{"gene":"MYLK","run_date":"2026-06-10T05:19:52","timeline":{"discoveries":[{"year":1991,"finding":"Telokin is the independently expressed C-terminal domain of smooth muscle MLCK, transcribed from a promoter located within an intron of the MLCK gene (in the 3' region encoding the calmodulin-binding domain), producing a protein identical to the last 155 residues of smMLCK.","method":"cDNA/genomic cloning, promoter mapping, Northern blot","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct molecular cloning with genomic structure determination, independently replicated across multiple labs","pmids":["1748667"],"is_preprint":false},{"year":1992,"finding":"The three-dimensional structure of telokin (C-terminal domain of smMLCK) at 2.8 Å resolution reveals seven strands of antiparallel beta-pleated sheet forming an immunoglobulin-like beta-barrel fold, structurally homologous to immunoglobulin constant domain CH2.","method":"X-ray crystallography","journal":"Journal of molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure solved to 2.8 Å with R-factor 19.5%","pmids":["1404391"],"is_preprint":false},{"year":1996,"finding":"Telokin protein and mRNA are expressed exclusively in smooth muscle tissues and cells (not in non-muscle cells), driven by a smooth muscle cell-specific promoter located within an intron of the smMLCK gene; transgenic mice confirmed this tissue specificity in vivo.","method":"Western blot, Northern blot, in vitro transfection reporter assays, transgenic mice with SV40 T-antigen reporter","journal":"The American journal of physiology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods including in vivo transgenic validation, replicated across studies","pmids":["8764148"],"is_preprint":false},{"year":1997,"finding":"Serum response factor (SRF) binding to a CArG element proximal to the TATA sequence is required for high levels of telokin transcription in A10 smooth muscle cells; MEF2 binds the AT-rich region but is not required for activity.","method":"Luciferase reporter gene assays, gel mobility shift assays, deletion/mutation analysis","journal":"The American journal of physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reporter assays plus gel shifts in single lab, two orthogonal methods","pmids":["9142867"],"is_preprint":false},{"year":1997,"finding":"Telokin modulates the oligomeric state of smMLCK: it solubilizes large MLCK oligomers, shifts the distribution toward dimers, and reduces MLCK binding to filamentous myosin, thereby inhibiting phosphorylation of regulatory light chain in filamentous myosin substrates but not isolated light chain.","method":"In vitro phosphorylation assays, sedimentation, biochemical fractionation","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro reconstitution with multiple assays, single lab","pmids":["9078244"],"is_preprint":false},{"year":1997,"finding":"Telokin is phosphorylated by smMLCK itself (at a threonine residue, up to 0.25 mol/mol stoichiometry) in a Ca2+/calmodulin-dependent manner; this phosphorylation modulates kinase oligomer interactions with telokin and affects MLCK activity.","method":"In vitro kinase assay, enzyme kinetics","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro reconstituted kinase assay, single lab, single study","pmids":["9371697"],"is_preprint":false},{"year":1998,"finding":"Telokin induces Ca2+ desensitization in smooth muscle by enhancing myosin light chain phosphatase (MLCP) activity, accelerating MLC dephosphorylation and relaxation; phosphorylation of telokin by cAMP-dependent protein kinase (PKA) and cGMP-dependent protein kinase (PKG) potentiates this relaxant effect.","method":"Permeabilized smooth muscle preparations, recombinant protein, phosphorylation assays, force measurements","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution with recombinant protein in permeabilized muscle, multiple kinase substrates tested, replicated across labs","pmids":["9556631"],"is_preprint":false},{"year":1998,"finding":"A 310-bp promoter fragment (-163 to +147) is sufficient for smooth muscle cell-specific telokin expression in vivo; a distal estrogen response element (ERE) between -1447 and -1474 is additionally required for expression in uterine smooth muscle.","method":"Transgenic mice with reporter gene, luciferase assays, estrogen stimulation","journal":"The American journal of physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo transgenic confirmation plus in vitro reporter, single lab","pmids":["9612204"],"is_preprint":false},{"year":1999,"finding":"A single human MYLK gene produces both smooth muscle and nonmuscle MLCK isoforms through alternative splicing of a single pre-mRNA precursor, yielding at least five nonmuscle isoforms (MLCK1–4); MLCK2 is the dominant nonmuscle splice variant expressed in human tissues.","method":"cDNA cloning, genomic analysis, RT-PCR, Northern blot","journal":"Genomics","confidence":"High","confidence_rationale":"Tier 2 / Strong — comprehensive genomic and cDNA analysis with Northern blot and quantitative RT-PCR, foundational gene structure paper","pmids":["10198165"],"is_preprint":false},{"year":2000,"finding":"Telokin is phosphorylated in vivo at Ser-13 (the primary site) upon forskolin stimulation in rabbit ileum; portal vein telokin is phosphorylated at both Ser-13 and Ser-19 in vivo. Ser-13 mutation (to Ala) abolishes in vitro phosphorylation by cyclic nucleotide-dependent kinases.","method":"In vivo 32P-labeling with forskolin/GTPγS stimulation, phosphopeptide mapping, site-directed mutagenesis, in vitro kinase assays","journal":"FEBS letters","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vivo site identification combined with mutagenesis confirmation, multiple smooth muscle preparations","pmids":["10981712"],"is_preprint":false},{"year":2001,"finding":"Phosphorylation of telokin at Ser-13 (mimicked by S13D mutant) enhances its Ca2+-desensitizing activity in smooth muscle; the S13D mutant is more effective than wild-type, and its effect is not further enhanced by 8-Br-cGMP confirming Ser-13 specificity. The S19A mutant is more effective than wild-type, while S19D is not different. The C-terminal acidic tail (residues 142–155) is not required for relaxant activity.","method":"Site-directed mutagenesis, permeabilized smooth muscle force measurements, recombinant protein","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution with multiple phosphomimetic/non-phosphorylatable mutants in permeabilized smooth muscle","pmids":["11346659"],"is_preprint":false},{"year":2001,"finding":"Telokin mRNA expression is restricted to smooth muscle cell layers in adult and embryonic tissues; expression is first detected in mouse gut at embryonic day 11.5 and is absent from cardiac or skeletal muscle; telokin protein expression increases postnatally in male and female reproductive tract smooth muscle.","method":"In situ mRNA hybridization, Western blot, postnatal developmental analysis","journal":"American journal of physiology. Cell physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization by in situ hybridization with developmental time course, single lab","pmids":["11121372"],"is_preprint":false},{"year":2002,"finding":"Telokin translocates from cytosol to near the plasma membrane in living smooth muscle cells in response to cGMP signaling (sodium nitroprusside or 8-Br-cGMP); this translocation requires phosphorylation at Ser-13 and/or Ser-19 and is dependent on both PKG and MAPK activity.","method":"GFP-tagged telokin live imaging in primary cultured smooth muscle cells, pharmacological inhibitors, phosphorylation site mutants","journal":"American journal of physiology. Cell physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — live cell imaging with functional mutants, single lab","pmids":["12176732"],"is_preprint":false},{"year":2004,"finding":"The MLCK1 splice isoform (long MLCK) localizes specifically to the perijunctional actomyosin ring in well-differentiated intestinal absorptive enterocytes; siRNA knockdown of MLCK1 specifically reduces tight junction permeability during Na+-glucose cotransport-dependent regulation.","method":"Immunolocalization, siRNA knockdown, transepithelial resistance measurement, crypt-villus expression analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — specific subcellular localization linked to functional consequence via siRNA knockdown with defined permeability readout","pmids":["15507455"],"is_preprint":false},{"year":2004,"finding":"Thyrotroph embryonic factor (TEF) binds to the AT-rich region of the telokin promoter and specifically activates telokin gene transcription in a calcium-dependent manner in smooth muscle cells; TEF does not activate SM22α, smooth muscle α-actin, or smooth muscle myosin heavy chain promoters.","method":"Southwestern screen, gel mobility shift assay, luciferase reporter assays, adenoviral overexpression in A10 cells","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical binding combined with functional reporter assays and endogenous gene expression, single lab","pmids":["14702338"],"is_preprint":false},{"year":2005,"finding":"Telokin mediates Ca2+ desensitization by activating myosin light chain phosphatase (MLCP) in both phasic (ileum) and tonic (femoral artery) smooth muscle; the phospho-telokin mimetic S13D is more effective than wild-type; the extent of relaxation correlates with relative telokin content (27 μM in ileum vs 6 μM in femoral artery).","method":"Alpha-toxin permeabilized smooth muscle, caged-ATP photolysis, recombinant telokin (S13D mutant), force measurements","journal":"Journal of muscle research and cell motility","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — reconstitution with recombinant protein in permeabilized muscle from two tissue types, single lab","pmids":["15750850"],"is_preprint":false},{"year":2006,"finding":"Telokin null mice show ~30% decreased MLCP activity in ileal smooth muscle, increased Ca2+ sensitivity of force (leftward shift of Ca2+-force relationship), increased MLC phosphorylation, and ~50% attenuation of 8-Br-cGMP-induced Ca2+ desensitization; these effects are rescued by recombinant telokin. No difference was observed in aorta (low telokin content).","method":"Telokin knockout mice (congenic background), myosin light chain phosphatase activity assay, Ca2+-force relationship, permeabilized muscle force measurements, recombinant protein rescue","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic knockout with defined biochemical and functional phenotypes plus recombinant protein rescue in a rigorously controlled in vivo model","pmids":["16461919"],"is_preprint":false},{"year":2006,"finding":"The 130-kDa smMLCK isoform is transcribed from an internal CArG-dependent promoter within the mouse mylk gene; SRF binds to a CArG box at -167 to -151 both in vitro and in vivo (by ChIP); SRF knockdown reduces endogenous smMLCK expression; GATA-6 binding to a consensus GATA site inhibits smMLCK promoter activity and blocks myocardin-induced expression.","method":"Luciferase reporter assays, gel mobility shift assay, ChIP, siRNA knockdown, promoter deletion analysis","journal":"American journal of physiology. Cell physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP plus functional reporters plus siRNA, single lab","pmids":["16407417"],"is_preprint":false},{"year":2006,"finding":"The mylk1 gene encodes at least four protein products (two 220-kDa MLCK isoforms, 130-kDa MLCK, and telokin) from four independent promoters; these kinase products regulate nonmuscle and smooth muscle myosin II activity, controlling contractile processes including smooth muscle contraction, cell adhesion, migration, and proliferation.","method":"Review synthesizing promoter and expression studies","journal":"American journal of physiology. Cell physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — synthesis of multiple experimental studies, not a single primary experiment but summarizes established findings","pmids":["16774989"],"is_preprint":false},{"year":2012,"finding":"Phospho-telokin (S13D) activates myosin light chain phosphatase by directly interacting with and binding to inhibited phospho-MYPT1 (regulatory subunit of MLCP), facilitating its binding to phosphomyosin and accelerating MLC20 dephosphorylation; this mechanism does not involve changes in MYPT1 phosphorylation status at Thr-696/Thr-853 or Rho kinase activity.","method":"GST-MYPT1 pulldown, surface plasmon resonance, protein ligation assay, telokin-null mouse smooth muscle reconstitution, phosphatase activity assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — surface plasmon resonance, co-precipitation, and protein ligation assay combined with functional reconstitution in KO muscle, multiple orthogonal methods","pmids":["22544752"],"is_preprint":false},{"year":2015,"finding":"MYLK (MLCK) is identified as a novel target of miR-200c regulated by the ZEB1/miR-200 feedback loop; depletion of MYLK in breast cancer cells reduces invasion and invadopodia formation, identifying MYLK as a player in invadopodia formation downstream of ZEB1/EMT signaling.","method":"In silico screening, miR-200c target validation, siRNA knockdown, invasion/invadopodia assays","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — functional knockdown with invasion phenotype, but mechanistic placement relies partly on computational prediction","pmids":["26334100"],"is_preprint":false},{"year":2015,"finding":"Conditional oocyte-specific knockout of Mylk1 in mice causes subfertility with delayed morula-to-blastocyst transition (>1/3 embryos at morula at 3.5 dpc), while oocyte meiotic maturation, spindle organization, polarity establishment, and polar body extrusion are unaffected.","method":"Cre-loxP conditional knockout, embryo developmental staging in vivo and in vitro","journal":"Biology of reproduction","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional genetic knockout with defined developmental phenotype, single lab","pmids":["25761595"],"is_preprint":false},{"year":2015,"finding":"NMR spectroscopy and molecular modeling of the nmMLCK N-terminal domain show that disease-associated MYLK SNPs (P21H, S147P, V261A) localize to loops connecting immunoglobulin-like domains and adversely affect a 14-3-3 protein-protein interaction motif; 14-3-3 binding was confirmed by immunoprecipitation.","method":"1H-15N HSQC NMR spectroscopy, molecular modeling, immunoprecipitation/Western blot","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — NMR structural analysis with biochemical validation by co-IP, single lab","pmids":["26111161"],"is_preprint":false},{"year":2017,"finding":"Mechanical stress (18% cyclic stretch) increases nmMLCK2 splice variant expression in human lung endothelial cells; MYLK SNPs rs77323602 and rs147245669 alter exon 11 donor/acceptor sites to regulate alternative splicing, with rs147245669 favoring nmMLCK2 production; the splicing factor hnRNPA1 directly regulates MYLK exon 11 splicing.","method":"Minigene constructs, RT-PCR, cyclic stretch model, lymphoblastoid cell line validation, gel shift assay","journal":"American journal of respiratory cell and molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — minigene splicing assay with in silico and cellular validation, single lab","pmids":["27529643","29077485"],"is_preprint":false},{"year":2018,"finding":"Disease-associated MYLK coding SNPs (Pro21His, Ser147Pro, Val261Ala) reduce S1P-induced phosphorylation at Y464 (key regulatory site for nmMLCK enzymatic activation), attenuate nmMLCK translocation to the cell periphery, and retard S1P-induced lamellipodial protrusion in human lung endothelial cells.","method":"Cell immunofluorescence assays, kymographic assays, MYLK transgene expression with SNP variants","journal":"Pulmonary circulation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional cellular assays with defined SNP transgenes, multiple readouts, single lab","pmids":["29480069"],"is_preprint":false},{"year":2018,"finding":"MYLK missense variants associated with familial thoracic aortic disease decrease MLCK kinase activity; loss-of-function MYLK variants cause heritable thoracic aortic dissection. Functional kinase assays demonstrated that at least one missense variant reduces kinase activity, though functional assays alone fail to identify all pathogenic variants.","method":"Kinase activity assays on purified variant proteins, clinical cohort analysis","journal":"Genetics in medicine","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay for variant proteins, but limited to a subset of variants and single lab","pmids":["29925964"],"is_preprint":false},{"year":2018,"finding":"The MYLK missense variant Ala1491Ser reduces kinase activity compared to wild-type protein and causes thoracic aortic aneurysm/dissection with a genotype-phenotype correlation: homozygous carriers have severe early-onset fatal disease while heterozygous carriers have mild late-onset disease.","method":"Kinase activity assay on recombinant variant protein, family clinical