{"gene":"KLC2","run_date":"2026-06-10T02:59:49","timeline":{"discoveries":[{"year":2002,"finding":"14-3-3 protein directly binds to kinesin heterodimers through interaction with KLC2, and this interaction is phosphorylation-dependent. Mass spectrometry identified Ser575 as the phosphorylation site on KLC2 responsible for the in vivo interaction with 14-3-3.","method":"Proteomic pulldown from PC12 cells expressing myc-tagged 14-3-3eta, SDS-PAGE/mass spectrometry, interaction studies with KLC2 variants in cultured cells","journal":"Biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS-identified phosphorylation site with cell-based interaction studies, single lab, two orthogonal methods (pulldown + MS site mapping)","pmids":["11969417"],"is_preprint":false},{"year":2008,"finding":"Conventional kinesin holoenzymes are composed of kinesin-1 homodimers (not heterodimers), and KLC subunits also homodimerize. No specificity was found between kinesin-1 isoforms and KLC1/KLC2, suggesting six variant forms of kinesin exist. Different variants associate with biochemically distinct membrane-bounded organelles (MBOs), suggesting kinesin-1 heavy chains target the holoenzyme to specific cargoes.","method":"Immunoprecipitation from brain tissue, subcellular fractionation","journal":"Biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP from brain tissue with fractionation, single lab, two orthogonal methods","pmids":["18361505"],"is_preprint":false},{"year":2010,"finding":"GSK-3β phosphorylates KLC2 on serine residues upon AMPA stimulation, causing dissociation of the GluR1/KLC2 protein complex and release of AMPA-containing vesicles from the kinesin cargo system. A peptide inhibitor of KLC2 phosphorylation (TAT-KLCpCDK) reduced long-term depression formation.","method":"Phosphorylation assays, co-immunoprecipitation, peptide inhibitor experiments in neuronal cells, behavioral assays in mice","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP demonstrating complex dissociation upon phosphorylation, peptide inhibitor with cellular and behavioral readouts, single lab","pmids":["20534517"],"is_preprint":false},{"year":2011,"finding":"LMTK2 signals via protein phosphatase-1C (PP1C) to increase inhibitory phosphorylation of GSK-3β on serine-9, which reduces KLC2 phosphorylation by GSK-3β and promotes binding of the cargo Smad2 to KLC2. siRNA knockdown of LMTK2 reduces Smad2 binding to KLC2 and inhibits TGFβ-induced Smad2 nuclear signalling.","method":"siRNA knockdown, co-immunoprecipitation, phosphorylation assays, TGFβ signaling readouts","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and siRNA knockdown with defined signaling readout, single lab, two orthogonal methods","pmids":["21996745"],"is_preprint":false},{"year":2011,"finding":"A bipartite tryptophan-based (W-acidic) motif present in vaccinia protein A36 and in over 450 human proteins mediates binding to KLC1 and KLC2. Different proteins containing this motif show distinct preferences for KLC1 versus KLC2. Regions containing this motif from cellular proteins can functionally recruit KLC and promote kinesin-1-dependent virus transport.","method":"Bioinformatic analysis, functional transport assays using vaccinia as surrogate cargo, co-immunoprecipitation outside infection context","journal":"The EMBO journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus functional transport assays, single lab, two orthogonal methods identifying a shared KLC binding motif","pmids":["21915095"],"is_preprint":false},{"year":2012,"finding":"Crystal structures of the TPR domains of KLC1 and KLC2 were determined by X-ray crystallography. KLC2 residue S328 (corresponding to N343 in KLC1) lacks the ability to form a 'carboxylate clamp' for JIP1 binding, explaining why KLC2, unlike KLC1, does not interact with JIP1. A common groove in both KLC1 and KLC2 TPR domains mediates binding of shared cargoes.","method":"X-ray crystallography, isothermal titration calorimetry","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure determination combined with ITC binding measurements, directly explaining differential cargo specificity between KLC1 and KLC2","pmids":["22470497"],"is_preprint":false},{"year":2009,"finding":"In C. elegans, the KASH protein UNC-83 interacts with kinesin-1 light chain KLC-2 (identified by yeast two-hybrid and confirmed by in vitro assays), recruits KLC-2 to the nuclear envelope in heterologous tissue culture, and acts as a cargo adaptor for kinesin-1-dependent nuclear migration. A synthetic KLC-2::KASH fusion protein could partially bypass the requirement for UNC-83 in nuclear migration.","method":"Yeast two-hybrid, in vitro binding assay, heterologous tissue culture recruitment assay, genetic epistasis with mutant phenotype analysis, synthetic rescue experiment","journal":"Development (Cambridge, England)","confidence":"High","confidence_rationale":"Tier 2 / Strong — yeast two-hybrid confirmed by in vitro assay plus functional genetic epistasis and synthetic rescue, multiple orthogonal methods in a single study","pmids":["19605495"],"is_preprint":false},{"year":2004,"finding":"In C. elegans, UNC-116/KHC and KLC-2 form a complex orthologous to kinesin-1. KLC-2 also binds UNC-16 (JIP3/JSAP1 orthologue) and the UNC-14 RUN domain protein. Localization of UNC-16 and UNC-14 depends on kinesin-1 (UNC-116 and KLC-2). Double mutant analysis places unc-116, klc-2, unc-16, and unc-14 in the same pathway controlling synaptic vesicle component localization.","method":"Co-immunoprecipitation, genetic epistasis (double-mutant analysis), fluorescent marker localization in mutant backgrounds","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, double-mutant epistasis, and localization studies across multiple labs confirming conserved kinesin-1 pathway","pmids":["15563606"],"is_preprint":false},{"year":2015,"finding":"Rab1A on melanosomes recruits SKIP/PLEKHM2 as a Rab1A-specific effector, and Rab1A, SKIP, and a kinesin-1/(KIF5b+KLC2) motor form a transport complex that mediates anterograde melanosome transport in melanocytes.","method":"Co-immunoprecipitation, knockdown with transport phenotype readout, fluorescence microscopy of melanosome movement","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP identifying complex plus knockdown with defined transport phenotype, single lab, two orthogonal methods","pmids":["25649263"],"is_preprint":false},{"year":2017,"finding":"Crystal structures of both KLC1 and KLC2 TPR domains including the N-terminal capping helix show that this helix adopts two distinct orientations relative to the TPR domain, generating a hydrophobic pocket and electrostatic variations at the groove surface. Ligand binding in the groove can be specific to one or the other N-terminal capping helix orientation, and the capping helix may serve as a protein-protein interaction site.","method":"X-ray crystallography, structural comparative analysis","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — crystal structures determined but functional validation of the capping helix orientations is structural/comparative rather than by mutagenesis in this study","pmids":["29036226"],"is_preprint":false},{"year":2021,"finding":"KLC2 is required for transport