binding small molecules can activate LKB1 kinase activity in a target-dependent manner in cancer cell lines, demonstrating that pharmacological engagement of the STRAD subunit of the LKB1-STRAD-MO25 complex is sufficient to stimulate the complex.\",\n      \"method\": \"Small-molecule screen, LKB1 kinase activity assay, cancer cell line target-dependent proliferation assay\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 — preprint, single lab, limited mechanistic detail on MO25 role specifically\",\n      \"pmids\": [\"bio_10.1101_2024.12.17.628051\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"CAB39 (MO25) is an armadillo-repeat scaffold protein that allosterically activates LKB1 by stabilizing the trimeric LKB1-STRADα-MO25 complex (with STRADα acting as a pseudosubstrate and MO25 contacting the LKB1 activation loop), and independently activates multiple STE20-family kinases (SPAK, OSR1, MST3, MST4, STK25) by rotating their αC helix into an active conformation; through SPAK/OSR1 activation, CAB39 is required for apical-membrane localization of SPAK and consequent phosphorylation of ion co-transporters (NKCC1, NCC), and through the LKB1-AMPK axis it regulates cellular energy sensing, autophagy, and polarity.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"CAB39 (MO25) is an armadillo-repeat scaffold protein that allosterically activates two major kinase systems: the LKB1–STRADα tumor-suppressor complex and multiple STE20/GCK-family kinases (SPAK, OSR1, MST3, MST4, STK25). In the LKB1 pathway, CAB39 stabilizes the heterotrimeric LKB1–STRADα–MO25 complex by engaging the STRADα C-terminal WEF motif and directly contacting the LKB1 activation loop, thereby activating LKB1 in a phosphorylation-independent manner to drive downstream AMPK signaling, energy sensing, polarity, and autophagy [PMID:14511394, PMID:19892943, PMID:14730349]. CAB39 independently binds and potently activates SPAK/OSR1 (~100-fold) and MST3/MST4/STK25 (3–4-fold) by rotating the αC helix of these kinases into an active conformation; through SPAK/OSR1, CAB39 is required for apical membrane targeting of SPAK and consequent phosphorylation of the ion co-transporters NKCC1 and NCC, and loss of CAB39 in mouse distal convoluted tubule abolishes NCC phosphorylation, producing a Gitelman-like salt-wasting phenotype [PMID:21423148, PMID:23434407, PMID:24746913, PMID:38258567]. CAB39 expression is post-transcriptionally regulated by miR-451 and miR-195, linking its abundance to AMPK pathway output in cardiac and glioma contexts [PMID:22179124, PMID:22844503].\",\n  \"teleology\": [\n    {\n      \"year\": 2003,\n      \"claim\": \"Identification of CAB39 (MO25) as an obligate component of the LKB1–STRAD complex that constitutes the long-sought AMPK kinase (AMPKK) established that LKB1 kinase activity toward AMPK requires both STRAD and MO25 cofactors.\",\n      \"evidence\": \"Biochemical purification from rat liver, reconstitution with recombinant proteins, kinase assays, and genetic rescue in LKB1-null cells\",\n      \"pmids\": [\"14511394\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How MO25 contacts LKB1 and STRAD at the structural level was unknown\", \"Whether MO25 has activating roles independent of LKB1 was not addressed\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Structural and mutagenesis studies revealed that MO25α is an armadillo-repeat protein with two distinct binding surfaces: one engages the STRAD C-terminal WEF motif via a hydrophobic pocket and the other faces LKB1, and MO25 binding dramatically increases STRADα–LKB1 affinity without requiring LKB1 T-loop phosphorylation, establishing MO25 as an allosteric activator.\",\n      \"evidence\": \"X-ray crystallography of MO25α–STRADα peptide complex, site-directed mutagenesis, in vitro kinase and binding assays\",\n      \"pmids\": [\"14730349\", \"15561763\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full heterotrimeric structure was not yet resolved\", \"How MO25 engages the LKB1 activation loop remained speculative\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"The crystal structure of the complete LKB1–STRADα–MO25α heterotrimer showed that STRADα acts as a pseudosubstrate locked in an active-kinase-like conformation and that MO25α directly contacts the LKB1 activation loop, providing the structural basis for phosphorylation-independent allosteric activation of LKB1.\",\n      \"evidence\": \"X-ray crystallography of the full heterotrimer with mutagenesis and kinase assays\",\n      \"pmids\": [\"19892943\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Regulation of the complex by metabolites or post-translational modifications of MO25 itself was not resolved\", \"In vivo tissue-specific requirements of MO25 were not tested\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Discovery that MO25 directly binds and relocates the STE20 kinase MST4 from the Golgi to the subapical membrane — where MST4 phosphorylates Ezrin to initiate brush border formation — revealed the first LKB1-independent function of MO25 as an activator of GCK-family kinases in epithelial polarity.