analysis","journal":"Orphanet journal of rare diseases","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro kinase activity measurement, single lab","pmids":["29544503"],"is_preprint":false},{"year":2019,"finding":"A unique domain within MLCK1 (not present in other isoforms) directs its recruitment to the perijunctional actomyosin ring (PAMR); a small molecule 'divertin' blocks this domain-mediated MLCK1 recruitment without inhibiting enzymatic activity, thereby preventing TNF-induced MLC phosphorylation and barrier loss in vitro and in vivo.","method":"Domain deletion/mutagenesis, small molecule screen, live cell imaging, MLC phosphorylation assays, mouse barrier/diarrhea models, experimental IBD models","journal":"Nature medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — isoform-specific domain identified functionally, validated with specific small molecule inhibitor in multiple in vitro and in vivo models","pmids":["30936544"],"is_preprint":false},{"year":2019,"finding":"LPS increases intestinal epithelial tight junction permeability via TLR4/MyD88 activation of TAK-1, which activates NF-κB (p50/p65), leading to upregulation of MLCK (MYLK) expression and subsequent increased TJ permeability; siRNA silencing of TAK-1 or p65/p50 inhibited LPS-induced MLCK upregulation.","method":"siRNA knockdown of TAK-1, p65, p50, MLCK in Caco-2 cells and mouse models, TER/permeability assays","journal":"The American journal of pathology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis by sequential siRNA knockdown defining pathway, with functional permeability readout","pmids":["30711488"],"is_preprint":false},{"year":2018,"finding":"MYLK promotes hepatocellular carcinoma cell migration and invasion by phosphorylating myosin light chain (MLC), organizing F-actin stress fibers and cytoskeletal architecture, and facilitating epithelial-mesenchymal transition (increased Vimentin, N-cadherin, Snail; decreased E-cadherin).","method":"shRNA knockdown, wound-healing assay, Matrigel invasion, Western blot for phospho-MLC, F-actin staining","journal":"Clinical and experimental medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with defined cytoskeletal and phosphorylation readouts, single lab","pmids":["29855744"],"is_preprint":false},{"year":2022,"finding":"SIK2 directly phosphorylates MYLK at Ser343, activating MYLK's downstream effector MYL2 (myosin light chain 2) phosphorylation, thereby promoting ovarian cancer cell motility and metastasis; adipocytes induce SIK2 phosphorylation at Ser358 and MYLK phosphorylation at Ser343 to enhance cancer cell motility.","method":"In vitro kinase assay, phospho-site identification, in vitro and in vivo migration/metastasis assays","journal":"Molecular oncology","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay identifying phosphorylation site, with functional in vitro and in vivo validation, single lab","pmids":["35278271"],"is_preprint":false},{"year":2022,"finding":"The m6A methyltransferase METTL3 promotes stability of MYLK mRNA through m6A modification, thereby promoting trophoblast cell invasion; MYLK overexpression rescues the impaired invasion caused by METTL3 depletion.","method":"m6A sequencing, shRNA knockdown, MYLK overexpression rescue, xCELLigence invasion assay","journal":"Placenta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — m6A-seq with rescue experiment, defines post-transcriptional regulation mechanism, single lab","pmids":["36170767"],"is_preprint":false},{"year":2024,"finding":"miR-1204 directly targets MYLK mRNA, reducing MYLK expression in vascular smooth muscle cells (VSMCs), leading to acquisition of a senescence-associated secretory phenotype (SASP) and loss of contractile phenotype; MYLK overexpression reverses miR-1204-induced VSMC senescence, SASP, and contractile phenotypic changes and restores TGF-β signaling.","method":"miRNA target validation, MYLK overexpression rescue, mouse AAD models, angiotensin II and BAPN-induced models","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct miRNA targeting with rescue experiments in multiple mouse models, single lab","pmids":["39013850"],"is_preprint":false},{"year":2024,"finding":"IGF2BP3 (m6A reader) binds MYLK mRNA in an m6A-dependent manner to extend its half-life and increase MYLK protein levels; elevated MYLK inhibits ERK1/2 phosphorylation, thereby repressing MSC adipogenesis.","method":"m6A-RIP, mRNA stability assay, siRNA/shRNA knockdown, ERK1/2 phosphorylation assay, adipogenesis assay, AAV overexpression in mice","journal":"Cellular and molecular life sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — m6A-dependent RNA binding with mRNA stability measurement, functional rescue in vitro and in vivo, single lab","pmids":["38196046"],"is_preprint":false},{"year":2018,"finding":"In C. elegans, MLCK-1 (ortholog of MYLK) relocalizes from apical cell boundaries to basal actomyosin bundles during spermathecae contraction, stabilizing myosin downstream of calcium signaling; MLCK and ROCK act in distinct cell subsets to coordinate the timing of contraction.","method":"Live imaging, GFP-tagging, genetic analysis in C. elegans","journal":"Molecular biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct live-cell localization linked to functional consequence in a well-validated invertebrate ortholog system","pmids":["30088798"],"is_preprint":false}],"current_model":"MYLK encodes multiple isoforms of myosin light chain kinase (including nonmuscle nmMLCK1/2, smooth muscle smMLCK of 130 and 220 kDa, and the independently transcribed C-terminal fragment telokin) from a single gene via alternative promoters and splicing; the kinase isoforms phosphorylate the myosin II regulatory light chain in a Ca2+/calmodulin-dependent manner to control smooth muscle contraction, epithelial tight junction permeability, and cell motility, while telokin acts as a non-catalytic modulator that activates myosin light chain phosphatase (by directly binding phospho-MYPT1) to promote smooth muscle relaxation—an effect potentiated by PKA/PKG phosphorylation of telokin at Ser-13—and the subcellular localization of MLCK1 to the perijunctional actomyosin ring, directed by a unique isoform-specific domain, is essential for its barrier-regulatory function."},"narrative":{"mechanistic_narrative":"MYLK is a single gene that produces multiple functionally distinct products—smooth muscle and nonmuscle myosin light chain kinase isoforms (220-kDa and 130-kDa smMLCK, nonmuscle MLCK1–4) and the non-catalytic C-terminal fragment telokin—through alternative promoters and splicing, thereby controlling actomyosin contractility across smooth muscle contraction, epithelial barrier function, and cell motility [PMID:10198165, PMID:16774989]. The kinase isoforms phosphorylate the myosin II regulatory light chain to organize F-actin and drive contraction, migration, and epithelial-mesenchymal transition [PMID:29855744], and their enzymatic activation depends on regulatory phosphorylation (e.g., S1P-induced Y464) that, when impaired by coding variants, attenuates peripheral translocation and lamellipodial protrusion [PMID:29480069]. Telokin is transcribed from an intronic, smooth-muscle-specific promoter as a protein identical to the smMLCK C-terminus, folds as an immunoglobulin-like beta-barrel, and acts as a relaxant modulator that induces Ca2+ desensitization by activating myosin light chain phosphatase [PMID:1748667, PMID:1404391, PMID:9556631]; mechanistically, phospho-telokin (Ser-13) binds inhibited phospho-MYPT1 directly, facilitating phosphatase engagement of phosphomyosin and accelerating MLC dephosphorylation, with PKA/PKG phosphorylation at Ser-13 potentiating this effect [PMID:11346659, PMID:22544752]. Telokin-null mice confirm this physiological role through reduced phosphatase activity and increased Ca2+ sensitivity that is rescued by recombinant telokin [PMID:16461919]. The MLCK1 splice isoform is uniquely targeted to the perijunctional actomyosin ring by an isoform-specific domain, and this recruitment—blockable by the small molecule divertin without inhibiting catalysis—is required for cytokine-induced tight junction permeability loss [PMID:15507455, PMID:30936544]. MYLK expression is induced via TLR4/MyD88/TAK-1/NF-κB signaling and tuned post-transcriptionally by m6A machinery and miRNAs [PMID:30711488, PMID:36170767, PMID:38196046, PMID:39013850]. Loss-of-function MYLK missense variants that reduce kinase activity cause heritable thoracic aortic aneurysm and dissection [PMID:29925964, PMID:29544503].","teleology":[{"year":1991,"claim":"Established that telokin is not a separate gene but the independently transcribed C-terminal domain of smMLCK, defining the multi-product architecture of the locus.","evidence":"cDNA/genomic cloning and promoter mapping with Northern blot","pmids":["1748667"],"confidence":"High","gaps":["Did not define telokin's biochemical function","Did not enumerate the full set of gene products"]},{"year":1992,"claim":"Resolved telokin's fold as an immunoglobulin-like beta-barrel, providing the structural basis for its protein-interaction modulatory role.","evidence":"X-ray crystallography at 2.8 Å","pmids":["1404391"],"confidence":"High","gaps":["Structure alone did not assign binding partners or function"]},{"year":1996,"claim":"Showed telokin expression is smooth-muscle-specific and driven by an intronic promoter, distinguishing it from the kinase isoforms.","evidence":"Western/Northern blot, reporter assays, transgenic mice","pmids":["8764148"],"confidence":"High","gaps":["Did not identify the transcription factors driving specificity"]},{"year":1997,"claim":"Identified telokin's biochemical activity as a modulator of smMLCK oligomeric state and a substrate of smMLCK itself, linking telokin to kinase regulation.","evidence":"In vitro phosphorylation, sedimentation, kinase assays","pmids":["9078244","9371697"],"confidence":"Medium","gaps":["In vitro reconstitution did not establish physiological relevance","Threonine phosphorylation site not mapped to residue"]},{"year":1998,"claim":"Defined telokin's core physiological function as inducing Ca2+ desensitization by enhancing myosin light chain phosphatase activity, potentiated by PKA/PKG phosphorylation.","evidence":"Permeabilized smooth muscle with recombinant protein and force measurements","pmids":["9556631"],"confidence":"High","gaps":["Molecular target of telokin within the phosphatase complex unknown","Phosphorylation site not yet identified"]},{"year":2000,"claim":"Mapped Ser-13 as the primary in vivo cyclic-nucleotide-dependent phosphorylation site, providing the molecular handle for telokin's relaxant regulation.","evidence":"In vivo 32P-labeling, phosphopeptide mapping, site-directed mutagenesis","pmids":["10981712"],"confidence":"High","gaps":["Functional consequence of Ser-13 phosphorylation not yet demonstrated"]},{"year":2001,"claim":"Demonstrated that Ser-13 phosphorylation (mimicked by S13D) enhances telokin's Ca2+-desensitizing activity, linking the phosphosite to function.","evidence":"Phosphomimetic mutants in permeabilized smooth muscle force assays","pmids":["11346659"],"confidence":"High","gaps":["Did not identify the phosphatase subunit telokin engages"]},{"year":1999,"claim":"Established that smooth muscle and nonmuscle MLCK isoforms arise from a single MYLK gene by alternative splicing, defining the isoform repertoire.","evidence":"cDNA cloning, genomic analysis, RT-PCR, Northern blot","pmids":["10198165"],"confidence":"High","gaps":["Functional distinctions among nonmuscle isoforms not resolved"]},{"year":2006,"claim":"Genetic knockout proved telokin's in vivo role in maintaining phosphatase activity and Ca2+ desensitization, with recombinant rescue confirming causality.","evidence":"Telokin knockout mice, phosphatase and Ca2+-force assays, recombinant rescue","pmids":["16461919"],"confidence":"High","gaps":["Direct phosphatase-binding mechanism still unresolved at this stage"]},{"year":2004,"claim":"Localized the MLCK1 long splice isoform to the perijunctional actomyosin ring and linked it functionally to tight junction permeability regulation.","evidence":"Immunolocalization, siRNA knockdown, transepithelial resistance","pmids":["15507455"],"confidence":"High","gaps":["Targeting determinant for PAMR recruitment not yet identified"]},{"year":2012,"claim":"Resolved the molecular mechanism of telokin action: phospho-telokin directly binds inhibited phospho-MYPT1 to facilitate phosphatase engagement of phosphomyosin.","evidence":"GST pulldown, surface plasmon resonance, protein ligation assay, KO muscle reconstitution","pmids":["22544752"],"confidence":"High","gaps":["Structural basis of the telokin–MYPT1 interaction not solved"]},{"year":2019,"claim":"Identified an isoform-unique domain that recruits MLCK1 to the perijunctional actomyosin ring and showed this recruitment, not catalysis, mediates cytokine-induced barrier loss—targetable by divertin.","evidence":"Domain mutagenesis, small molecule screen, live imaging, mouse barrier/IBD models","pmids":["30936544"],"confidence":"High","gaps":["Binding partner of the recruitment domain at the PAMR not defined"]},{"year":2018,"claim":"Established MYLK as a heritable thoracic aortic disease gene through loss-of-function missense variants that reduce kinase activity.","evidence":"Kinase assays on purified variant proteins and clinical cohort/family analysis","pmids":["29925964","29544503"],"confidence":"Medium","gaps":["Functional assays fail to classify all pathogenic variants","Vascular mechanism downstream of reduced kinase activity not detailed"]},{"year":2022,"claim":"Showed MYLK is itself a kinase substrate, phosphorylated by SIK2 at Ser343 to drive MYL2 phosphorylation and cancer cell motility, expanding upstream regulation.","evidence":"In vitro kinase assay, phosphosite identification, migration/metastasis assays","pmids":["35278271"],"confidence":"Medium","gaps":["Single lab","Generality of Ser343 regulation across isoforms unknown"]},{"year":2024,"claim":"Defined post-transcriptional control of MYLK by m6A machinery and miRNAs governing vascular smooth muscle phenotype and stem-cell differentiation.","evidence":"m6A-RIP, mRNA stability, miRNA target validation, rescue in mouse models","pmids":["38196046","39013850","36170767"],"confidence":"Medium","gaps":["Whether these regulators act on specific MYLK isoforms not resolved","Single-lab studies per regulator"]},{"year":null,"claim":"How distinct MYLK isoforms are differentially deployed and regulated across smooth muscle, endothelium, epithelium, and cancer to produce tissue-specific contractile and barrier outcomes remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified structural model of isoform-specific targeting domains","Endogenous binding partners of the MLCK1 recruitment domain unknown","Isoform-resolved contributions to disease phenotypes not dissected"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[29,25,26,30]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[5,29,30]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[4,6,19]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[4,29]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[12]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[12,24]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[13,27,29]}],"pathway":[{"term_id":"R-HSA-397014","term_label":"Muscle contraction","supporting_discovery_ids":[6,16,18]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[6,19,30]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[25,26,27]}],"complexes":[],"partners":["MYPT1","CALMODULIN","14-3-3","SIK2","HNRNPA1","IGF2BP3","METTL3"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q15746","full_name":"Myosin light chain kinase, smooth muscle","aliases":["Kinase-related protein","KRP","Telokin"],"length_aa":1914,"mass_kda":210.7,"function":"Calcium/calmodulin-dependent myosin light chain kinase implicated in smooth muscle contraction via phosphorylation of myosin light chains (MLC). 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Mediates Giardia spp.-mediated reduced epithelial barrier function during giardiasis intestinal infection via reorganization of cytoskeletal F-actin and tight junctional ZO-1. Necessary for hypotonicity-induced Ca(2+) entry and subsequent activation of volume-sensitive organic osmolyte/anion channels (VSOAC) in cervical cancer cells. 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the independently expressed C-terminal domain of smooth muscle MLCK, transcribed from a promoter located within an intron of the MLCK gene (in the 3' region encoding the calmodulin-binding domain), producing a protein identical to the last 155 residues of smMLCK.