of NIS (sodium/iodide symporter) beyond the endoplasmic reticulum to the plasma membrane via a tryptophan-acidic (W-acidic) motif adjacent to G561 in NIS. A G561E NIS variant impairs recognition of this motif by KLC2. Knockdown of Klc2 in rat thyroid cells causes defective NIS maturation and decreased iodide accumulation; morpholino knockdown of klc2 in zebrafish causes hypothyroidism.","method":"Siever sequencing, iodide uptake assays, biochemical interaction assays, siRNA knockdown in thyroid cells, morpholino knockdown in zebrafish, structural bioinformatic analysis","journal":"The Journal of clinical endocrinology and metabolism","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical interaction assays plus loss-of-function in two model systems with defined transport/functional phenotypes, single lab","pmids":["33912899"],"is_preprint":false},{"year":2021,"finding":"KLC2 deficiency in mice causes abnormal mitochondrial transport and downregulation of the GABAA receptor family in cochlear hair cells, leading to low-frequency sensorineural hearing loss. AAV-mediated delivery of wild-type Klc2 cDNA rescued hearing thresholds and reduced outer hair cell loss in Klc2-null mice.","method":"Klc2 knockout mouse model, ABR threshold measurement, immunostaining, AAV gene rescue","journal":"Molecular neurobiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — knockout mouse with defined cellular and functional phenotypes plus AAV rescue, single lab","pmids":["34014435"],"is_preprint":false},{"year":2019,"finding":"KLC2 interacts with Nup358 through a W-acidic motif in Nup358 that is highly conserved among vertebrates. KLC2 and Nup358 form predominantly monomers alone, but their interaction produces 2:2 complexes. The dynein adaptor BicD2 and KLC2 interact simultaneously with Nup358, forming 2:2:2 complexes, suggesting simultaneous recruitment of kinesin-1 and dynein to the nuclear pore.","method":"In vitro reconstitution, biochemical binding assays, analytical ultracentrifugation or similar biophysical characterization","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstituted minimal complex in vitro with stoichiometry determination, W-acidic motif mutagenesis validating the interaction, single lab","pmids":["31756096"],"is_preprint":false},{"year":2025,"finding":"In C. elegans, UNC-83c isoform binds KLC-2 with high affinity to promote kinesin-1 activation for plus-end nuclear movement, while UNC-83a/b isoforms contain an N-terminal inhibitory domain that directly binds kinesin heavy chain UNC-116, reducing its affinity for KLC-2 and allowing dynein-mediated transport. AlphaFold predictions identify spectrin-like repeats in the inhibitory domain, genetically confirmed to be essential for dynein-dependent P cell migration.","method":"Genetic epistasis (C. elegans mutant analysis), in vitro binding assays, AlphaFold structural prediction with genetic validation, isoform-specific functional analysis","journal":"Current biology : CB","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis combined with in vitro binding assays and structural prediction with experimental validation, single lab","pmids":["40925371"],"is_preprint":false},{"year":2018,"finding":"Silencing of KLC1 and KLC2 in neurons inhibited the majority of anterograde HSV enveloped virion transport in axons, while kinesin-1 heavy chain proteins KIF5A, -5B, and -5C also colocalized with HSV particles and were required for transport. Kinesin-3 (KIF1A) silencing had little effect.","method":"siRNA silencing, fluorescence colocalization, live imaging of anterograde transport in neuronal axons","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA knockdown with defined transport phenotype and colocalization, single lab, two orthogonal methods","pmids":["30068641"],"is_preprint":false},{"year":2008,"finding":"UPEC type 1 pilus-mediated invasion of bladder cells requires kinesin-1 light chain KLC2, as well as HDAC6 and microtubules. Silencing KLC2 inhibited host cell invasion by UPEC.","method":"siRNA silencing of KLC2 with invasion assay readout","journal":"The Journal of biological chemistry","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single siRNA knockdown experiment with invasion phenotype, no molecular mechanism for KLC2 action elucidated, single lab","pmids":["18996840"],"is_preprint":false},{"year":2015,"finding":"A homozygous 216-bp deletion in the non-coding upstream region of KLC2 causes SPOAN syndrome by increasing KLC2 expression 48–74% above wild-type levels, as confirmed by luciferase reporter assays in constructs bearing the deletion. Both knockdown and overexpression of klc2 in zebrafish produced a curly-tail phenotype suggestive of neuromuscular disorder.","method":"Whole-genome sequencing, luciferase reporter assay, klc2 knockdown and overexpression in zebrafish, expression analysis in patient fibroblasts and iPSC-derived motor neurons","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reporter assay confirming regulatory effect of deletion, zebrafish gain- and loss-of-function phenotypes, patient-derived cell expression data, single lab with multiple methods","pmids":["26385635"],"is_preprint":false},{"year":2005,"finding":"In C. elegans, UNC-33 (CRMP orthologue) interacts with UNC-14 and KLC-2 in vivo, and its localization to neurites requires UNC-116 (kinesin heavy chain) and KLC-2. Mutations in unc-116 and klc-2 mislocalize UNC-33 to the cell body, implicating kinesin-1 (UNC-116/KLC-2 complex) in axonal transport of UNC-33 for neurite outgrowth.","method":"Co-immunoprecipitation in vivo (C. elegans), mutant localization analysis, genetic screening","journal":"Journal of neurochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo Co-IP plus genetic localization analysis, single lab, two orthogonal methods","pmids":["16236031"],"is_preprint":false},{"year":2025,"finding":"Structural characterization by cryo-EM and SAXS of a minimal KLC2/Nup358/BicD2 complex reveals a rod-like KLC2/Nup358 structure. Addition of BicD2 increases the complex thickness and shifts stoichiometry toward 2:2:2, suggesting cooperative recruitment of kinesin-1 and dynein to Nup358 modulated by oligomeric state.","method":"Cryo-electron microscopy, small angle X-ray scattering (SAXS), reconstituted minimal complex","journal":"bioRxiv : the preprint server for biology","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — cryo-EM and SAXS structural characterization of reconstituted complex, preprint, single lab","pmids":["41648486"],"is_preprint":true},{"year":2025,"finding":"In C. elegans, reduced-function mutation in klc-2 alters the superdiffusive retrograde movement of dense core vesicles (DCVs) in ALA neurons, demonstrating that KLC-2 (kinesin-1 light chain) influences DCV transport dynamics even in the retrograde direction.","method":"Live imaging of DCV trajectories in C. elegans neurons, mathematical modelling of transport statistics across three genetic strains","journal":"Scientific reports","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single imaging study using reduced-function allele, no molecular mechanism identified, single lab","pmids":["40016327"],"is_preprint":false},{"year":2025,"finding":"KLC2 mutants identified