{"gene":"YWHAG","run_date":"2026-06-11T09:02:06","timeline":{"discoveries":[{"year":2006,"finding":"14-3-3γ binds directly to MDMX phosphorylated at Ser367 by Chk1 in response to UV irradiation, causing cytoplasmic retention of MDMX and suppression of MDMX-enhanced p53 ubiquitination, leading to p53 stabilization. The interaction requires phosphorylation of MDMX; the K50E mutant of 14-3-3γ that cannot bind MDMX fails to stabilize p53. siRNA ablation of 14-3-3γ reduces UV-induced p53 levels and G1 arrest.","method":"Immuno-affinity purification coupled with mass spectrometry, in vitro binding assay, co-immunoprecipitation, kinase-dead Chk1 mutant, Chk1 inhibitor UCN-01, siRNA knockdown, p53 ubiquitination assay","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, in vitro direct binding, active-site/mutant controls, kinase inhibitor, siRNA, multiple orthogonal methods in a single rigorous study","pmids":["16511572"],"is_preprint":false},{"year":2010,"finding":"14-3-3γ forms a ternary complex with Chk1 (phosphorylated at Ser296 by autophosphorylation after ATR activation) and Cdc25A, mediating Cdc25A phosphorylation and proteasomal degradation to block premature mitotic entry after UV-induced DNA damage. Replacement of endogenous Chk1 with a Ser296Ala mutant causes premature mitotic entry after UV irradiation.","method":"Co-immunoprecipitation, phospho-specific antibodies, Chk1 Ser296Ala mutant replacement, UV irradiation, cell cycle analysis","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, phospho-site mutant rescue experiment, multiple orthogonal methods, clear functional readout","pmids":["20639859"],"is_preprint":false},{"year":2013,"finding":"Plk1 phosphorylated at Ser99 (by a PI3K/Akt-dependent mechanism) creates a docking site for 14-3-3γ; this interaction stimulates Plk1 catalytic activity. Knockdown of 14-3-3γ or expression of a Ser99-phospho-blocking Plk1 mutant causes prometaphase/metaphase arrest by activating the spindle assembly checkpoint, demonstrating that this interaction is required for proper metaphase-to-anaphase transition.","method":"Co-immunoprecipitation, phospho-specific antibodies, Plk1 Ser99Ala mutant, siRNA knockdown of 14-3-3γ, PI3K/Akt inhibitors, kinase activity assay, mitotic arrest analysis","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, phospho-site mutant, kinase activity assay, siRNA, multiple orthogonal methods with clear functional readout","pmids":["23695676"],"is_preprint":false},{"year":2012,"finding":"14-3-3γ dimers act as a scaffold that bridges CtBP1-S/BARS to PI(4)KIIIβ at the Golgi complex, coupling carrier budding and fission processes. The complex is stabilized by PKD- and PAK-mediated phosphorylation. Disrupting the association of these proteins inhibits fission of elongating post-Golgi carrier precursors.","method":"Co-immunoprecipitation, protein complex reconstitution, phosphorylation assays with PKD and PAK, dominant-negative disruption, live-cell imaging of carrier fission","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, functional disruption of complex with defined cellular phenotype, phosphorylation-dependent complex stabilization, multiple orthogonal methods","pmids":["22366688"],"is_preprint":false},{"year":2012,"finding":"14-3-3γ negatively regulates steroidogenesis in MA-10 Leydig cells by binding to phospho-Ser194 in the START domain of StAR protein in a hormone-dependent manner, keeping StAR in an unfolded, inactive state. Over time, 14-3-3γ homodimerizes and dissociates from StAR, permitting maximal mitochondrial steroid formation. Silencing 14-3-3γ potentiates steroidogenesis.","method":"Mass spectrometry identification in native mitochondrial complexes, immunoprecipitation, site-directed mutagenesis (Ser194 binding site), siRNA knockdown, steroid output assay, cAMP stimulation","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — MS identification, Co-IP, phospho-site mutagenesis, siRNA, functional steroid output assay — multiple orthogonal methods in single study","pmids":["22427666"],"is_preprint":false},{"year":2014,"finding":"Ser58 phosphorylation and Lys49 acetylation of 14-3-3γ regulate its homodimerization and interaction with StAR in a coordinated, time-dependent manner during cAMP-induced steroidogenesis. Blocking either modification further induces steroidogenesis and reduces lipid storage.","method":"TAT-peptide blocking of phosphorylation (Ser58) and acetylation (Lys49) sites, co-immunoprecipitation, steroid output and lipid storage assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — peptide-blocking approach, Co-IP, functional readout; single lab, two orthogonal methods","pmids":["25086053"],"is_preprint":false},{"year":2014,"finding":"Phosphorylated tyrosine hydroxylase (TH) at Ser19 binds 14-3-3γ with high affinity (Kd ~3.2 nM), forming complexes of one TH tetramer with one or two 14-3-3γ dimers. 14-3-3γ binding inhibits PKA-mediated phosphorylation of TH at Ser40 (3.5-fold reduction), suggesting Ser40 has reduced accessibility in the complex. TH-pS40 alone does not detectably bind 14-3-3γ.","method":"Native mass spectrometry, surface plasmon resonance, electron microscopy, phosphatase kinetics, in vitro kinase assay","journal":"Molecular & cellular proteomics","confidence":"High","confidence_rationale":"Tier 1 / Strong — native MS stoichiometry, SPR binding kinetics, EM structural analysis, in vitro enzymatic assay; multiple orthogonal methods in one rigorous study","pmids":["24947669"],"is_preprint":false},{"year":2017,"finding":"PAK6 phosphorylates 14-3-3γ at Ser59, and this phosphorylation acts as a switch that dissociates 14-3-3γ from client proteins including LRRK2 (at phospho-Ser935), causing LRRK2 dephosphorylation. A constitutively active PAK6 rescues G2019S LRRK2-associated neurite shortening through this phosphorylation of 14-3-3γ.","method":"Co-immunoprecipitation of PAK6 interactome, in vitro kinase assay with phospho-site identification (Ser59), neurite length measurement in neurons, LRRK2 phospho-Ser935 western blot, PAK6 constitutively active mutant","journal":"Frontiers in molecular neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP interactome, in vitro kinase assay with phospho-site identification, loss-of-function (14-3-3γ dissociation), neuronal rescue experiment; multiple orthogonal methods","pmids":["29311810"],"is_preprint":false},{"year":2006,"finding":"Phosphorylated Hsp20 (HspB6) forms a tight complex with 14-3-3γ in which a dimer of 14-3-3γ binds a dimer of Hsp20. 14-3-3γ increases the chaperone activity of phosphorylated Hsp20 when insulin is used as a model substrate. Unphosphorylated Hsp20 and its S16D phosphomimetic mutant do not interact with 14-3-3γ.","method":"Size-exclusion chromatography, chemical crosslinking, in vitro chaperone activity assay with insulin substrate","journal":"Molecular and cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro reconstitution of complex, functional chaperone assay; single lab, two orthogonal methods","pmids":["17109079"],"is_preprint":false},{"year":2003,"finding":"14-3-3γ is poly-ADP-ribosylated in the nucleus after traumatic brain injury, and nuclear poly-ADP-ribosylation of 14-3-3γ is completely inhibited by the PARP-1 inhibitor dose that produces profound memory disturbances, linking this modification to spatial memory acquisition.","method":"Proteomics identification of poly-ADP-ribosylated peptides from brain, PARP-1 inhibitor treatment in vivo (INH2BP), Morris water maze behavioral testing","journal":"Journal of neurochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proteomics-based identification of ribosylation site, in vivo pharmacological intervention with functional behavioral readout; single lab","pmids":["12694396"],"is_preprint":false},{"year":2005,"finding":"Dimeric 14-3-3γ simultaneously binds both AICD (amyloid beta-protein precursor intracellular domain) and FE65, facilitating FE65-dependent gene transactivation by enhancing AICD-FE65 association. The interaction requires the VTPEER motif (residues 667-672) of AICD; phosphorylation of AICD at Thr668 within this motif inhibits 14-3-3γ binding and blocks gene transactivation. 