\",\n      \"method\": \"cDNA/genomic cloning, promoter mapping, Northern blot\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct molecular cloning with genomic structure determination, independently replicated across multiple labs\",\n      \"pmids\": [\"1748667\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1992,\n      \"finding\": \"The three-dimensional structure of telokin (C-terminal domain of smMLCK) at 2.8 Å resolution reveals seven strands of antiparallel beta-pleated sheet forming an immunoglobulin-like beta-barrel fold, structurally homologous to immunoglobulin constant domain CH2.\",\n      \"method\": \"X-ray crystallography\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure solved to 2.8 Å with R-factor 19.5%\",\n      \"pmids\": [\"1404391\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1996,\n      \"finding\": \"Telokin protein and mRNA are expressed exclusively in smooth muscle tissues and cells (not in non-muscle cells), driven by a smooth muscle cell-specific promoter located within an intron of the smMLCK gene; transgenic mice confirmed this tissue specificity in vivo.\",\n      \"method\": \"Western blot, Northern blot, in vitro transfection reporter assays, transgenic mice with SV40 T-antigen reporter\",\n      \"journal\": \"The American journal of physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods including in vivo transgenic validation, replicated across studies\",\n      \"pmids\": [\"8764148\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"Serum response factor (SRF) binding to a CArG element proximal to the TATA sequence is required for high levels of telokin transcription in A10 smooth muscle cells; MEF2 binds the AT-rich region but is not required for activity.\",\n      \"method\": \"Luciferase reporter gene assays, gel mobility shift assays, deletion/mutation analysis\",\n      \"journal\": \"The American journal of physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reporter assays plus gel shifts in single lab, two orthogonal methods\",\n      \"pmids\": [\"9142867\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"Telokin modulates the oligomeric state of smMLCK: it solubilizes large MLCK oligomers, shifts the distribution toward dimers, and reduces MLCK binding to filamentous myosin, thereby inhibiting phosphorylation of regulatory light chain in filamentous myosin substrates but not isolated light chain.\",\n      \"method\": \"In vitro phosphorylation assays, sedimentation, biochemical fractionation\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro reconstitution with multiple assays, single lab\",\n      \"pmids\": [\"9078244\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"Telokin is phosphorylated by smMLCK itself (at a threonine residue, up to 0.25 mol/mol stoichiometry) in a Ca2+/calmodulin-dependent manner; this phosphorylation modulates kinase oligomer interactions with telokin and affects MLCK activity.\",\n      \"method\": \"In vitro kinase assay, enzyme kinetics\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro reconstituted kinase assay, single lab, single study\",\n      \"pmids\": [\"9371697\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Telokin induces Ca2+ desensitization in smooth muscle by enhancing myosin light chain phosphatase (MLCP) activity, accelerating MLC dephosphorylation and relaxation; phosphorylation of telokin by cAMP-dependent protein kinase (PKA) and cGMP-dependent protein kinase (PKG) potentiates this relaxant effect.\",\n      \"method\": \"Permeabilized smooth muscle preparations, recombinant protein, phosphorylation assays, force measurements\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution with recombinant protein in permeabilized muscle, multiple kinase substrates tested, replicated across labs\",\n      \"pmids\": [\"9556631\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"A 310-bp promoter fragment (-163 to +147) is sufficient for smooth muscle cell-specific telokin expression in vivo; a distal estrogen response element (ERE) between -1447 and -1474 is additionally required for expression in uterine smooth muscle.\",\n      \"method\": \"Transgenic mice with reporter gene, luciferase assays, estrogen stimulation\",\n      \"journal\": \"The American journal of physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo transgenic confirmation plus in vitro reporter, single lab\",\n      \"pmids\": [\"9612204\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"A single human MYLK gene produces both smooth muscle and nonmuscle MLCK isoforms through alternative splicing of a single pre-mRNA precursor, yielding at least five nonmuscle isoforms (MLCK1–4); MLCK2 is the dominant nonmuscle splice variant expressed in human tissues.\",\n      \"method\": \"cDNA cloning, genomic analysis, RT-PCR, Northern blot\",\n      \"journal\": \"Genomics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — comprehensive genomic and cDNA analysis with Northern blot and quantitative RT-PCR, foundational gene structure paper\",\n      \"pmids\": [\"10198165\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Telokin is phosphorylated in vivo at Ser-13 (the primary site) upon forskolin stimulation in rabbit ileum; portal vein telokin is phosphorylated at both Ser-13 and Ser-19 in vivo. Ser-13 mutation (to Ala) abolishes in vitro phosphorylation by cyclic nucleotide-dependent kinases.\",\n      \"method\": \"In vivo 32P-labeling with forskolin/GTPγS stimulation, phosphopeptide mapping, site-directed mutagenesis, in vitro kinase assays\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vivo site identification combined with mutagenesis confirmation, multiple smooth muscle preparations\",\n      \"pmids\": [\"10981712\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Phosphorylation of telokin at Ser-13 (mimicked by S13D mutant) enhances its Ca2+-desensitizing activity in smooth muscle; the S13D mutant is more effective than wild-type, and its effect is not further enhanced by 8-Br-cGMP confirming Ser-13 specificity. The S19A mutant is more effective than wild-type, while S19D is not different. The C-terminal acidic tail (residues 142–155) is not required for relaxant activity.\",\n      \"method\": \"Site-directed mutagenesis, permeabilized smooth muscle force measurements, recombinant protein\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution with multiple phosphomimetic/non-phosphorylatable mutants in permeabilized smooth muscle\",\n      \"pmids\": [\"11346659\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Telokin mRNA expression is restricted to smooth muscle cell layers in adult and embryonic tissues; expression is first detected in mouse gut at embryonic day 11.5 and is absent from cardiac or skeletal muscle; telokin protein expression increases postnatally in male and female reproductive tract smooth muscle.\",\n      \"method\": \"In situ mRNA hybridization, Western blot, postnatal developmental analysis\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization by in situ hybridization with developmental time course, single lab\",\n      \"pmids\": [\"11121372\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Telokin translocates from cytosol to near the plasma membrane in living smooth muscle cells in response to cGMP signaling (sodium nitroprusside or 8-Br-cGMP); this translocation requires phosphorylation at Ser-13 and/or Ser-19 and is dependent on both PKG and MAPK activity.\",\n      \"method\": \"GFP-tagged telokin live imaging in primary cultured smooth muscle cells, pharmacological inhibitors, phosphorylation site mutants\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — live cell imaging with functional mutants, single lab\",\n      \"pmids\": [\"12176732\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"The MLCK1 splice isoform (long MLCK) localizes specifically to the perijunctional actomyosin ring in well-differentiated intestinal absorptive enterocytes; siRNA knockdown of MLCK1 specifically reduces tight junction permeability during Na+-glucose cotransport-dependent regulation.\",\n      \"method\": \"Immunolocalization, siRNA knockdown, transepithelial resistance measurement, crypt-villus expression analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — specific subcellular localization linked to functional consequence via siRNA knockdown with defined permeability readout\",\n      \"pmids\": [\"15507455\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Thyrotroph embryonic factor (TEF) binds to the AT-rich region of the telokin promoter and specifically activates telokin gene transcription in a calcium-dependent manner in smooth muscle cells; TEF does not activate SM22α, smooth muscle α-actin, or smooth muscle myosin heavy chain promoters.