in CML myeloid blast phase patients promote cell proliferation, decrease imatinib sensitivity, and impair TGF-β-mediated SMAD2/3 activation while enhancing STAT3 phosphorylation. Both wild-type and mutant KLC2 interact with SMAD2, but mutant KLC2 disrupts TGF-β/SMAD2 signaling.","method":"Immunoprecipitation (KLC2-SMAD2 interaction), immunoblot for STAT3/SMAD2/3 phosphorylation, cell proliferation/apoptosis assays, xenograft mouse model","journal":"Oncology research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP confirming SMAD2 interaction with both WT and mutant KLC2 plus signaling pathway analysis and in vivo xenograft, single lab","pmids":["41502514"],"is_preprint":false}],"current_model":"KLC2 is the cargo-binding light chain subunit of kinesin-1 that recruits diverse cargoes (including AMPA receptors, Smad2, NIS, melanosomes, viral particles, and nuclear envelope proteins) to the motor via its TPR domain's W-acidic motif-binding groove; cargo binding and release are regulated by GSK-3β-mediated phosphorylation of KLC2 (at serine residues), which is in turn controlled upstream by LMTK2 via PP1C-dependent inhibitory phosphorylation of GSK-3β, and by 14-3-3 binding to phospho-Ser575 of KLC2; structural studies reveal that differential residues in the KLC1 vs. KLC2 TPR groove (N343 vs. S328) confer distinct cargo specificities, while a structurally plastic N-terminal capping helix further modulates cargo binding versatility."},"narrative":{"mechanistic_narrative":"KLC2 is the cargo-binding light chain subunit of the kinesin-1 microtubule motor, coupling diverse membrane and protein cargoes to the heavy chain for directional transport along axons and within cells [PMID:15563606, PMID:18361505]. Cargo recognition is mediated by the TPR domain of KLC2, which binds bipartite tryptophan-based (W-acidic) motifs present in over 450 human proteins and in viral proteins, with individual cargoes showing distinct preferences for KLC1 versus KLC2 [PMID:21915095]. Crystal structures of the KLC2 TPR domain define a common cargo-binding groove and explain isoform-specific selectivity: residue S328 (corresponding to KLC1 N343) lacks the carboxylate clamp required for JIP1 binding, and a structurally plastic N-terminal capping helix adopts alternative orientations that modulate the groove surface and cargo versatility [PMID:22470497, PMID:29036226]. Through this groove KLC2 recruits cargoes including AMPA-receptor (GluR1) vesicles [PMID:20534517], the TGF-β effector Smad2 [PMID:21996745, PMID:41502514], the sodium/iodide symporter NIS [PMID:33912899], melanosomes via Rab1A/SKIP [PMID:25649263], and the nucleoporin Nup358, where KLC2 forms a 2:2:2 complex with the dynein adaptor BicD2 to permit simultaneous bidirectional motor recruitment to the nuclear pore [PMID:31756096]. Cargo loading is switched by phosphorylation: GSK-3β phosphorylates KLC2 on serine residues to dissociate cargo such as GluR1 vesicles, an event controlled upstream by LMTK2, which acts through PP1C to increase inhibitory phosphorylation of GSK-3β and thereby promote Smad2 binding to KLC2 [PMID:20534517, PMID:21996745]; 14-3-3 binds KLC2 in a phosphorylation-dependent manner at Ser575 [PMID:11969417]. KLC2 supports kinesin-1–dependent anterograde transport of herpesvirus particles in axons [PMID:30068641], and in C. elegans the KLC-2 orthologue partners with the KASH protein UNC-83 and adaptors UNC-16/UNC-14/UNC-33 to drive nuclear migration and synaptic-cargo localization, with distinct UNC-83 isoforms toggling between kinesin-1 activation and dynein-mediated transport [PMID:19605495, PMID:15563606, PMID:40925371]. A homozygous regulatory deletion that elevates KLC2 expression causes SPOAN syndrome, and Klc2 loss in mice produces sensorineural hearing loss rescuable by gene delivery [PMID:26385635, PMID:34014435].","teleology":[{"year":2002,"claim":"Established that KLC2 is regulated post-translationally by phosphorylation-dependent adaptor binding, linking the motor light chain to signaling control.","evidence":"Proteomic pulldown and MS site mapping of 14-3-3 binding to KLC2 in PC12 cells","pmids":["11969417"],"confidence":"Medium","gaps":["Functional consequence of Ser575 phosphorylation for cargo transport not defined","Kinase responsible for Ser575 phosphorylation not identified"]},{"year":2004,"claim":"Defined a conserved kinesin-1 cargo pathway by placing KLC-2 with the heavy chain and the adaptors UNC-16 and UNC-14 in one genetic pathway controlling synaptic-vesicle localization.","evidence":"Reciprocal Co-IP, double-mutant epistasis, and marker localization in C. elegans","pmids":["15563606"],"confidence":"High","gaps":["Direct binding interface on KLC-2 not mapped","Conservation of adaptor binding to human KLC2 not tested"]},{"year":2005,"claim":"Extended KLC-2 cargo range to a CRMP-family protein required for neurite outgrowth, showing kinesin-1 carries cytoskeletal regulators.","evidence":"In vivo Co-IP and mutant mislocalization analysis in C. elegans","pmids":["16236031"],"confidence":"Medium","gaps":["Whether UNC-33 binds KLC-2 directly or via UNC-14 unresolved"]},{"year":2008,"claim":"Clarified holoenzyme architecture, showing kinesin-1 forms homodimers with homodimerized KLC subunits and that heavy-chain isoforms, not KLC1/KLC2 choice, drive organelle targeting.","evidence":"Reciprocal Co-IP and subcellular fractionation from brain tissue","pmids":["18361505"],"confidence":"Medium","gaps":["Functional distinction between KLC1- and KLC2-containing holoenzymes not resolved here"]},{"year":2010,"claim":"Identified the phosphorylation switch that releases cargo, showing GSK-3β phosphorylation of KLC2 dissociates AMPA-receptor vesicles and modulates synaptic plasticity.","evidence":"Phosphorylation and Co-IP assays plus peptide inhibitor with behavioral readouts in mice","pmids":["20534517"],"confidence":"Medium","gaps":["Exact GSK-3β target serines on KLC2 not fully mapped","Generality of phospho-release across cargoes untested"]},{"year":2011,"claim":"Mapped the upstream signaling that controls KLC2 cargo binding, showing LMTK2–PP1C inhibition of GSK-3β promotes Smad2 loading and TGF-β nuclear signaling.","evidence":"siRNA knockdown, Co-IP, and TGF-β signaling readouts","pmids":["21996745"],"confidence":"Medium","gaps":["Direct phosphosite linkage between GSK-3β activity and Smad2 binding not resolved"]},{"year":2011,"claim":"Defined the general cargo-recognition code, identifying a bipartite W-acidic motif that binds KLC1/KLC2 with isoform preference and is sufficient to recruit kinesin-1 to cargo.","evidence":"Bioinformatics, Co-IP, and functional transport assays using vaccinia as surrogate cargo","pmids":["21915095"],"confidence":"Medium","gaps":["Structural basis of KLC1 vs KLC2 preference not yet resolved in this study"]},{"year":2012,"claim":"Provided the structural basis for KLC isoform-specific cargo selectivity, defining a shared TPR groove and the S328-vs-N343 difference that excludes JIP1 from KLC2.","evidence":"X-ray crystallography of KLC1/KLC2 TPR domains with ITC binding","pmids":["22470497"],"confidence":"High","gaps":["Capping helix contribution not yet characterized","Phospho-regulation not captured in the