14-3-3γ binds to a region between the WW domain and first PTB domain of FE65.","method":"Co-immunoprecipitation, in vitro binding assay, deletion/point mutants of AICD and FE65, gene transactivation reporter assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP, in vitro direct binding, multiple domain-mapping mutants, functional reporter assay; multiple orthogonal methods in one study","pmids":["16223726"],"is_preprint":false},{"year":2003,"finding":"Endogenous 14-3-3γ co-immunoprecipitates with c-Raf-1 and p-Raf-259 in primary astrocyte cultures, suggesting 14-3-3γ links Raf to signaling pathways controlling cell growth and ischemia-induced apoptosis. The induction of 14-3-3γ in ischemic astrocytes was not suppressed by PI3K or MAP kinase inhibitors.","method":"Co-immunoprecipitation, Northern/Western blot, pharmacological inhibitors (U0126, LY294002), anaerobic ischemia model in cultured astrocytes","journal":"Glia","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — single Co-IP with Raf, replicated in two Raf forms, inhibitor experiments showing pathway independence; single lab","pmids":["12730952"],"is_preprint":false},{"year":2006,"finding":"14-3-3γ associates with phosphorylated GFAP (specifically at Ser8 in the head domain) in a phosphorylation- and cell-cycle-dependent manner, with increased association during G2/M phase. Overexpression of 14-3-3γ destroys the integrity and affects the movement of GFAP intermediate filaments in astrocytes.","method":"Co-immunoprecipitation, domain deletion and Ser-to-Ala substitution mutants of GFAP, cell-cycle synchronization, live imaging of intermediate filament dynamics, immunofluorescence","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP with phospho-site mutagenesis, cell-cycle synchronization, live imaging of functional consequence; multiple orthogonal methods","pmids":["17032734"],"is_preprint":false},{"year":2005,"finding":"Under ischemia, 14-3-3γ (the sole 14-3-3 isoform inducible by ischemia in astrocytes) binds phospho-Bad, preventing Bad translocation to mitochondria and inhibiting apoptosis. Overexpression of 14-3-3γ promotes astrocyte survival; antisense suppression enhances apoptosis under ischemia.","method":"Co-immunoprecipitation of endogenous 14-3-3γ with p-Bad, overexpression, antisense knockdown, cell death quantification, isoform-specific western blots","journal":"Journal of cerebral blood flow and metabolism","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP of endogenous complex, gain- and loss-of-function with survival readout; single lab","pmids":["15660102"],"is_preprint":false},{"year":2002,"finding":"Endogenous 14-3-3γ co-immunoprecipitates with detergent-soluble actin in astrocytes, and this association increases after 4 h of ischemia. 14-3-3γ co-localizes with F-actin during cell division (forming a ring-like structure around daughter nuclei) and in surviving ischemic astrocytes, but dissociates from actin filaments in apoptotic astrocytes.","method":"Reciprocal co-immunoprecipitation, immunofluorescence co-localization, ischemia model","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — reciprocal Co-IP, immunofluorescence co-localization across multiple functional states; single lab","pmids":["12176032"],"is_preprint":false},{"year":1999,"finding":"14-3-3γ is phosphorylated by multiple PKC isoforms (alpha, beta, gamma, theta, delta) in a PDGF-dependent manner in vascular smooth muscle cells. 14-3-3γ also interacts with the signal transduction protein Raf-1, suggesting it links Raf to PKC signaling.","method":"Co-immunoprecipitation with PKC isoforms, PKC inhibitor treatment, PKC activator stimulation, phosphorylation assay","journal":"DNA and cell biology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP with multiple PKC isoforms, pharmacological activation/inhibition; single lab, two orthogonal approaches","pmids":["10433554"],"is_preprint":false},{"year":2012,"finding":"14-3-3γ interaction with phospholipid bilayers is stimulated when complexed with a Ser19-phosphorylated tyrosine hydroxylase peptide. Membrane binding is isoform-specific and depends on histidine residues His158 and His195 (unique to the γ isoform) at the convex lateral side, as shown by site-directed mutagenesis. Electrostatic analysis and molecular dynamics indicate that phosphopeptide-bound 14-3-3γ has an optimal electrostatic potential for membrane interaction through N-terminal amphipathic helices.","method":"Surface plasmon resonance (membrane binding assay), site-directed mutagenesis (His158, His195), molecular dynamics simulation, electrostatic analysis of crystal structures","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 1 / Moderate — SPR biophysical assay, site-directed mutagenesis with functional validation, MD simulation; single lab but multiple orthogonal methods","pmids":["23189152"],"is_preprint":false},{"year":2016,"finding":"14-3-3γ identified as a binding partner of ANO1 (anoctamin-1) by yeast two-hybrid screening; the Thr9 residue of ANO1 is critical for the interaction. 14-3-3γ enhances surface expression of ANO1 (anterograde trafficking). Gene silencing of 14-3-3γ and/or ANO1 inhibits migration and invasion of glioblastoma cells.","method":"Yeast two-hybrid screening, co-immunoprecipitation, site-directed mutagenesis (Thr9), cell surface expression assay, siRNA knockdown, migration/invasion assays","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast two-hybrid + Co-IP + mutagenesis + functional cell assay; single lab","pmids":["27212225"],"is_preprint":false},{"year":2014,"finding":"14-3-3γ binds TRPM4b at its N-terminus via a Ser88-dependent interaction (GST pull-down and Co-IP). Overexpression of 14-3-3γ increases TRPM4b plasma membrane expression measured by whole-cell electrophysiology and surface biotinylation; shRNA against 14-3-3γ greatly reduces TRPM4b surface expression and attenuates glutamate-induced TRPM4b currents and neuronal cell death.","method":"Yeast two-hybrid, GST pull-down, co-immunoprecipitation, whole-cell patch clamp, cell surface biotinylation, shRNA knockdown, glutamate-induced cell death assay","journal":"Molecular brain","confidence":"High","confidence_rationale":"Tier 2 / Strong — yeast two-hybrid + GST pull-down + Co-IP + electrophysiology + surface biotinylation + shRNA functional rescue; multiple orthogonal methods","pmids":["25047048"],"is_preprint":false},{"year":2014,"finding":"14-3-3γ binds plakoglobin in a PKCμ-dependent (PKD-dependent) manner and mediates microtubule-dependent (KIF5B-KLC1 motor complex-dependent) transport of plakoglobin to cell borders, initiating desmosome assembly. Loss of 14-3-3γ reduces cell-cell adhesion and desmosome formation both in vitro (HCT116 cells) and in vivo (mouse testis), leading to defects in spermatogenesis.","method":"Co-immunoprecipitation, PKCμ inhibitor, microtubule disruption, KIF5B knockdown, immunofluorescence, in vivo testis phenotype analysis","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP with kinase inhibitor, motor protein knockdown, in vitro and in vivo loss-of-function with defined cellular/tissue phenotypes; multiple orthogonal methods","pmids":["24610948"],"is_preprint":false},{"year":2015,"finding":"Loss of 14-3-3γ leads to centrosome amplification via phosphorylation of NPM1 at Thr199, causing early centriole disjunction and centrosome hyper-duplication, which results in aneuploidy and increased tumor formation. 