\",\n      \"method\": \"Southwestern screen, gel mobility shift assay, luciferase reporter assays, adenoviral overexpression in A10 cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical binding combined with functional reporter assays and endogenous gene expression, single lab\",\n      \"pmids\": [\"14702338\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Telokin mediates Ca2+ desensitization by activating myosin light chain phosphatase (MLCP) in both phasic (ileum) and tonic (femoral artery) smooth muscle; the phospho-telokin mimetic S13D is more effective than wild-type; the extent of relaxation correlates with relative telokin content (27 μM in ileum vs 6 μM in femoral artery).\",\n      \"method\": \"Alpha-toxin permeabilized smooth muscle, caged-ATP photolysis, recombinant telokin (S13D mutant), force measurements\",\n      \"journal\": \"Journal of muscle research and cell motility\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstitution with recombinant protein in permeabilized muscle from two tissue types, single lab\",\n      \"pmids\": [\"15750850\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Telokin null mice show ~30% decreased MLCP activity in ileal smooth muscle, increased Ca2+ sensitivity of force (leftward shift of Ca2+-force relationship), increased MLC phosphorylation, and ~50% attenuation of 8-Br-cGMP-induced Ca2+ desensitization; these effects are rescued by recombinant telokin. No difference was observed in aorta (low telokin content).\",\n      \"method\": \"Telokin knockout mice (congenic background), myosin light chain phosphatase activity assay, Ca2+-force relationship, permeabilized muscle force measurements, recombinant protein rescue\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic knockout with defined biochemical and functional phenotypes plus recombinant protein rescue in a rigorously controlled in vivo model\",\n      \"pmids\": [\"16461919\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"The 130-kDa smMLCK isoform is transcribed from an internal CArG-dependent promoter within the mouse mylk gene; SRF binds to a CArG box at -167 to -151 both in vitro and in vivo (by ChIP); SRF knockdown reduces endogenous smMLCK expression; GATA-6 binding to a consensus GATA site inhibits smMLCK promoter activity and blocks myocardin-induced expression.\",\n      \"method\": \"Luciferase reporter assays, gel mobility shift assay, ChIP, siRNA knockdown, promoter deletion analysis\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP plus functional reporters plus siRNA, single lab\",\n      \"pmids\": [\"16407417\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"The mylk1 gene encodes at least four protein products (two 220-kDa MLCK isoforms, 130-kDa MLCK, and telokin) from four independent promoters; these kinase products regulate nonmuscle and smooth muscle myosin II activity, controlling contractile processes including smooth muscle contraction, cell adhesion, migration, and proliferation.\",\n      \"method\": \"Review synthesizing promoter and expression studies\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — synthesis of multiple experimental studies, not a single primary experiment but summarizes established findings\",\n      \"pmids\": [\"16774989\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Phospho-telokin (S13D) activates myosin light chain phosphatase by directly interacting with and binding to inhibited phospho-MYPT1 (regulatory subunit of MLCP), facilitating its binding to phosphomyosin and accelerating MLC20 dephosphorylation; this mechanism does not involve changes in MYPT1 phosphorylation status at Thr-696/Thr-853 or Rho kinase activity.\",\n      \"method\": \"GST-MYPT1 pulldown, surface plasmon resonance, protein ligation assay, telokin-null mouse smooth muscle reconstitution, phosphatase activity assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — surface plasmon resonance, co-precipitation, and protein ligation assay combined with functional reconstitution in KO muscle, multiple orthogonal methods\",\n      \"pmids\": [\"22544752\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"MYLK (MLCK) is identified as a novel target of miR-200c regulated by the ZEB1/miR-200 feedback loop; depletion of MYLK in breast cancer cells reduces invasion and invadopodia formation, identifying MYLK as a player in invadopodia formation downstream of ZEB1/EMT signaling.\",\n      \"method\": \"In silico screening, miR-200c target validation, siRNA knockdown, invasion/invadopodia assays\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — functional knockdown with invasion phenotype, but mechanistic placement relies partly on computational prediction\",\n      \"pmids\": [\"26334100\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Conditional oocyte-specific knockout of Mylk1 in mice causes subfertility with delayed morula-to-blastocyst transition (>1/3 embryos at morula at 3.5 dpc), while oocyte meiotic maturation, spindle organization, polarity establishment, and polar body extrusion are unaffected.\",\n      \"method\": \"Cre-loxP conditional knockout, embryo developmental staging in vivo and in vitro\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional genetic knockout with defined developmental phenotype, single lab\",\n      \"pmids\": [\"25761595\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"NMR spectroscopy and molecular modeling of the nmMLCK N-terminal domain show that disease-associated MYLK SNPs (P21H, S147P, V261A) localize to loops connecting immunoglobulin-like domains and adversely affect a 14-3-3 protein-protein interaction motif; 14-3-3 binding was confirmed by immunoprecipitation.\",\n      \"method\": \"1H-15N HSQC NMR spectroscopy, molecular modeling, immunoprecipitation/Western blot\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — NMR structural analysis with biochemical validation by co-IP, single lab\",\n      \"pmids\": [\"26111161\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Mechanical stress (18% cyclic stretch) increases nmMLCK2 splice variant expression in human lung endothelial cells; MYLK SNPs rs77323602 and rs147245669 alter exon 11 donor/acceptor sites to regulate alternative splicing, with rs147245669 favoring nmMLCK2 production; the splicing factor hnRNPA1 directly regulates MYLK exon 11 splicing.\",\n      \"method\": \"Minigene constructs, RT-PCR, cyclic stretch model, lymphoblastoid cell line validation, gel shift assay\",\n      \"journal\": \"American journal of respiratory cell and molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — minigene splicing assay with in silico and cellular validation, single lab\",\n      \"pmids\": [\"27529643\", \"29077485\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Disease-associated MYLK coding SNPs (Pro21His, Ser147Pro, Val261Ala) reduce S1P-induced phosphorylation at Y464 (key regulatory site for nmMLCK enzymatic activation), attenuate nmMLCK translocation to the cell periphery, and retard S1P-induced lamellipodial protrusion in human lung endothelial cells.\",\n      \"method\": \"Cell immunofluorescence assays, kymographic assays, MYLK transgene expression with SNP variants\",\n      \"journal\": \"Pulmonary circulation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional cellular assays with defined SNP transgenes, multiple readouts, single lab\",\n      \"pmids\": [\"29480069\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"MYLK missense variants associated with familial thoracic aortic disease decrease MLCK kinase activity; loss-of-function MYLK variants cause heritable thoracic aortic dissection. Functional kinase assays demonstrated that at least one missense variant reduces kinase activity, though functional assays alone fail to identify all pathogenic variants.\",\n      \"method\": \"Kinase activity assays on purified variant proteins, clinical cohort analysis\",\n      \"journal\": \"Genetics in medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay for variant proteins, but limited to a subset of variants and single lab\",\n      \"pmids\": [\"29925964\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"The MYLK missense variant Ala1491Ser reduces kinase activity compared to wild-type protein and causes thoracic aortic aneurysm/dissection with a genotype-phenotype correlation: homozygous carriers have severe early-onset fatal disease while heterozygous carriers have mild late-onset disease.