structures"]},{"year":2015,"claim":"Showed KLC2 mediates organelle transport, recruiting melanosomes via a Rab1A/SKIP effector chain.","evidence":"Co-IP, knockdown with transport phenotype, and melanosome live imaging","pmids":["25649263"],"confidence":"Medium","gaps":["Direct W-acidic motif on SKIP engaging KLC2 not mapped here"]},{"year":2015,"claim":"Linked KLC2 dosage to human disease, establishing that a regulatory deletion raising KLC2 expression causes SPOAN syndrome.","evidence":"Whole-genome sequencing, luciferase reporter, and zebrafish gain/loss-of-function with patient cell expression","pmids":["26385635"],"confidence":"Medium","gaps":["Mechanism by which KLC2 overexpression injures motor neurons unknown","Affected cargo(es) driving pathology not identified"]},{"year":2017,"claim":"Revealed an additional modulator of cargo binding, showing the N-terminal capping helix of the TPR domain adopts two orientations that reshape the binding groove.","evidence":"X-ray crystallography and comparative structural analysis","pmids":["29036226"],"confidence":"Medium","gaps":["Functional role of capping-helix orientations not tested by mutagenesis","Which cargoes select each orientation unknown"]},{"year":2018,"claim":"Demonstrated KLC2 (with KLC1 and KIF5 heavy chains) is required for anterograde axonal transport of an enveloped virus, distinguishing kinesin-1 from kinesin-3 dependence.","evidence":"siRNA silencing, colocalization, and live imaging of HSV transport in axons","pmids":["30068641"],"confidence":"Medium","gaps":["Viral cargo motif engaging KLC2 not defined"]},{"year":2019,"claim":"Reconstituted bidirectional motor recruitment, showing KLC2 and the dynein adaptor BicD2 bind Nup358 simultaneously to form a 2:2:2 complex at the nuclear pore.","evidence":"In vitro reconstitution, W-acidic motif mutagenesis, and stoichiometry determination","pmids":["31756096"],"confidence":"High","gaps":["How opposing motors are coordinated in vivo not established"]},{"year":2021,"claim":"Identified KLC2 as essential for cargo maturation and surface delivery, showing it transports NIS via a W-acidic motif and that loss causes hypothyroidism.","evidence":"Interaction assays, iodide uptake, siRNA in thyroid cells, and zebrafish morpholino knockdown","pmids":["33912899"],"confidence":"Medium","gaps":["Structural basis of NIS motif recognition by KLC2 not solved"]},{"year":2021,"claim":"Established a physiological requirement for KLC2 in cochlear function via mitochondrial transport and GABAA-receptor maintenance, rescuable by gene delivery.","evidence":"Klc2 knockout mouse, ABR thresholds, immunostaining, and AAV rescue","pmids":["34014435"],"confidence":"Medium","gaps":["Direct KLC2 cargo responsible for the hair-cell phenotype not identified"]},{"year":2025,"claim":"Resolved isoform-level regulation of motor selection, showing UNC-83 isoforms either activate kinesin-1 by binding KLC-2 or inhibit it to favor dynein transport.","evidence":"C. elegans genetic epistasis, in vitro binding, and AlphaFold prediction with genetic validation","pmids":["40925371"],"confidence":"Medium","gaps":["Conservation of this isoform switch to human KLC2 untested"]},{"year":2025,"claim":"Connected KLC2 mutation to cancer signaling, showing CML blast-phase mutants disrupt TGF-β/SMAD2 signaling and enhance STAT3 phosphorylation despite retaining SMAD2 binding.","evidence":"Co-IP, signaling immunoblots, proliferation/apoptosis assays, and xenograft","pmids":["41502514"],"confidence":"Medium","gaps":["How mutant KLC2 redirects SMAD2 transport mechanistically unresolved"]},{"year":null,"claim":"How phospho-switches, the capping helix, and competing W-acidic cargoes are integrated to set KLC2 cargo priority in living human cells remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structure of KLC2 bound to a physiological cargo motif in the human context","Phospho-regulatory map across all KLC2 cargoes incomplete","In vivo coordination of kinesin-1 vs dynein recruitment not directly observed in mammalian cells"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[4,5,6,12]},{"term_id":"GO:0038024","term_label":"cargo receptor activity","supporting_discovery_ids":[4,8,10]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[1,7]}],"localization":[{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[7,14]},{"term_id":"GO:0005635","term_label":"nuclear envelope","supporting_discovery_ids":[6,12]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[1,2]}],"pathway":[{"term_id":"R-HSA-9609507","term_label":"Protein localization","supporting_discovery_ids":[4,10,12]},{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[8,14]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[3,20]}],"complexes":["kinesin-1"],"partners":["KIF5B","SMAD2","NUP358","BICD2","14-3-3","UNC-83","GLUR1","SKIP"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9H0B6","full_name":"Kinesin light chain 2","aliases":[],"length_aa":622,"mass_kda":68.9,"function":"Kinesin is a microtubule-associated force-producing protein that plays a role in organelle transport. The light chain functions in coupling of cargo to the heavy chain or in the modulation of its ATPase activity (Probable). Through binding with PLEKHM2 and ARL8B, recruits kinesin-1 to lysosomes and hence direct lysosomes movement toward microtubule plus ends (PubMed:22172677)","subcellular_location":"Cytoplasm, cytoskeleton; Lysosome membrane","url":"https://www.uniprot.org/uniprotkb/Q9H0B6/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/KLC2","classification":"Common Essential","n_dependent_lines":758,"n_total_lines":1208,"dependency_fraction":0.6274834437086093},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000174996","cell_line_id":"CID001432","localizations":[{"compartment":"centrosome","grade":3},{"compartment":"cytoplasmic","grade":3}],"interactors":[{"gene":"KIF5B","stoichiometry":10.0},{"gene":"KLC1","stoichiometry":10.0},{"gene":"KLC4","stoichiometry":4.0},{"gene":"ACTR2","stoichiometry":0.2},{"gene":"ARL6IP6","stoichiometry":0.2},{"gene":"ARL8B","stoichiometry":0.2},{"gene":"KIF5C","stoichiometry":0.2},{"gene":"MTOR","stoichiometry":0.2},{"gene":"KIF5A","stoichiometry":0.2},{"gene":"NAA40","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001432","total_profiled":1310},"omim":[{"mim_id":"620909","title":"KINESIN LIGHT CHAIN 4; KLC4","url":"https://www.omim.org/entry/620909"},{"mim_id":"611729","title":"KINESIN LIGHT CHAIN 2; KLC2","url":"https://www.omim.org/entry/611729"},{"mim_id":"609541","title":"SPASTIC PARAPLEGIA, OPTIC ATROPHY, AND NEUROPATHY; SPOAN","url":"https://www.omim.org/entry/609541"},{"mim_id":"605104","title":"RNA-BINDING FOX1 HOMOLOG 1; RBFOX1","url":"https://www.omim.org/entry/605104"},{"mim_id":"601366","title":"SMAD FAMILY MEMBER 2; SMAD2","url":"https://www.omim.org/entry/601366"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Plasma membrane","reliability":"Supported"},{"location":"Cytosol","reliability":"Supported"},{"location":"Nucleoplasm","reliability":"Additional"},{"location":"Mitochondria","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"brain","ntpm":147.1}],"url":"https://www.proteinatlas.org/search/KLC2"},"hgnc":{"alias_symbol":["FLJ12387"],"prev_symbol":[]},"alphafold":{"accession":"Q9H0B6","domains":[{"cath_id":"-","chopping":"387-422_442-478","consensus_level":"medium","plddt":83.4927,"start":387,"end":478},{"cath_id":"1.20.5","chopping":"15-43_58-137","consensus_level":"high","plddt":93.8644,"start":15,"end":137}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H0B6","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H0B6-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H0B6-F1-predicted_aligned_error_v6.png","plddt_mean":71.88},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=KLC2","jax_strain_url":"https://www.jax.org/strain/search?query=KLC2"},"sequence":{"accession":"Q9H0B6","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9H0B6.