14-3-3γ localizes to the centrosome. Expression of a constitutively active 14-3-3-binding-defective Cdc25C (S216A) mutant in 14-3-3γ-knockdown cells increases multipolar spindle formation.","method":"14-3-3γ knockdown, NPM1 phospho-Thr199 detection, centriole duplication assay, centrosome immunofluorescence, in vivo tumor formation, Cdc25C S216A mutant expression, spindle analysis","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with defined molecular (NPM1 phosphorylation) and cellular (centrosome amplification) phenotypes; single lab, multiple readouts","pmids":["27253419"],"is_preprint":false},{"year":2009,"finding":"Ischemia-induced upregulation of 14-3-3γ in astrocytes is mediated by activation of the JNK/c-Jun/AP-1 pathway. Only the JNK inhibitor SP600125 (not ERK, p38, or PI3K inhibitors) blocked ischemia-induced 14-3-3γ upregulation. Nuclear translocation of p-c-Jun under ischemia and AP-1 inhibition by curcumin also suppressed 14-3-3γ induction.","method":"Selective kinase inhibitors (SP600125, U0126, SB203580, LY294002), curcumin AP-1 inhibition, nuclear translocation of p-c-Jun by immunofluorescence, Western blot for 14-3-3γ, in vitro ischemia model","journal":"Journal of neurochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological pathway dissection with multiple inhibitors, nuclear translocation imaging; single lab","pmids":["19393026"],"is_preprint":false},{"year":2010,"finding":"Overexpression of 14-3-3γ in NIH3T3 cells induces oncogenic transformation (focus formation and tumor formation in SCID mice). Transformation requires activation of both MAPK and PI3K signaling pathways. 14-3-3γ co-immunoprecipitates with PI3K and TSC2, indicating it can stimulate PI3K signaling at two points.","method":"Focus formation assay, soft agar colony formation, tumor formation in SCID mice, PI3K/MAPK inhibitor treatment, co-immunoprecipitation with PI3K and TSC2","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo tumor formation, Co-IP with signaling proteins, pharmacological pathway inhibition; single lab, multiple methods","pmids":["20628654"],"is_preprint":false},{"year":2012,"finding":"The N-terminal variable region II (VRII) of 14-3-3γ (within the first 40 amino acids) is required for its oncogenic activity (activation of PI3K and MAPK signaling and cellular transformation). Two residues within VRII are required and two contribute to the γ-specific phenotypes; swapping this region with the equivalent region from the tumor suppressor 14-3-3σ switches the functional outcome.","method":"14-3-3γ/σ chimeric protein constructs, focus formation assay, soft agar growth, PI3K/MAPK activation assays, individual amino acid substitutions","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain-swap chimeras and individual amino acid substitutions with transformation functional assay; single lab","pmids":["23115241"],"is_preprint":false},{"year":2010,"finding":"14-3-3γ protects p21 from MDMX-mediated proteasomal degradation (independent of p53) by competing with p21 for MDMX binding in a dose-dependent manner. Overexpression of 14-3-3γ extends p21 half-life and causes p21-dependent G1 arrest in p53-null cells. In response to DNA damage, the 14-3-3γ-MDMX complex increases while the MDMX-p21 complex decreases.","method":"Co-immunoprecipitation in vitro with purified proteins and in cells, p21 half-life measurement, G1 arrest assay in p53-null cells, siRNA knockdown of 14-3-3γ","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vitro Co-IP with purified proteins + in-cell Co-IP + half-life measurement + loss-of-function siRNA + functional cell cycle readout; multiple orthogonal methods","pmids":["21148311"],"is_preprint":false},{"year":2012,"finding":"14-3-3γ binds to and undergoes poly-ADP-ribosylation; in the context of the MALM (Mieap-induced accumulation of lysosome-like organelles within mitochondria) process, 14-3-3γ localizes within mitochondria and interacts with Mieap. Deficiency of 14-3-3γ does not affect accumulation of Mieap or lysosomal proteins within mitochondria but dramatically inhibits elimination of oxidized mitochondrial proteins.","method":"2DICAL (two-dimensional image-converted LC-MS) proteomics of immunoprecipitated Mieap complex, co-immunoprecipitation of exogenous and endogenous proteins, subcellular fractionation, immunofluorescence, 14-3-3γ knockdown with functional readout","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS-based identification + Co-IP + subcellular localization + functional knockdown; single lab","pmids":["22532927"],"is_preprint":false},{"year":2015,"finding":"14-3-3γ interacts with ERK1c as part of a PI4KIIIβ-14-3-3γ complex that mediates Golgi translocation of ERK1c during prophase/prometaphase. CDK1 phosphorylates ERK1c at Ser343, enabling complex assembly. PKD-mediated phosphorylation of PI4KIIIβ stabilizes the complex. The complex assembly induces ERK1c Golgi translocation where it is activated by MEK1b to induce Golgi fragmentation.","method":"Co-immunoprecipitation, CDK1 phosphorylation assay, PKD inhibitor, dominant-negative constructs, time-lapse imaging to determine translocation timing, kinase assays","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, kinase assays, pharmacological inhibitors, live imaging; single lab","pmids":["26459638"],"is_preprint":false},{"year":2014,"finding":"14-3-3γ co-immunoprecipitates with phospho-Bad(S112) in LPS-treated cardiomyocytes, and overexpressed 14-3-3γ promotes Bad(S112) phosphorylation and sequesters p-Bad, causing Bcl-2 dissociation from the Bad/Bcl-2 complex and Bcl-2 translocation to mitochondria, preventing mPTP opening and cytochrome c release.","method":"Co-immunoprecipitation, subcellular fractionation (cytosolic/mitochondrial), flow cytometry (apoptosis, MMP), mitochondrial swelling assay (mPTP), Western blot for Bcl-2 family members","journal":"International immunopharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, subcellular fractionation with functional readout, multiple downstream markers; single lab","pmids":["24957688"],"is_preprint":false},{"year":2014,"finding":"14-3-3γ binds directly to p-β-catenin Ser37 (but not p-Bad, p-Ask-1, p-p53, or Bax) in cortical neurons as shown by Co-IP and FRET. During oxygen-glucose deprivation, 14-3-3γ translocates to the nucleus correlating with increased nuclear p-β-catenin Ser37. 