\",\n      \"method\": \"Kinase activity assay on recombinant variant protein, family clinical analysis\",\n      \"journal\": \"Orphanet journal of rare diseases\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase activity measurement, single lab\",\n      \"pmids\": [\"29544503\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"A unique domain within MLCK1 (not present in other isoforms) directs its recruitment to the perijunctional actomyosin ring (PAMR); a small molecule 'divertin' blocks this domain-mediated MLCK1 recruitment without inhibiting enzymatic activity, thereby preventing TNF-induced MLC phosphorylation and barrier loss in vitro and in vivo.\",\n      \"method\": \"Domain deletion/mutagenesis, small molecule screen, live cell imaging, MLC phosphorylation assays, mouse barrier/diarrhea models, experimental IBD models\",\n      \"journal\": \"Nature medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — isoform-specific domain identified functionally, validated with specific small molecule inhibitor in multiple in vitro and in vivo models\",\n      \"pmids\": [\"30936544\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"LPS increases intestinal epithelial tight junction permeability via TLR4/MyD88 activation of TAK-1, which activates NF-κB (p50/p65), leading to upregulation of MLCK (MYLK) expression and subsequent increased TJ permeability; siRNA silencing of TAK-1 or p65/p50 inhibited LPS-induced MLCK upregulation.\",\n      \"method\": \"siRNA knockdown of TAK-1, p65, p50, MLCK in Caco-2 cells and mouse models, TER/permeability assays\",\n      \"journal\": \"The American journal of pathology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis by sequential siRNA knockdown defining pathway, with functional permeability readout\",\n      \"pmids\": [\"30711488\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"MYLK promotes hepatocellular carcinoma cell migration and invasion by phosphorylating myosin light chain (MLC), organizing F-actin stress fibers and cytoskeletal architecture, and facilitating epithelial-mesenchymal transition (increased Vimentin, N-cadherin, Snail; decreased E-cadherin).\",\n      \"method\": \"shRNA knockdown, wound-healing assay, Matrigel invasion, Western blot for phospho-MLC, F-actin staining\",\n      \"journal\": \"Clinical and experimental medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with defined cytoskeletal and phosphorylation readouts, single lab\",\n      \"pmids\": [\"29855744\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"SIK2 directly phosphorylates MYLK at Ser343, activating MYLK's downstream effector MYL2 (myosin light chain 2) phosphorylation, thereby promoting ovarian cancer cell motility and metastasis; adipocytes induce SIK2 phosphorylation at Ser358 and MYLK phosphorylation at Ser343 to enhance cancer cell motility.\",\n      \"method\": \"In vitro kinase assay, phospho-site identification, in vitro and in vivo migration/metastasis assays\",\n      \"journal\": \"Molecular oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay identifying phosphorylation site, with functional in vitro and in vivo validation, single lab\",\n      \"pmids\": [\"35278271\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"The m6A methyltransferase METTL3 promotes stability of MYLK mRNA through m6A modification, thereby promoting trophoblast cell invasion; MYLK overexpression rescues the impaired invasion caused by METTL3 depletion.\",\n      \"method\": \"m6A sequencing, shRNA knockdown, MYLK overexpression rescue, xCELLigence invasion assay\",\n      \"journal\": \"Placenta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — m6A-seq with rescue experiment, defines post-transcriptional regulation mechanism, single lab\",\n      \"pmids\": [\"36170767\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"miR-1204 directly targets MYLK mRNA, reducing MYLK expression in vascular smooth muscle cells (VSMCs), leading to acquisition of a senescence-associated secretory phenotype (SASP) and loss of contractile phenotype; MYLK overexpression reverses miR-1204-induced VSMC senescence, SASP, and contractile phenotypic changes and restores TGF-β signaling.\",\n      \"method\": \"miRNA target validation, MYLK overexpression rescue, mouse AAD models, angiotensin II and BAPN-induced models\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct miRNA targeting with rescue experiments in multiple mouse models, single lab\",\n      \"pmids\": [\"39013850\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"IGF2BP3 (m6A reader) binds MYLK mRNA in an m6A-dependent manner to extend its half-life and increase MYLK protein levels; elevated MYLK inhibits ERK1/2 phosphorylation, thereby repressing MSC adipogenesis.\",\n      \"method\": \"m6A-RIP, mRNA stability assay, siRNA/shRNA knockdown, ERK1/2 phosphorylation assay, adipogenesis assay, AAV overexpression in mice\",\n      \"journal\": \"Cellular and molecular life sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — m6A-dependent RNA binding with mRNA stability measurement, functional rescue in vitro and in vivo, single lab\",\n      \"pmids\": [\"38196046\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"In C. elegans, MLCK-1 (ortholog of MYLK) relocalizes from apical cell boundaries to basal actomyosin bundles during spermathecae contraction, stabilizing myosin downstream of calcium signaling; MLCK and ROCK act in distinct cell subsets to coordinate the timing of contraction.\",\n      \"method\": \"Live imaging, GFP-tagging, genetic analysis in C. elegans\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct live-cell localization linked to functional consequence in a well-validated invertebrate ortholog system\",\n      \"pmids\": [\"30088798\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MYLK encodes multiple isoforms of myosin light chain kinase (including nonmuscle nmMLCK1/2, smooth muscle smMLCK of 130 and 220 kDa, and the independently transcribed C-terminal fragment telokin) from a single gene via alternative promoters and splicing; the kinase isoforms phosphorylate the myosin II regulatory light chain in a Ca2+/calmodulin-dependent manner to control smooth muscle contraction, epithelial tight junction permeability, and cell motility, while telokin acts as a non-catalytic modulator that activates myosin light chain phosphatase (by directly binding phospho-MYPT1) to promote smooth muscle relaxation—an effect potentiated by PKA/PKG phosphorylation of telokin at Ser-13—and the subcellular localization of MLCK1 to the perijunctional actomyosin ring, directed by a unique isoform-specific domain, is essential for its barrier-regulatory function.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MYLK is a single gene that produces multiple functionally distinct products—smooth muscle and nonmuscle myosin light chain kinase isoforms (220-kDa and 130-kDa smMLCK, nonmuscle MLCK1–4) and the non-catalytic C-terminal fragment telokin—through alternative promoters and splicing, thereby controlling actomyosin contractility across smooth muscle contraction, epithelial barrier function, and cell motility [#8, #18]. The kinase isoforms phosphorylate the myosin II regulatory light chain to organize F-actin and drive contraction, migration, and epithelial-mesenchymal transition [#29], and their enzymatic activation depends on regulatory phosphorylation (e.g., S1P-induced Y464) that, when impaired by coding variants, attenuates peripheral translocation and lamellipodial protrusion [#24]. Telokin is transcribed from an intronic, smooth-muscle-specific promoter as a protein identical to the smMLCK C-terminus, folds as an immunoglobulin-like beta-barrel, and acts as a relaxant modulator that induces Ca2+ desensitization by activating myosin light chain phosphatase [#0, #1, #6]; mechanistically, phospho-telokin (Ser-13) binds inhibited phospho-MYPT1 directly, facilitating phosphatase engagement of phosphomyosin and accelerating MLC dephosphorylation, with PKA/PKG phosphorylation at Ser-13 potentiating this effect [#10, #19]. Telokin-null mice confirm this physiological role through reduced phosphatase activity and increased Ca2+ sensitivity that is rescued by recombinant telokin [#16]. The MLCK1 splice isoform is uniquely targeted to the perijunctional actomyosin ring by an isoform-specific domain, and this recruitment—blockable by the small molecule divertin without inhibiting catalysis—is required for cytokine-induced tight junction permeability loss [#13, #27]. MYLK expression is induced via TLR4/MyD88/TAK-1/NF-κB signaling and tuned post-transcriptionally by m6A machinery and miRNAs [#28, #31, #33, #32]. Loss-of-function MYLK missense variants that reduce kinase activity cause heritable thoracic aortic aneurysm and dissection [#25, #26].