\",\n      \"evidence\": \"Co-immunoprecipitation, live-cell imaging, kinase assays, MST4 loss-of-function in polarizing epithelial cells\",\n      \"pmids\": [\"19386264\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether MO25 activates MST4 catalytically or only controls its localization was ambiguous\", \"Generality to other GCK kinases was not established\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"MO25 was shown to directly and potently activate SPAK/OSR1 (~100-fold) and MST3/MST4/YSK1 (3–4-fold), with SPAK/OSR1 activation driving phosphorylation of ion co-transporters NKCC1/NKCC2/NCC; siRNA knockdown of MO25 reduced endogenous NKCC1 phosphorylation, establishing MO25 as a master activator of multiple STE20 kinases beyond LKB1.\",\n      \"evidence\": \"In vitro kinase assays with recombinant proteins, siRNA knockdown and rescue, mass spectrometry phosphosite mapping\",\n      \"pmids\": [\"21423148\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How MO25 structurally engages SPAK/OSR1 versus STRAD was not resolved\", \"In vivo physiological significance for renal ion transport was not tested\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Crystal structures of MO25 in complex with MST4, MST3, and STK25 revealed a unified activation mechanism: MO25 rotates the αC helix of GCK-family kinases into an active conformation, analogous to but distinct from the two-step mechanism used for LKB1 (where MO25 first activates pseudokinase STRAD, which then activates LKB1).\",\n      \"evidence\": \"X-ray crystallography of MO25α–MST4, MO25β–MST3, and MO25α–STK25 complexes with mutagenesis and kinase assays\",\n      \"pmids\": [\"23434407\", \"23296203\", \"24746913\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether αC-helix rotation is sufficient or also requires dimerization-dependent trans-autophosphorylation in vivo was not fully resolved\", \"No structural data for MO25–SPAK/OSR1 complexes\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"WNK-catalyzed phosphorylation of the C-terminal WEWS motif of SPAK/OSR1 was found to enhance their binding to MO25, providing a mechanism by which upstream WNK signaling primes SPAK/OSR1 for MO25-mediated activation; Drosophila studies confirmed that Mo25 is required for stimulated (but not basal) transepithelial ion flux via the WNK–SPAK axis.\",\n      \"evidence\": \"Phosphomimetic mutagenesis, in vitro kinase assays, co-immunoprecipitation, Drosophila Mo25 knockdown with ion flux measurements\",\n      \"pmids\": [\"30060950\", \"29602832\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Relative contributions of WNK phosphorylation versus chloride sensing in regulating MO25–SPAK interaction in vivo were not dissected\", \"No mammalian in vivo validation at this stage\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Double knockout of Cab39/Cab39l in mouse distal convoluted tubule abolished NCC phosphorylation and produced a Gitelman-like phenotype, directly demonstrating that MO25 proteins are essential for SPAK apical membrane localization and NCC activation in vivo; without MO25, phosphorylated SPAK accumulates in cytoplasmic condensates that contain WNK4 and L-WNK1 but cannot reach the apical membrane.\",\n      \"evidence\": \"Conditional and global double-knockout mice, immunofluorescence, immunoblotting, electrolyte measurements, triple-KO genetic epistasis with KS-WNK1\",\n      \"pmids\": [\"38258567\", \"41903110\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity and biophysical nature of the SPAK-containing cytoplasmic condensates are not fully characterized\", \"Whether MO25 loss affects other renal transport pathways beyond NCC/NKCC remains open\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key open questions include: (1) how MO25 selectively partitions between the LKB1–STRAD complex and free GCK kinases in different tissues; (2) whether post-translational modifications of MO25 itself regulate its activity or localization; (3) the structural basis of the MO25–SPAK/OSR1 interaction; and (4) whether therapeutic targeting of the MO25 interface can selectively modulate AMPK versus ion-transport signaling.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structure of MO25–SPAK/OSR1 complex exists\", \"No post-translational modification map for MO25 has been reported\", \"Tissue-specific stoichiometric competition between LKB1 and GCK kinases for MO25 has not been measured\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 1, 2, 4, 6, 9, 10, 12]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 4, 6, 16]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [0, 6, 18]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [5, 16]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 6, 8, 13, 14, 17]},\n      {\"term_id\": \"R-HSA-382551\", \"supporting_discovery_ids\": [6, 13, 16, 18]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [17]}\n    ],\n    \"complexes\": [\n      \"LKB1–STRADα–MO25α heterotrimer\"\n    ],\n    \"partners\": [\n      \"LKB1\",\n      \"STRADα\",\n      \"SPAK\",\n      \"OSR1\",\n      \"MST3\",\n      \"MST4\",\n      \"STK25\",\n      \"WNK4\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}