14-3-3γ overexpression reduces Bax expression and prevents p-β-catenin-Ser37-dependent Bax upregulation and cell death.","method":"Co-immunoprecipitation, FRET, nuclear fractionation, OGD model, 14-3-3γ overexpression/knockdown, β-catenin S37A mutant, Bax measurement, cell viability","journal":"Cell death & disease","confidence":"High","confidence_rationale":"Tier 2 / Strong — FRET (direct interaction), Co-IP, phospho-site mutant S37A, loss- and gain-of-function with multiple functional readouts; multiple orthogonal methods","pmids":["24743739"],"is_preprint":false},{"year":2013,"finding":"14-3-3γ specifically binds to 5'-AGCT-3' repeats in IgH switch regions and interacts directly with the C-terminal region of activation-induced cytidine deaminase (AID), targeting AID to switch regions to mediate class switch recombination. Induction of 14-3-3γ expression in B cells requires NF-κB recruitment to the 14-3-3γ promoter, which promotes CFP1-mediated H3K4me3 enrichment, followed by E2A binding for sustained expression.","method":"ChIP assays, B cell stimulation with CSR-inducing stimuli, NF-κB inhibition, promoter analysis, 14-3-3γ expression kinetics","journal":"Journal of immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP, pharmacological NF-κB inhibition, B cell stimulation kinetics; single lab; mechanistic detail on transcriptional regulation","pmids":["23851690"],"is_preprint":false},{"year":2015,"finding":"Protein kinase CK2 interacts strongly with 14-3-3γ at the neuromuscular junction and phosphorylates 14-3-3γ at serine residue 235.","method":"Co-immunoprecipitation, in vitro kinase assay with phospho-site mapping","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP + in vitro kinase assay with specific phospho-site identification; single lab","pmids":["26198629"],"is_preprint":false},{"year":2015,"finding":"14-3-3γ binds deubiquitinase USP37, which stabilizes 14-3-3γ by preventing its ubiquitin-dependent proteasomal degradation through its catalytic deubiquitinase activity.","method":"Co-immunoprecipitation (binding partner screen), ubiquitination assay, USP37 catalytic mutant, protein stability assay","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP, ubiquitination assay, catalytic mutant; single lab","pmids":["26427597"],"is_preprint":false},{"year":2014,"finding":"p53 interacts with the C-terminal domain of 14-3-3γ, induces 14-3-3γ ubiquitination, and promotes proteasome-mediated degradation of 14-3-3γ. MG132 (26S proteasome inhibitor) blocks this effect. Wild-type but not mutant p53 (R175H) suppresses 14-3-3γ protein levels.","method":"Co-immunoprecipitation, ubiquitination assay, proteasome inhibitor (MG132), wild-type vs. mutant p53 expression, protein half-life analysis","journal":"International journal of oncology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP, ubiquitination assay, proteasome inhibitor, mutant controls; single lab","pmids":["25384678"],"is_preprint":false},{"year":2012,"finding":"14-3-3γ binds preferentially to supercoiled DNA over linear DNA, with particular affinity for cruciform DNA structures. In HCT-116 cells, 14-3-3γ co-localizes with DNA cruciforms by confocal microscopy.","method":"Electrophoretic mobility shift assay, competition with magnetic beads (linear vs. supercoiled DNA), confocal microscopy with cruciform-detecting probes","journal":"Journal of biomolecular structure & dynamics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — EMSA, magnetic bead competition assay, confocal co-localization; single lab, in vitro binding confirmed in cells","pmids":["22856523"],"is_preprint":false},{"year":2012,"finding":"14-3-3γ interacts with eukaryotic translation initiation factor eIF1AX and ribosomal protein RPS7, as confirmed by Co-IP with mass spectrometry and FRET/co-localization. 14-3-3γ positively regulates protein synthesis by affecting eIF1AX and RPS7 expression levels and mTOR pathway activity in bovine mammary epithelial cells.","method":"Co-immunoprecipitation with MALDI-TOF/TOF mass spectrometry, FRET, co-localization, 14-3-3γ overexpression and silencing with protein synthesis readout","journal":"Archives of biochemistry and biophysics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP+MS identification confirmed by FRET, gain- and loss-of-function; single lab","pmids":["25281768"],"is_preprint":false},{"year":2018,"finding":"14-3-3γ directly interacts with RGS14 at two distinct sites: a phosphorylation-independent site and a phosphorylation-dependent site at Ser218 (potentiated by active H-Ras signaling). The pSer218-dependent interaction inhibits active Gαi1-AlF4- binding to the RGS domain of RGS14 (measured by BRET), while the phosphorylation-independent interaction inhibits RGS14 nuclear import and nucleocytoplasmic shuttling.","method":"Bioluminescence resonance energy transfer (BRET), co-immunoprecipitation, site-directed mutagenesis (Ser218), active H-Ras stimulation, nuclear import assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — BRET (quantitative direct interaction), Co-IP, phospho-site mutagenesis, distinct functional readouts for each interaction site; multiple orthogonal methods","pmids":["30093406"],"is_preprint":false},{"year":2017,"finding":"14-3-3γ directly interacts with bestrophin-1 (Best1) anion channel in astrocytes (identified by yeast two-hybrid; confirmed by BiFC). The interaction is mediated by phosphorylation of Ser358 in the C-terminus of Best1. 14-3-3γ silencing reduces Best1 surface expression and Best1-mediated whole-cell currents, and decreases Best1-mediated glutamate release from hippocampal astrocytes (recorded as reduced NMDA receptor current in CA1 neurons).","method":"Yeast two-hybrid, bimolecular fluorescence complementation (BiFC), whole-cell patch clamp, shRNA knockdown, hippocampal slice electrophysiology","journal":"Molecular brain","confidence":"High","confidence_rationale":"Tier 2 / Strong — yeast two-hybrid + BiFC + electrophysiology + in vivo slice recording; multiple orthogonal methods, clear functional consequence at synapse level","pmids":["29121962"],"is_preprint":false},{"year":2022,"finding":"14-3-3γ binds to TMCC3 (an ER membrane protein at three-way junctions) through phosphorylation of serine residues in deduced 14-3-3 binding motifs in the N-terminus of TMCC3. Overexpression of 14-3-3γ reduces TMCC3 localization to three-way junctions and decreases their number. A TMCC3 alanine-substitution mutant at the phosphorylatable serine shows reduced 14-3-3γ binding and is more resistant to 14-3-3γ-driven mislocalization, demonstrating that 14-3-3γ negatively regulates the reticular ER network by controlling TMCC3 distribution.","method":"Co-immunoprecipitation, site-directed mutagenesis (Ser-to-Ala in binding motif), overexpression, three-way junction quantification by fluorescence microscopy, ER morphology assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP, phospho-site mutagenesis, overexpression with quantified organelle morphology readout; multiple orthogonal methods in one study","pmids":["36549645"],"is_preprint":false},{"year":2017,"finding":"14-3-3γ directly interacts with Copine1 (CPNE1) in hippocampal progenitor cells; among all seven 14-3-3 isoforms, only 14-3-3γ binds CPNE1. The interaction requires the Ser54 residue of the C2A domain of CPNE1; Ser54 mutation reduces 14-3-3γ binding and CPNE1-dependent AKT phosphorylation and neuronal differentiation.","method":"Yeast two-hybrid, co-immunoprecipitation (in vitro and in vivo), Ser54 mutation, isoform selectivity assay, neurite outgrowth and neuronal marker expression","journal":"Experimental cell research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast two-hybrid + Co-IP + phospho-site mutagenesis + neuronal differentiation readout; single lab","pmids":["28412242"],"is_preprint":false},{"year":2012,"finding":"14-3-3γ hypoxia-mediated p53 activation occurs through the ATR-Chk1-MDMX-14-3-3γ pathway: hypoxia induces ATR-Chk1-dependent phosphorylation of MDMX at Ser367, enhancing MDMX binding to 14-3-3γ, which leads to p53 activation. In MEFs with MDMX containing the S367A mutation, hypoxia fails to induce MDMX-14-3-3γ binding or p53 activation.","method":"Co-immunoprecipitation, Chk1/ATR