\",\n  \"teleology\": [\n    {\n      \"year\": 1991,\n      \"claim\": \"Established that telokin is not a separate gene but the independently transcribed C-terminal domain of smMLCK, defining the multi-product architecture of the locus.\",\n      \"evidence\": \"cDNA/genomic cloning and promoter mapping with Northern blot\",\n      \"pmids\": [\"1748667\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define telokin's biochemical function\", \"Did not enumerate the full set of gene products\"]\n    },\n    {\n      \"year\": 1992,\n      \"claim\": \"Resolved telokin's fold as an immunoglobulin-like beta-barrel, providing the structural basis for its protein-interaction modulatory role.\",\n      \"evidence\": \"X-ray crystallography at 2.8 Å\",\n      \"pmids\": [\"1404391\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure alone did not assign binding partners or function\"]\n    },\n    {\n      \"year\": 1996,\n      \"claim\": \"Showed telokin expression is smooth-muscle-specific and driven by an intronic promoter, distinguishing it from the kinase isoforms.\",\n      \"evidence\": \"Western/Northern blot, reporter assays, transgenic mice\",\n      \"pmids\": [\"8764148\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify the transcription factors driving specificity\"]\n    },\n    {\n      \"year\": 1997,\n      \"claim\": \"Identified telokin's biochemical activity as a modulator of smMLCK oligomeric state and a substrate of smMLCK itself, linking telokin to kinase regulation.\",\n      \"evidence\": \"In vitro phosphorylation, sedimentation, kinase assays\",\n      \"pmids\": [\"9078244\", \"9371697\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"In vitro reconstitution did not establish physiological relevance\", \"Threonine phosphorylation site not mapped to residue\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Defined telokin's core physiological function as inducing Ca2+ desensitization by enhancing myosin light chain phosphatase activity, potentiated by PKA/PKG phosphorylation.\",\n      \"evidence\": \"Permeabilized smooth muscle with recombinant protein and force measurements\",\n      \"pmids\": [\"9556631\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular target of telokin within the phosphatase complex unknown\", \"Phosphorylation site not yet identified\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Mapped Ser-13 as the primary in vivo cyclic-nucleotide-dependent phosphorylation site, providing the molecular handle for telokin's relaxant regulation.\",\n      \"evidence\": \"In vivo 32P-labeling, phosphopeptide mapping, site-directed mutagenesis\",\n      \"pmids\": [\"10981712\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of Ser-13 phosphorylation not yet demonstrated\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Demonstrated that Ser-13 phosphorylation (mimicked by S13D) enhances telokin's Ca2+-desensitizing activity, linking the phosphosite to function.\",\n      \"evidence\": \"Phosphomimetic mutants in permeabilized smooth muscle force assays\",\n      \"pmids\": [\"11346659\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify the phosphatase subunit telokin engages\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Established that smooth muscle and nonmuscle MLCK isoforms arise from a single MYLK gene by alternative splicing, defining the isoform repertoire.\",\n      \"evidence\": \"cDNA cloning, genomic analysis, RT-PCR, Northern blot\",\n      \"pmids\": [\"10198165\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional distinctions among nonmuscle isoforms not resolved\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Genetic knockout proved telokin's in vivo role in maintaining phosphatase activity and Ca2+ desensitization, with recombinant rescue confirming causality.\",\n      \"evidence\": \"Telokin knockout mice, phosphatase and Ca2+-force assays, recombinant rescue\",\n      \"pmids\": [\"16461919\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct phosphatase-binding mechanism still unresolved at this stage\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Localized the MLCK1 long splice isoform to the perijunctional actomyosin ring and linked it functionally to tight junction permeability regulation.\",\n      \"evidence\": \"Immunolocalization, siRNA knockdown, transepithelial resistance\",\n      \"pmids\": [\"15507455\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Targeting determinant for PAMR recruitment not yet identified\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Resolved the molecular mechanism of telokin action: phospho-telokin directly binds inhibited phospho-MYPT1 to facilitate phosphatase engagement of phosphomyosin.\",\n      \"evidence\": \"GST pulldown, surface plasmon resonance, protein ligation assay, KO muscle reconstitution\",\n      \"pmids\": [\"22544752\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of the telokin–MYPT1 interaction not solved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified an isoform-unique domain that recruits MLCK1 to the perijunctional actomyosin ring and showed this recruitment, not catalysis, mediates cytokine-induced barrier loss—targetable by divertin.\",\n      \"evidence\": \"Domain mutagenesis, small molecule screen, live imaging, mouse barrier/IBD models\",\n      \"pmids\": [\"30936544\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Binding partner of the recruitment domain at the PAMR not defined\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Established MYLK as a heritable thoracic aortic disease gene through loss-of-function missense variants that reduce kinase activity.\",\n      \"evidence\": \"Kinase assays on purified variant proteins and clinical cohort/family analysis\",\n      \"pmids\": [\"29925964\", \"29544503\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional assays fail to classify all pathogenic variants\", \"Vascular mechanism downstream of reduced kinase activity not detailed\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Showed MYLK is itself a kinase substrate, phosphorylated by SIK2 at Ser343 to drive MYL2 phosphorylation and cancer cell motility, expanding upstream regulation.\",\n      \"evidence\": \"In vitro kinase assay, phosphosite identification, migration/metastasis assays\",\n      \"pmids\": [\"35278271\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Generality of Ser343 regulation across isoforms unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined post-transcriptional control of MYLK by m6A machinery and miRNAs governing vascular smooth muscle phenotype and stem-cell differentiation.\",\n      \"evidence\": \"m6A-RIP, mRNA stability, miRNA target validation, rescue in mouse models\",\n      \"pmids\": [\"38196046\", \"39013850\", \"36170767\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether these regulators act on specific MYLK isoforms not resolved\", \"Single-lab studies per regulator\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How distinct MYLK isoforms are differentially deployed and regulated across smooth muscle, endothelium, epithelium, and cancer to produce tissue-specific contractile and barrier outcomes remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified structural model of isoform-specific targeting domains\", \"Endogenous binding partners of the MLCK1 recruitment domain unknown\", \"Isoform-resolved contributions to disease phenotypes not dissected\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [29, 25, 26, 30]},\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [5, 29, 30]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [4, 6, 19]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [4, 29]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [12]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [12, 24]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [13, 27, 29]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-397014\", \"supporting_discovery_ids\": [6, 16, 18]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [6, 19, 30]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [25, 26, 27]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"MYPT1\", \"calmodulin\", \"14-3-3\", \"SIK2\", \"hnRNPA1\", \"IGF2BP3\", \"METTL3\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}