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9H0B6/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H0B6"}},"corpus_meta":[{"pmid":"15563606","id":"PMC_15563606","title":"The 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Mass spectrometry identified Ser575 as the phosphorylation site on KLC2 responsible for the in vivo interaction with 14-3-3.\",\n      \"method\": \"Proteomic pulldown from PC12 cells expressing myc-tagged 14-3-3eta, SDS-PAGE/mass spectrometry, interaction studies with KLC2 variants in cultured cells\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS-identified phosphorylation site with cell-based interaction studies, single lab, two orthogonal methods (pulldown + MS site mapping)\",\n      \"pmids\": [\"11969417\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Conventional kinesin holoenzymes are composed of kinesin-1 homodimers (not heterodimers), and KLC subunits also homodimerize. No specificity was found between kinesin-1 isoforms and KLC1/KLC2, suggesting six variant forms of kinesin exist. Different variants associate with biochemically distinct membrane-bounded organelles (MBOs), suggesting kinesin-1 heavy chains target the holoenzyme to specific cargoes.\",\n      \"method\": \"Immunoprecipitation from brain tissue, subcellular fractionation\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP from brain tissue with fractionation, single lab, two orthogonal methods\",\n      \"pmids\": [\"18361505\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"GSK-3β phosphorylates KLC2 on serine residues upon AMPA stimulation, causing dissociation of the GluR1/KLC2 protein complex and release of AMPA-containing vesicles from the kinesin cargo system. A peptide inhibitor of KLC2 phosphorylation (TAT-KLCpCDK) reduced long-term depression formation.\",\n      \"method\": \"Phosphorylation assays, co-immunoprecipitation, peptide inhibitor experiments in neuronal cells, behavioral assays in mice\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP demonstrating complex dissociation upon phosphorylation, peptide inhibitor with cellular and behavioral readouts, single lab\",\n      \"pmids\": [\"20534517\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"LMTK2 signals via protein phosphatase-1C (PP1C) to increase inhibitory phosphorylation of GSK-3β on serine-9, which reduces KLC2 phosphorylation by GSK-3β and promotes binding of the cargo Smad2 to KLC2. siRNA knockdown of LMTK2 reduces Smad2 binding to KLC2 and inhibits TGFβ-induced Smad2 nuclear signalling.\",\n      \"method\": \"siRNA knockdown, co-immunoprecipitation, phosphorylation assays, TGFβ signaling readouts\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and siRNA knockdown with defined signaling readout, single lab, two orthogonal methods\",\n      \"pmids\": [\"21996745\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"A bipartite tryptophan-based (W-acidic) motif present in vaccinia protein A36 and in over 450 human proteins mediates binding to KLC1 and KLC2. Different proteins containing this motif show distinct preferences for KLC1 versus KLC2. Regions containing this motif from cellular proteins can functionally recruit KLC and promote kinesin-1-dependent virus transport.\",\n      \"method\": \"Bioinformatic analysis, functional transport assays using vaccinia as surrogate cargo, co-immunoprecipitation outside infection context\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus functional transport assays, single lab, two orthogonal methods identifying a shared KLC binding motif\",\n      \"pmids\": [\"21915095\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Crystal structures of the TPR domains of KLC1 and KLC2 were determined by X-ray crystallography. KLC2 residue S328 (corresponding to N343 in KLC1) lacks the ability to form a 'carboxylate clamp' for JIP1 binding, explaining why KLC2, unlike KLC1, does not interact with JIP1. A common groove in both KLC1 and KLC2 TPR domains mediates binding of shared cargoes.\",\n      \"method\": \"X-ray crystallography, isothermal titration calorimetry\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure determination combined with ITC binding measurements, directly explaining differential cargo specificity between KLC1 and KLC2\",\n      \"pmids\": [\"22470497\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"In C. elegans, the KASH protein UNC-83 interacts with kinesin-1 light chain KLC-2 (identified by yeast two-hybrid and confirmed by in vitro assays), recruits KLC-2 to the nuclear envelope in heterologous tissue culture, and acts as a cargo adaptor for kinesin-1-dependent nuclear migration. A synthetic KLC-2::KASH fusion protein could partially bypass the requirement for UNC-83 in nuclear migration.\",\n      \"method\": \"Yeast two-hybrid, in vitro binding assay, heterologous tissue culture recruitment assay, genetic epistasis with mutant phenotype analysis, synthetic rescue experiment\",\n      \"journal\": \"Development (Cambridge, England)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — yeast two-hybrid confirmed by in vitro assay plus functional genetic epistasis and synthetic rescue, multiple orthogonal methods in a single study\",\n      \"pmids\": [\"19605495\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"In C. elegans, UNC-116/KHC and KLC-2 form a complex orthologous to kinesin-1. KLC-2 also binds UNC-16 (JIP3/JSAP1 orthologue) and the UNC-14 RUN domain protein. Localization of UNC-16 and UNC-14 depends on kinesin-1 (UNC-116 and KLC-2). Double mutant analysis places unc-116, klc-2, unc-16, and unc-14 in the same pathway controlling synaptic vesicle component localization.\",\n      \"method\": \"Co-immunoprecipitation, genetic epistasis (double-mutant analysis), fluorescent marker localization in mutant backgrounds\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, double-mutant epistasis, and localization studies across multiple labs confirming conserved kinesin-1 pathway\",\n      \"pmids\": [\"15563606\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Rab1A on melanosomes recruits SKIP/PLEKHM2 as a Rab1A-specific effector, and Rab1A, SKIP, and a kinesin-1/(KIF5b+KLC2) motor form a transport complex that mediates anterograde melanosome transport in melanocytes.