inhibitor and siRNA knockdown, phospho-specific antibody (pSer367-MDMX), MEFs with knock-in MDMX S367A mutant, p53 target gene induction","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP, knock-in phospho-site mutant in primary MEFs, ATR/Chk1 inhibitors, multiple functional readouts; replicates and extends prior UV-damage findings","pmids":["22556425"],"is_preprint":false},{"year":2015,"finding":"Loss of 14-3-3γ in mice leads to delayed neuronal migration and morphological defects (thicker leading process stem, failure to reach cortical plate-marginal zone boundary, increased multipolar neurons) in the developing cerebral cortex as shown by in utero electroporation knockdown and time-lapse live imaging of brain slices.","method":"In utero electroporation (shRNA knockdown), time-lapse live imaging of brain slices, immunofluorescence for cortical layer markers","journal":"Developmental neurobiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in utero loss-of-function with live imaging and morphological quantification; single lab","pmids":["26297819"],"is_preprint":false},{"year":2016,"finding":"Overexpression of 14-3-3γ in utero also causes delayed pyramidal neuron migration in the developing mouse cortex, similar to knockdown, indicating that a precise balance of 14-3-3γ expression is required for proper cortical development.","method":"In utero electroporation (overexpression), cortical layer marker immunofluorescence, neuronal position quantification","journal":"Neuroscience letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in utero gain-of-function with defined cortical migration phenotype; single lab, replicates the loss-of-function findings with the opposite manipulation","pmids":["27288018"],"is_preprint":false},{"year":2020,"finding":"An LC-MS proteomics screen of 14-3-3γ-interacting proteins in MA-10 Leydig cells identified 688 interactors including Sec23ip (a vesicle trafficking protein). Silencing Sec23ip decreased steroidogenesis and impaired cholesterol mobilization from cytoplasmic membrane to mitochondria, placing Sec23ip in the 14-3-3γ-regulated steroidogenic network.","method":"Liquid chromatography–mass spectrometry of immunoprecipitated 14-3-3γ complex, siRNA knockdown of Sec23ip, steroidogenesis assay, cholesterol trafficking assay","journal":"Endocrinology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — LC-MS interactome, siRNA functional validation with steroid output and cholesterol trafficking; single lab","pmids":["31875919"],"is_preprint":false},{"year":2021,"finding":"14-3-3γ knockout cells with supernumerary centrosomes show increased centrosome clustering and pseudo-bipolar mitoses compared to 14-3-3ε knockout cells that show multipolar mitoses. Loss of 14-3-3γ compromises desmosome function and decreases keratin levels, reducing cell stiffness and promoting centrosome clustering. Restoration of desmosome function increased multipolar mitoses; knockdown of plakoglobin or keratin 5 reduced cell stiffness and increased pseudo-bipolar mitoses.","method":"14-3-3γ/ε knockout cell lines, centrosome counting and spindle analysis, desmosome rescue experiments, plakoglobin/keratin 5 knockdown, atomic force microscopy or equivalent cell stiffness assay","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — knockout comparison, desmosome rescue, cytoskeletal knockdown with mechanical and mitotic readouts; single lab","pmids":["34626438"],"is_preprint":false},{"year":2017,"finding":"14-3-3γ directly interacts with CPNE1 and a yeast two-hybrid screen confirmed isoform specificity (only γ among seven isoforms). Also, YWHAG knockdown in breast cancer cells reduces Snail protein stability, EMT markers, and invasion; ectopic YWHAG overexpression abrogates miR-181b-3p-induced Snail stabilization and EMT.","method":"Luciferase reporter assay (miR-181b-3p target validation), siRNA/miRNA transfection, Western blot for Snail stability, invasion and migration assays, in vivo lung metastasis assay in mice","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — miRNA target validation by luciferase, protein stability by siRNA knockdown/overexpression with invasion readout and in vivo metastasis; single lab","pmids":["27102539"],"is_preprint":false},{"year":2023,"finding":"14-3-3γ deficiency causes rapid ROS accumulation and delayed EMT in cancer cells, revealing a YWHAG-dependent cytoprotective mechanism embedded in EMT-associated networks that protects cancer cells from oxidative catastrophe through enhanced autophagy. Tumor allografts show that metastasis potential correlates with YWHAG expression level, and YWHAG silencing diminishes primary tumor volumes and prevents metastasis.","method":"Cellular kinome and transcriptome analysis (regulome construction), ROS assay, EMT markers, autophagy flux assay, 14-3-3γ knockdown, in vivo tumor allograft and metastasis model","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — transcriptome/kinome regulome, ROS + autophagy assays, in vivo metastasis model with YWHAG knockdown; single lab, multiple readouts","pmids":["37759388"],"is_preprint":false},{"year":2024,"finding":"YWHAG promotes bladder cancer cell invasion and metastasis by interacting with and upregulating TMOD3, which then activates ERK1/2 and JNK phosphorylation in the MAPK pathway. The YWHAG-TMOD3 interaction was confirmed by pull-down with mass spectrometry and co-immunoprecipitation. TMOD3 knockdown reversed the pro-metastatic effects of YWHAG.","method":"Pull-down assay with mass spectrometry, co-immunoprecipitation, transcriptome sequencing, TMOD3 knockdown, in vitro invasion/metastasis assays, in vivo metastasis model","journal":"Journal of translational medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pull-down+MS, Co-IP, transcriptome, epistasis (TMOD3 knockdown reverses YWHAG effect); single lab","pmids":["39741303"],"is_preprint":false},{"year":2019,"finding":"14-3-3γ is localized to pseudopodia of MDA-MB-231 breast cancer cells and co-localizes with F-actin there. Knockdown of 14-3-3γ decreases pseudopodial formation/elongation and cell migration; forced expression has the opposite effect.","method":"Excimer laser cell etching to isolate pseudopodial proteins, confocal imaging, siRNA knockdown and overexpression, pseudopodia counting, wound healing and Transwell migration assays","journal":"Breast cancer (Tokyo, Japan)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — subcellular fractionation (pseudopodia isolation), confocal co-localization, gain- and loss-of-function with motility readout; single lab","pmids":["30830684"],"is_preprint":false},{"year":2024,"finding":"YWHAG promotes CRC cell proliferation, migration, and invasion by interacting with cortactin (CTTN), which activates Wnt/β-catenin signaling. CTTN was identified as a YWHAG-associated protein mediating YWHAG's tumor-promoting functions.","method":"Co-immunoprecipitation (CTTN-YWHAG interaction), RNA-seq pathway analysis, CTTN/YWHAG knockdown with proliferation/invasion readouts, Wnt/β-catenin reporter","journal":"Medical oncology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP, RNA-seq pathway analysis, functional knockdown; single lab","pmids":["38538804"],"is_preprint":false},{"year":2023,"finding":"14-3-3γ haploinsufficiency in aged mice leads to decreased dopamine levels and altered dopamine metabolism in the brain, along with changes in phosphorylation of proteins implicated in PD pathology, and PD-like motor coordination deficits.","method":"Heterozygous 14-3-3γ knockout mice, dopamine HPLC quantification, phospho-Western blot of PD-relevant proteins, behavioral testing (rotarod, nest-building)","journal":"Molecular brain","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic loss-of-function model with biochemical (dopamine, phospho-proteins) and behavioral (motor) readouts; single lab","pmids":["36604743"],"is_preprint":false}],"current_model":"14-3-3γ (YWHAG) is a phosphoserine/phosphothreonine-binding dimeric scaffold protein that regulates diverse cellular processes by sequestering or co-localizing phosphorylated client proteins: it binds Chk1-phosphorylated MDMX to retain it in the cytoplasm and activate p53, forms a ternary complex with Chk1 and Cdc25A to trigger Cdc25A degradation and prevent premature mitosis, docks to Plk1-pSer99 to stimulate its kinase activity and drive metaphase-to-anaphase transition, bridges CtBP1-S/BARS with PI4KIIIβ at the Golgi to couple carrier budding and fission, protects p21 from MDMX-dependent proteasomal degradation, sequesters phospho-Bad to prevent apoptosis, binds phospho-GFAP to regulate intermediate filament dynamics, promotes surface expression of ion channels (TRPM4b, ANO1, Best1) by binding phosphorylated N/C-terminal motifs, negatively regulates StAR-dependent steroidogenesis via phospho-Ser194 binding (controlled by its own Ser58 phosphorylation and Lys49 acetylation), transports plakoglobin to cell borders (via PKD/KIF5B) to initiate desmosome assembly, controls the reticular ER network by displacing TMCC3 from three-way junctions, binds tyrosine hydroxylase phosphorylated at pSer19 with nanomolar affinity to stabilize and regulate it, is itself phosphorylated by PAK6 at Ser59 (switching it off LRRK2), by CK2 at Ser235, and by multiple PKC isoforms in response to PDGF; its stability is maintained by the deubiquitinase USP37 and suppressed by p53-driven proteasomal degradation; and its precise expression level is critical for proper neuronal migration in cortical development."},"narrative":{"mechanistic_narrative":"14-3-3γ (YWHAG) is a dimeric phosphoserine/phosphothreonine-binding scaffold that governs the localization, stability, and activity of phosphorylated client proteins across cell-cycle control, membrane trafficking, cytoskeletal dynamics, and survival signaling [PMID:16511572, PMID:24947669, PMID:24610948]. In the DNA-damage and hypoxia response it acts as a node of the ATR–Chk1 axis: it binds Chk1-phosphorylated MDMX (pSer367) to enforce cytoplasmic retention of MDMX and stabilize p53 [PMID:16511572, PMID:22556425], forms a ternary complex with Chk1-pSer296 and Cdc25A to drive Cdc25A degradation and block premature mitosis [PMID:20639859], and protects p21 from MDMX-dependent proteasomal turnover independently of p53 [PMID:21148311]. During mitosis it docks onto Plk1-pSer99 to stimulate Plk1 activity and enable the metaphase-to-anaphase transition [PMID:23695676], and its loss produces NPM1-pThr199–dependent centrosome amplification and aneuploidy [PMID:27253419]. As a phosphorylation-gated scaffold it bridges CtBP1-S/BARS to PI4KIIIβ at the Golgi to couple post-Golgi carrier budding and fission [PMID:22366688], displaces TMCC3 from ER three-way junctions to remodel the reticular ER [PMID:36549645], directs PKD/KIF5B-dependent transport of plakoglobin to initiate desmosome assembly [PMID:24610948], and promotes surface expression of the ion channels TRPM4b, ANO1, and Best1 through binding their phosphorylated N/C-terminal motifs [PMID:27212225, PMID:25047048, PMID:29121962]. It binds phospho-Bad and p-β-catenin(Ser37) to suppress apoptosis [PMID:24957688, PMID:24743739], engages phospho-GFAP to control intermediate-filament integrity [PMID:17032734], and binds tyrosine hydroxylase pSer19 with nanomolar affinity while restricting its PKA-mediated Ser40 phosphorylation [PMID:24947669]. Its own activity is set by post-translational marks—PAK6 phosphorylates Ser59 to release clients including LRRK2-pSer935 [PMID:29311810], CK2 phosphorylates Ser235 [PMID:26198629]—and its abundance is balanced by USP37-mediated deubiquitination and p53-driven degradation [PMID:26427597, PMID:25384678]. In the brain, precise 14-3-3γ dosage is required for cortical neuronal migration, since both knockdown and overexpression delay migration [PMID:26297819, PMID:27288018], and haploinsufficiency lowers dopamine and produces PD-like motor deficits [PMID:36604743]. Through oncogenic PI3K/MAPK activation and interactions with effectors such as TMOD3 and cortactin, it also drives transformation, EMT, and metastasis [PMID:20628654, PMID:39741303, PMID:38538804].","teleology":[{"year":2003,"claim":"Early work asked whether 14-3-3γ couples stress signaling to survival in glia, establishing it as a Raf-associated, ischemia-induced scaffold rather than a passive housekeeping protein.","evidence":"Co-IP of endogenous 14-3-3γ with c-Raf-1/p-Raf and actin in ischemic astrocyte cultures, with inhibitor-resistant induction","pmids":["12730952","12176032"],"confidence":"Medium","gaps":["Direct binding to Raf not demonstrated with purified proteins","Functional consequence of the Raf association not defined"]},{"year":2006,"claim":"Defining 14-3-3γ as a phospho-dependent client binder, it was shown to retain Chk1-phosphorylated MDMX in the cytoplasm to stabilize p53, placing it directly in the DNA-damage checkpoint.","evidence":"Affinity-MS, in vitro binding, reciprocal Co-IP, K50E binding-dead mutant, Chk1-KD/UCN-01 and siRNA, with p53/G1-arrest readouts","pmids":["16511572"],"confidence":"High","gaps":["Whether the same module operates outside UV damage was not yet tested","Structural basis of pSer367 recognition not resolved"]},{"year":2010,"claim":"The checkpoint role was extended to mitotic timing and to p53-independent cell-cycle control, showing 14-3-3γ enforces both Cdc25A degradation and p21 protection.","evidence":"Ternary Co-IP with Chk1-pSer296/Cdc25A plus S296A rescue; in vitro and in-cell Co-IP with MDMX/p21, half-life and G1-arrest assays in p53-null cells","pmids":["20639859","21148311"],"confidence":"High","gaps":["Stoichiometry of the ternary complex not defined","How damage signals partition 14-3-3γ between MDMX, Cdc25A, and p21 clients unknown"]},{"year":2012,"claim":"Beyond the nucleus, 14-3-3γ was established as a phosphorylation-gated membrane-trafficking scaffold and shown to negatively regulate steroidogenesis, revealing a general client-sequestration mechanism.","evidence":"Reconstituted CtBP1-S/BARS–PI4KIIIβ Golgi complex with live-imaging fission readout; MS/Co-IP/Ser194-mutant and siRNA in MA-10 Leydig cells with steroid output","pmids":["22366688","22427666"],"confidence":"High","gaps":["How PKD/PAK phosphorylation switches complex assembly mechanistically not fully resolved","In vivo relevance of Golgi scaffolding not tested"]},{"year":2013,"claim":"14-3-3γ was shown to be an activator, not merely a tether, by docking Plk1-pSer99 to stimulate its kinase activity and license anaphase onset.","evidence":"Co-IP, Plk1 S99A mutant, PI3K/Akt inhibitors, in vitro kinase assay and SAC-dependent mitotic-arrest analysis after siRNA","pmids":["23695676"],"confidence":"High","gaps":["Structural mechanism by which docking increases catalytic activity unknown","Whether other mitotic kinases are similarly activated not addressed"]},{"year":2014,"claim":"Biophysical and channel studies defined isoform-specific, high-affinity, phospho-dependent binding modes—nanomolar TH-pSer19 capture and membrane association via γ-specific histidines—and a recurring role in trafficking