\",\n      \"method\": \"Co-immunoprecipitation, knockdown with transport phenotype readout, fluorescence microscopy of melanosome movement\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP identifying complex plus knockdown with defined transport phenotype, single lab, two orthogonal methods\",\n      \"pmids\": [\"25649263\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Crystal structures of both KLC1 and KLC2 TPR domains including the N-terminal capping helix show that this helix adopts two distinct orientations relative to the TPR domain, generating a hydrophobic pocket and electrostatic variations at the groove surface. Ligand binding in the groove can be specific to one or the other N-terminal capping helix orientation, and the capping helix may serve as a protein-protein interaction site.\",\n      \"method\": \"X-ray crystallography, structural comparative analysis\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — crystal structures determined but functional validation of the capping helix orientations is structural/comparative rather than by mutagenesis in this study\",\n      \"pmids\": [\"29036226\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"KLC2 is required for transport of NIS (sodium/iodide symporter) beyond the endoplasmic reticulum to the plasma membrane via a tryptophan-acidic (W-acidic) motif adjacent to G561 in NIS. A G561E NIS variant impairs recognition of this motif by KLC2. Knockdown of Klc2 in rat thyroid cells causes defective NIS maturation and decreased iodide accumulation; morpholino knockdown of klc2 in zebrafish causes hypothyroidism.\",\n      \"method\": \"Siever sequencing, iodide uptake assays, biochemical interaction assays, siRNA knockdown in thyroid cells, morpholino knockdown in zebrafish, structural bioinformatic analysis\",\n      \"journal\": \"The Journal of clinical endocrinology and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical interaction assays plus loss-of-function in two model systems with defined transport/functional phenotypes, single lab\",\n      \"pmids\": [\"33912899\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"KLC2 deficiency in mice causes abnormal mitochondrial transport and downregulation of the GABAA receptor family in cochlear hair cells, leading to low-frequency sensorineural hearing loss. AAV-mediated delivery of wild-type Klc2 cDNA rescued hearing thresholds and reduced outer hair cell loss in Klc2-null mice.\",\n      \"method\": \"Klc2 knockout mouse model, ABR threshold measurement, immunostaining, AAV gene rescue\",\n      \"journal\": \"Molecular neurobiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knockout mouse with defined cellular and functional phenotypes plus AAV rescue, single lab\",\n      \"pmids\": [\"34014435\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"KLC2 interacts with Nup358 through a W-acidic motif in Nup358 that is highly conserved among vertebrates. KLC2 and Nup358 form predominantly monomers alone, but their interaction produces 2:2 complexes. The dynein adaptor BicD2 and KLC2 interact simultaneously with Nup358, forming 2:2:2 complexes, suggesting simultaneous recruitment of kinesin-1 and dynein to the nuclear pore.\",\n      \"method\": \"In vitro reconstitution, biochemical binding assays, analytical ultracentrifugation or similar biophysical characterization\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstituted minimal complex in vitro with stoichiometry determination, W-acidic motif mutagenesis validating the interaction, single lab\",\n      \"pmids\": [\"31756096\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In C. elegans, UNC-83c isoform binds KLC-2 with high affinity to promote kinesin-1 activation for plus-end nuclear movement, while UNC-83a/b isoforms contain an N-terminal inhibitory domain that directly binds kinesin heavy chain UNC-116, reducing its affinity for KLC-2 and allowing dynein-mediated transport. AlphaFold predictions identify spectrin-like repeats in the inhibitory domain, genetically confirmed to be essential for dynein-dependent P cell migration.\",\n      \"method\": \"Genetic epistasis (C. elegans mutant analysis), in vitro binding assays, AlphaFold structural prediction with genetic validation, isoform-specific functional analysis\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis combined with in vitro binding assays and structural prediction with experimental validation, single lab\",\n      \"pmids\": [\"40925371\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Silencing of KLC1 and KLC2 in neurons inhibited the majority of anterograde HSV enveloped virion transport in axons, while kinesin-1 heavy chain proteins KIF5A, -5B, and -5C also colocalized with HSV particles and were required for transport. Kinesin-3 (KIF1A) silencing had little effect.\",\n      \"method\": \"siRNA silencing, fluorescence colocalization, live imaging of anterograde transport in neuronal axons\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA knockdown with defined transport phenotype and colocalization, single lab, two orthogonal methods\",\n      \"pmids\": [\"30068641\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"UPEC type 1 pilus-mediated invasion of bladder cells requires kinesin-1 light chain KLC2, as well as HDAC6 and microtubules. Silencing KLC2 inhibited host cell invasion by UPEC.\",\n      \"method\": \"siRNA silencing of KLC2 with invasion assay readout\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single siRNA knockdown experiment with invasion phenotype, no molecular mechanism for KLC2 action elucidated, single lab\",\n      \"pmids\": [\"18996840\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"A homozygous 216-bp deletion in the non-coding upstream region of KLC2 causes SPOAN syndrome by increasing KLC2 expression 48–74% above wild-type levels, as confirmed by luciferase reporter assays in constructs bearing the deletion. Both knockdown and overexpression of klc2 in zebrafish produced a curly-tail phenotype suggestive of neuromuscular disorder.\",\n      \"method\": \"Whole-genome sequencing, luciferase reporter assay, klc2 knockdown and overexpression in zebrafish, expression analysis in patient fibroblasts and iPSC-derived motor neurons\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reporter assay confirming regulatory effect of deletion, zebrafish gain- and loss-of-function phenotypes, patient-derived cell expression data, single lab with multiple methods\",\n      \"pmids\": [\"26385635\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"In C. elegans, UNC-33 (CRMP orthologue) interacts with UNC-14 and KLC-2 in vivo, and its localization to neurites requires UNC-116 (kinesin heavy chain) and KLC-2. Mutations in unc-116 and klc-2 mislocalize UNC-33 to the cell body, implicating kinesin-1 (UNC-116/KLC-2 complex) in axonal transport of UNC-33 for neurite outgrowth.\",\n      \"method\": \"Co-immunoprecipitation in vivo (C. elegans), mutant localization analysis, genetic screening\",\n      \"journal\": \"Journal of neurochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo Co-IP plus genetic localization analysis, single lab, two orthogonal methods\",\n      \"pmids\": [\"16236031\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Structural characterization by cryo-EM and SAXS of a minimal KLC2/Nup358/BicD2 complex reveals a rod-like KLC2/Nup358 structure. Addition of BicD2 increases the complex thickness and shifts stoichiometry toward 2:2:2, suggesting cooperative recruitment of kinesin-1 and dynein to Nup358 modulated by oligomeric state.