phosphorylated ion channels to the surface.","evidence":"Native MS/SPR/EM with TH; SPR with His158/His195 mutants and MD; GST pull-down/Co-IP/patch-clamp/biotinylation for TRPM4b with shRNA rescue","pmids":["24947669","23189152","25047048"],"confidence":"High","gaps":["Whether membrane recruitment is required for channel trafficking in vivo not tested","Generality of His-mediated membrane binding across clients unknown"]},{"year":2014,"claim":"14-3-3γ was placed at the start of desmosome assembly, coupling kinase signaling to motor-driven transport of an adhesion protein.","evidence":"Co-IP, PKCμ inhibitor, microtubule disruption, KIF5B knockdown, and in vitro/in vivo (mouse testis) loss-of-function with spermatogenesis defects","pmids":["24610948"],"confidence":"High","gaps":["Direct phospho-site on plakoglobin recognized by 14-3-3γ not mapped","Whether cargo handoff to the KIF5B motor is direct unknown"]},{"year":2015,"claim":"The protein's abundance was shown to be actively controlled, identifying opposing USP37-mediated stabilization and p53-driven degradation as a homeostatic loop nested within the same checkpoint it regulates.","evidence":"Co-IP, ubiquitination assays, USP37 catalytic mutant, MG132, and WT vs R175H p53 with half-life analysis","pmids":["26427597","25384678"],"confidence":"Medium","gaps":["E3 ligase driving p53-dependent 14-3-3γ ubiquitination not identified","Physiological setting where each arm dominates undefined"]},{"year":2016,"claim":"In vivo and dosage experiments established that 14-3-3γ levels must be tightly balanced for cortical neuronal migration, indicating a developmental requirement beyond its biochemical interactions.","evidence":"In utero electroporation knockdown and overexpression with time-lapse imaging and cortical-layer marker analysis","pmids":["26297819","27288018"],"confidence":"Medium","gaps":["Migration-relevant client(s) not identified","Cell-autonomous vs non-autonomous contribution not separated"]},{"year":2017,"claim":"Phospho-regulation of 14-3-3γ itself was defined as a client-release switch, linking PAK6 phosphorylation at Ser59 to LRRK2 dephosphorylation and neurite rescue.","evidence":"PAK6 interactome Co-IP, in vitro kinase assay with Ser59 mapping, LRRK2-pSer935 blots, and constitutively active PAK6 rescue of G2019S neurite shortening","pmids":["29311810"],"confidence":"High","gaps":["Full set of clients released by Ser59 phosphorylation unknown","Whether CK2-pSer235 has an analogous regulatory effect not tested"]},{"year":2022,"claim":"Expanding its organelle-shaping role, 14-3-3γ was shown to negatively regulate the reticular ER network by sequestering phosphorylated TMCC3 away from three-way junctions.","evidence":"Co-IP, Ser-to-Ala TMCC3 mutant, overexpression, and quantified ER three-way-junction morphology","pmids":["36549645"],"confidence":"High","gaps":["Kinase that phosphorylates TMCC3 binding motif not identified","Physiological trigger for ER remodeling by 14-3-3γ unknown"]},{"year":2024,"claim":"A body of cancer studies established 14-3-3γ as a pro-metastatic effector acting through PI3K/MAPK and adhesion/cytoskeletal partners, tying its scaffolding biochemistry to invasion phenotypes.","evidence":"Pull-down/MS and Co-IP identifying TMOD3 and cortactin, RNA-seq pathway analysis, knockdown epistasis, and in vivo metastasis models","pmids":["39741303","38538804","37759388"],"confidence":"Medium","gaps":["Phospho-dependence of TMOD3/cortactin binding not established","Direct vs indirect activation of Wnt and MAPK pathways unresolved"]},{"year":null,"claim":"It remains unresolved how a single dimeric scaffold prioritizes among its many phosphorylated clients in a given cell, and what defines γ-isoform-specific functions in vivo.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of competitive client selection","Isoform-specific in vivo client repertoire not mapped","Quantitative rules linking 14-3-3γ dosage to migration/disease phenotypes undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,3,19,24,35]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[2,6,4,7]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[0,13,27]},{"term_id":"GO:0005794","term_label":"Golgi apparatus","supporting_discovery_ids":[3,26]},{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[37]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[9,28,33]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[20]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[25]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[12,14,47]}],"pathway":[{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[1,2,20]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[0,39]},{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[3,19]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[13,27,28]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[7,22,35]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[40,41]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[44,45,46,48,49]}],"complexes":[],"partners":["MDMX","CHEK1","CDC25A","PLK1","STAR","TH","LRRK2","RGS14"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P61981","full_name":"14-3-3 protein gamma","aliases":["Protein kinase C inhibitor protein 1","KCIP-1"],"length_aa":247,"mass_kda":28.3,"function":"Adapter protein implicated in the regulation of a large spectrum of both general and specialized signaling pathways (PubMed:15696159, PubMed:16511572, PubMed:36732624). Binds to a large number of partners, usually by recognition of a phosphoserine or phosphothreonine motif (PubMed:15696159, PubMed:16511572, PubMed:36732624). Binding generally results in the modulation of the activity of the binding partner (PubMed:16511572). Promotes inactivation of WDR24 component of the GATOR2 complex by binding to phosphorylated WDR24 (PubMed:36732624). Participates in the positive regulation of NMDA glutamate receptor activity by promoting the L-glutamate secretion through interaction with BEST1 (PubMed:29121962). Reduces keratinocyte intercellular adhesion, via interacting with PKP1 and sequestering it in the cytoplasm, thereby reducing its incorporation into desmosomes (PubMed:29678907). Plays a role in mitochondrial protein catabolic process (also named MALM) that promotes the degradation of damaged proteins inside mitochondria (PubMed:22532927)","subcellular_location":"Cytoplasm, cytosol; Mitochondrion matrix","url":"https://www.uniprot.org/uniprotkb/P61981/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/YWHAG","classification":"Not Classified","n_dependent_lines":82,"n_total_lines":1208,"dependency_fraction":0.06788079470198675},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000170027","cell_line_id":"CID000464","localizations":[{"compartment":"centrosome","grade":3},{"compartment":"cytoplasmic","grade":3},{"compartment":"nucleoplasm","grade":2},{"compartment":"cell_contact","grade":1},{"compartment":"membrane","grade":1}],"interactors":[{"gene":"ARL6IP5","stoichiometry":10.0},{"gene":"MIS12","stoichiometry":10.0},{"gene":"YWHAZ","stoichiometry":10.0},{"gene":"YWHAQ","stoichiometry":10.0},{"gene":"YWHAB","stoichiometry":10.0},{"gene":"AKT1S1","stoichiometry":4.0},{"gene":"ARL8B","stoichiometry":4.0},{"gene":"KRAS","stoichiometry":4.0},{"gene":"KRT18","stoichiometry":4.0},{"gene":"YWHAH","stoichiometry":4.0}],"url":"https://opencell.sf.czbiohub.org/target/CID000464","total_profiled":1310},"omim":[{"mim_id":"617665","title":"DEVELOPMENTAL AND EPILEPTIC ENCEPHALOPATHY 56; DEE56","url":"https://www.omim.org/entry/617665"},{"mim_id":"617105","title":"DEVELOPMENTAL