\",\n      \"method\": \"Cryo-electron microscopy, small angle X-ray scattering (SAXS), reconstituted minimal complex\",\n      \"journal\": \"bioRxiv : the preprint server for biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — cryo-EM and SAXS structural characterization of reconstituted complex, preprint, single lab\",\n      \"pmids\": [\"41648486\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In C. elegans, reduced-function mutation in klc-2 alters the superdiffusive retrograde movement of dense core vesicles (DCVs) in ALA neurons, demonstrating that KLC-2 (kinesin-1 light chain) influences DCV transport dynamics even in the retrograde direction.\",\n      \"method\": \"Live imaging of DCV trajectories in C. elegans neurons, mathematical modelling of transport statistics across three genetic strains\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single imaging study using reduced-function allele, no molecular mechanism identified, single lab\",\n      \"pmids\": [\"40016327\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"KLC2 mutants identified in CML myeloid blast phase patients promote cell proliferation, decrease imatinib sensitivity, and impair TGF-β-mediated SMAD2/3 activation while enhancing STAT3 phosphorylation. Both wild-type and mutant KLC2 interact with SMAD2, but mutant KLC2 disrupts TGF-β/SMAD2 signaling.\",\n      \"method\": \"Immunoprecipitation (KLC2-SMAD2 interaction), immunoblot for STAT3/SMAD2/3 phosphorylation, cell proliferation/apoptosis assays, xenograft mouse model\",\n      \"journal\": \"Oncology research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP confirming SMAD2 interaction with both WT and mutant KLC2 plus signaling pathway analysis and in vivo xenograft, single lab\",\n      \"pmids\": [\"41502514\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"KLC2 is the cargo-binding light chain subunit of kinesin-1 that recruits diverse cargoes (including AMPA receptors, Smad2, NIS, melanosomes, viral particles, and nuclear envelope proteins) to the motor via its TPR domain's W-acidic motif-binding groove; cargo binding and release are regulated by GSK-3β-mediated phosphorylation of KLC2 (at serine residues), which is in turn controlled upstream by LMTK2 via PP1C-dependent inhibitory phosphorylation of GSK-3β, and by 14-3-3 binding to phospho-Ser575 of KLC2; structural studies reveal that differential residues in the KLC1 vs. KLC2 TPR groove (N343 vs. S328) confer distinct cargo specificities, while a structurally plastic N-terminal capping helix further modulates cargo binding versatility.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"KLC2 is the cargo-binding light chain subunit of the kinesin-1 microtubule motor, coupling diverse membrane and protein cargoes to the heavy chain for directional transport along axons and within cells [#7, #1]. Cargo recognition is mediated by the TPR domain of KLC2, which binds bipartite tryptophan-based (W-acidic) motifs present in over 450 human proteins and in viral proteins, with individual cargoes showing distinct preferences for KLC1 versus KLC2 [#4]. Crystal structures of the KLC2 TPR domain define a common cargo-binding groove and explain isoform-specific selectivity: residue S328 (corresponding to KLC1 N343) lacks the carboxylate clamp required for JIP1 binding, and a structurally plastic N-terminal capping helix adopts alternative orientations that modulate the groove surface and cargo versatility [#5, #9]. Through this groove KLC2 recruits cargoes including AMPA-receptor (GluR1) vesicles [#2], the TGF-\\u03b2 effector Smad2 [#3, #20], the sodium/iodide symporter NIS [#10], melanosomes via Rab1A/SKIP [#8], and the nucleoporin Nup358, where KLC2 forms a 2:2:2 complex with the dynein adaptor BicD2 to permit simultaneous bidirectional motor recruitment to the nuclear pore [#12]. Cargo loading is switched by phosphorylation: GSK-3\\u03b2 phosphorylates KLC2 on serine residues to dissociate cargo such as GluR1 vesicles, an event controlled upstream by LMTK2, which acts through PP1C to increase inhibitory phosphorylation of GSK-3\\u03b2 and thereby promote Smad2 binding to KLC2 [#2, #3]; 14-3-3 binds KLC2 in a phosphorylation-dependent manner at Ser575 [#0]. KLC2 supports kinesin-1\\u2013dependent anterograde transport of herpesvirus particles in axons [#14], and in C. elegans the KLC-2 orthologue partners with the KASH protein UNC-83 and adaptors UNC-16/UNC-14/UNC-33 to drive nuclear migration and synaptic-cargo localization, with distinct UNC-83 isoforms toggling between kinesin-1 activation and dynein-mediated transport [#6, #7, #13]. A homozygous regulatory deletion that elevates KLC2 expression causes SPOAN syndrome, and Klc2 loss in mice produces sensorineural hearing loss rescuable by gene delivery [#16, #11].\",\n  \"teleology\": [\n    {\n      \"year\": 2002,\n      \"claim\": \"Established that KLC2 is regulated post-translationally by phosphorylation-dependent adaptor binding, linking the motor light chain to signaling control.\",\n      \"evidence\": \"Proteomic pulldown and MS site mapping of 14-3-3 binding to KLC2 in PC12 cells\",\n      \"pmids\": [\"11969417\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of Ser575 phosphorylation for cargo transport not defined\", \"Kinase responsible for Ser575 phosphorylation not identified\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Defined a conserved kinesin-1 cargo pathway by placing KLC-2 with the heavy chain and the adaptors UNC-16 and UNC-14 in one genetic pathway controlling synaptic-vesicle localization.\",\n      \"evidence\": \"Reciprocal Co-IP, double-mutant epistasis, and marker localization in C. elegans\",\n      \"pmids\": [\"15563606\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct binding interface on KLC-2 not mapped\", \"Conservation of adaptor binding to human KLC2 not tested\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Extended KLC-2 cargo range to a CRMP-family protein required for neurite outgrowth, showing kinesin-1 carries cytoskeletal regulators.\",\n      \"evidence\": \"In vivo Co-IP and mutant mislocalization analysis in C. elegans\",\n      \"pmids\": [\"16236031\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether UNC-33 binds KLC-2 directly or via UNC-14 unresolved\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Clarified holoenzyme architecture, showing kinesin-1 forms homodimers with homodimerized KLC subunits and that heavy-chain isoforms, not KLC1/KLC2 choice, drive organelle targeting.\",\n      \"evidence\": \"Reciprocal Co-IP and subcellular fractionation from brain tissue\",\n      \"pmids\": [\"18361505\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional distinction between KLC1- and KLC2-containing holoenzymes not resolved here\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Identified the phosphorylation switch that releases cargo, showing GSK-3\\u03b2 phosphorylation of KLC2 dissociates AMPA-receptor vesicles and modulates synaptic plasticity.\",\n      \"evidence\": \"Phosphorylation and Co-IP assays plus peptide inhibitor with behavioral readouts in mice\",\n      \"pmids\": [\"20534517\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Exact GSK-3\\u03b2 target serines on KLC2 not fully mapped\", \"Generality of phospho-release across cargoes untested\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Mapped the upstream signaling that controls KLC2 cargo binding, showing LMTK2\\u2013PP1C inhibition of GSK-3\\u03b2 promotes Smad2 loading and TGF-\\u03b2 nuclear signaling.\",\n      \"evidence\": \"siRNA knockdown, Co-IP, and TGF-\\u03b2 signaling readouts\",\n      \"pmids\": [\"21996745\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct phosphosite linkage between GSK-3\\u03b2 activity and Smad2 binding not resolved\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Defined the general cargo-recognition code, identifying a bipartite W-acidic motif that binds KLC1/KLC2 with isoform preference and is sufficient to recruit kinesin-1 to cargo.\",\n      \"evidence\": \"Bioinformatics, Co-IP, and functional transport assays using vaccinia as surrogate cargo\",\n      \"pmids\": [\"21915095\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of KLC1 vs KLC2 preference not yet resolved in this study\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Provided the structural basis for KLC isoform-specific cargo selectivity, defining a shared TPR groove and the S328-vs-N343 difference that excludes JIP1 from KLC2.\",\n      \"evidence\": \"X-ray crystallography of KLC1/KLC2 TPR domains with ITC binding\",\n      \"pmids\": [\"22470497\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Capping helix contribution not yet characterized\", \"Phospho-regulation not captured in the structures\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Showed KLC2 mediates organelle transport, recruiting melanosomes via a Rab1A/SKIP effector chain.\",\n      \"evidence\": \"Co-IP, knockdown with transport phenotype, and melanosome live imaging\",\n      \"pmids\": [\"25649263\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct W-acidic motif on SKIP engaging KLC2 not mapped here\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Linked KLC2 dosage to human disease, establishing that a regulatory deletion raising KLC2 expression causes SPOAN syndrome.\",\n      \"evidence\": \"Whole-genome sequencing, luciferase reporter, and zebrafish gain/loss-of-function with patient cell expression\",\n      \"pmids\": [\"26385635\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which KLC2 overexpression injures motor neurons unknown\", \"Affected cargo(es) driving pathology not identified\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Revealed an additional modulator of cargo binding, showing the N-terminal capping helix of the TPR domain adopts two orientations that reshape the binding groove.\",\n      \"evidence\": \"X-ray crystallography and comparative structural analysis\",\n      \"pmids\": [\"29036226\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional role of capping-helix orientations not tested by mutagenesis\", \"Which cargoes select each orientation unknown\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Demonstrated KLC2 (with KLC1 and KIF5 heavy chains) is required for anterograde axonal transport of an enveloped virus, distinguishing kinesin-1 from kinesin-3 dependence.\",\n      \"evidence\": \"siRNA silencing, colocalization, and live imaging of HSV transport in axons\",\n      \"pmids\": [\"30068641\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Viral cargo motif engaging KLC2 not defined\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Reconstituted bidirectional motor recruitment, showing KLC2 and the dynein adaptor BicD2 bind Nup358 simultaneously to form a 2:2:2 complex at the nuclear pore.\",\n      \"evidence\": \"In vitro reconstitution, W-acidic motif mutagenesis, and stoichiometry determination\",\n      \"pmids\": [\"31756096\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How opposing motors are coordinated in vivo not established\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified KLC2 as essential for cargo maturation and surface delivery, showing it transports NIS via a W-acidic motif and that loss causes hypothyroidism.\",\n      \"evidence\": \"Interaction assays, iodide uptake, siRNA in thyroid cells, and zebrafish morpholino knockdown\",\n      \"pmids\": [\"33912899\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of NIS motif recognition by KLC2 not solved\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Established a physiological requirement for KLC2 in cochlear function via mitochondrial transport and GABAA-receptor maintenance, rescuable by gene delivery.\",\n      \"evidence\": \"Klc2 knockout mouse, ABR thresholds, immunostaining, and AAV rescue\",\n      \"pmids\": [\"34014435\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct KLC2 cargo responsible for the hair-cell phenotype not identified\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Resolved isoform-level regulation of motor selection, showing UNC-83 isoforms either activate kinesin-1 by binding KLC-2 or inhibit it to favor dynein transport.\",\n      \"evidence\": \"C. elegans genetic epistasis, in vitro binding, and AlphaFold prediction with genetic validation\",\n      \"pmids\": [\"40925371\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Conservation of this isoform switch to human KLC2 untested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Connected KLC2 mutation to cancer signaling, showing CML blast-phase mutants disrupt TGF-\\u03b2/SMAD2 signaling and enhance STAT3 phosphorylation despite retaining SMAD2 binding.\",\n      \"evidence\": \"Co-IP, signaling immunoblots, proliferation/apoptosis assays, and xenograft\",\n      \"pmids\": [\"41502514\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"How mutant KLC2 redirects SMAD2 transport mechanistically unresolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How phospho-switches, the capping helix, and competing W-acidic cargoes are integrated to set KLC2 cargo priority in living human cells remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structure of KLC2 bound to a physiological cargo motif in the human context\", \"Phospho-regulatory map across all KLC2 cargoes incomplete\", \"In vivo coordination of kinesin-1 vs dynein recruitment not directly observed in mammalian cells\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [4, 5, 6, 12]},\n      {\"term_id\": \"GO:0038024\", \"supporting_discovery_ids\": [4, 8, 10]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [1, 7]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [7, 14]},\n      {\"term_id\": \"GO:0005635\", \"supporting_discovery_ids\": [6, 12]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [1, 2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-9609507\", \"supporting_discovery_ids\": [4, 10, 12]},\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [8, 14]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [3, 20]}\n    ],\n    \"complexes\": [\"kinesin-1\"],\n    \"partners\": [\"KIF5B\", \"Smad2\", \"Nup358\", \"BicD2\", \"14-3-3\", \"UNC-83\", \"GluR1\", \"SKIP\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}