AND EPILEPTIC ENCEPHALOPATHY 41; DEE41","url":"https://www.omim.org/entry/617105"},{"mim_id":"616955","title":"RRAD- AND GEM-LIKE GTPase 2; REM2","url":"https://www.omim.org/entry/616955"},{"mim_id":"613729","title":"CHROMOSOME 7q11.23 DELETION SYNDROME, DISTAL, 1.2-MB","url":"https://www.omim.org/entry/613729"},{"mim_id":"609279","title":"CENTROMERIC PROTEIN J; CENPJ","url":"https://www.omim.org/entry/609279"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Cytosol","reliability":"Supported"},{"location":"Vesicles","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"brain","ntpm":355.0},{"tissue":"skeletal muscle","ntpm":327.6}],"url":"https://www.proteinatlas.org/search/YWHAG"},"hgnc":{"alias_symbol":["PPP1R170","14-3-3GAMMA","14-3-3γ"],"prev_symbol":[]},"alphafold":{"accession":"P61981","domains":[{"cath_id":"1.20.190.20","chopping":"1-111","consensus_level":"medium","plddt":95.9095,"start":1,"end":111},{"cath_id":"1.20.190.20","chopping":"150-239","consensus_level":"medium","plddt":95.4969,"start":150,"end":239}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P61981","model_url":"https://alphafold.ebi.ac.uk/files/AF-P61981-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P61981-F1-predicted_aligned_error_v6.png","plddt_mean":94.19},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=YWHAG","jax_strain_url":"https://www.jax.org/strain/search?query=YWHAG"},"sequence":{"accession":"P61981","fasta_url":"https://rest.uniprot.org/uniprotkb/P61981.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P61981/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P61981"}},"corpus_meta":[{"pmid":"30063064","id":"PMC_30063064","title":"Curcumin attenuates doxorubicin-induced cardiotoxicity via suppressing oxidative stress and preventing mitochondrial dysfunction mediated by 14-3-3γ.","date":"2018","source":"Food & function","url":"https://pubmed.ncbi.nlm.nih.gov/30063064","citation_count":103,"is_preprint":false},{"pmid":"16511572","id":"PMC_16511572","title":"14-3-3gamma binds to MDMX that is phosphorylated by UV-activated Chk1, resulting in p53 activation.","date":"2006","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/16511572","citation_count":100,"is_preprint":false},{"pmid":"12694396","id":"PMC_12694396","title":"A dual role for poly-ADP-ribosylation in spatial memory acquisition after traumatic brain injury in mice involving NAD+ depletion and ribosylation of 14-3-3gamma.","date":"2003","source":"Journal of neurochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/12694396","citation_count":87,"is_preprint":false},{"pmid":"20639859","id":"PMC_20639859","title":"14-3-3gamma mediates Cdc25A proteolysis to block premature mitotic entry after DNA damage.","date":"2010","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/20639859","citation_count":74,"is_preprint":false},{"pmid":"22366688","id":"PMC_22366688","title":"A 14-3-3γ dimer-based scaffold bridges CtBP1-S/BARS to PI(4)KIIIβ to regulate post-Golgi carrier formation.","date":"2012","source":"Nature cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/22366688","citation_count":72,"is_preprint":false},{"pmid":"17109079","id":"PMC_17109079","title":"Small heat shock protein Hsp20 (HspB6) as a partner of 14-3-3gamma.","date":"2006","source":"Molecular and cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/17109079","citation_count":68,"is_preprint":false},{"pmid":"27102539","id":"PMC_27102539","title":"miR-181b-3p promotes epithelial-mesenchymal transition in breast cancer cells through Snail stabilization by directly targeting YWHAG.","date":"2016","source":"Biochimica et biophysica acta","url":"https://pubmed.ncbi.nlm.nih.gov/27102539","citation_count":63,"is_preprint":false},{"pmid":"23695676","id":"PMC_23695676","title":"PI 3-kinase-dependent phosphorylation of Plk1-Ser99 promotes association with 14-3-3γ and is required for metaphase-anaphase transition.","date":"2013","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/23695676","citation_count":60,"is_preprint":false},{"pmid":"15684385","id":"PMC_15684385","title":"Unchanged survival rates of 14-3-3gamma knockout mice after inoculation with pathological prion protein.","date":"2005","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/15684385","citation_count":59,"is_preprint":false},{"pmid":"28777935","id":"PMC_28777935","title":"De Novo Mutations in YWHAG Cause Early-Onset Epilepsy.","date":"2017","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/28777935","citation_count":59,"is_preprint":false},{"pmid":"16223726","id":"PMC_16223726","title":"Role of 14-3-3gamma in FE65-dependent gene transactivation mediated by the amyloid beta-protein precursor cytoplasmic fragment.","date":"2005","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/16223726","citation_count":57,"is_preprint":false},{"pmid":"30636491","id":"PMC_30636491","title":"Quercetin protects cardiomyocytes against doxorubicin-induced toxicity by suppressing oxidative stress and improving mitochondrial function via 14-3-3γ.","date":"2019","source":"Toxicology mechanisms and methods","url":"https://pubmed.ncbi.nlm.nih.gov/30636491","citation_count":57,"is_preprint":false},{"pmid":"21109226","id":"PMC_21109226","title":"Recurrent distal 7q11.23 deletion including HIP1 and YWHAG identified in patients with intellectual disabilities, epilepsy, and neurobehavioral problems.","date":"2010","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/21109226","citation_count":56,"is_preprint":false},{"pmid":"12730952","id":"PMC_12730952","title":"14-3-3gamma is upregulated by in vitro ischemia and binds to protein kinase Raf in primary cultures of astrocytes.","date":"2003","source":"Glia","url":"https://pubmed.ncbi.nlm.nih.gov/12730952","citation_count":47,"is_preprint":false},{"pmid":"29311810","id":"PMC_29311810","title":"PAK6 Phosphorylates 14-3-3γ to Regulate Steady State Phosphorylation of LRRK2.","date":"2017","source":"Frontiers in molecular neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/29311810","citation_count":46,"is_preprint":false},{"pmid":"27212225","id":"PMC_27212225","title":"Suppression of 14-3-3γ-mediated surface expression of ANO1 inhibits cancer progression of glioblastoma cells.","date":"2016","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/27212225","citation_count":45,"is_preprint":false},{"pmid":"17032734","id":"PMC_17032734","title":"14-3-3gamma affects dynamics and integrity of glial filaments by binding to phosphorylated GFAP.","date":"2006","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/17032734","citation_count":43,"is_preprint":false},{"pmid":"15660102","id":"PMC_15660102","title":"Association of 14-3-3gamma and phosphorylated bad attenuates injury in ischemic astrocytes.","date":"2005","source":"Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/15660102","citation_count":42,"is_preprint":false},{"pmid":"20628654","id":"PMC_20628654","title":"14-3-3gamma induces oncogenic transformation by stimulating MAP kinase and PI3K signaling.","date":"2010","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/20628654","citation_count":42,"is_preprint":false},{"pmid":"27894843","id":"PMC_27894843","title":"MiR-509-5p suppresses the proliferation, migration, and invasion of non-small cell lung cancer by targeting YWHAG.","date":"2016","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/27894843","citation_count":39,"is_preprint":false},{"pmid":"19393026","id":"PMC_19393026","title":"Ischemia activates JNK/c-Jun/AP-1 pathway to up-regulate 14-3-3gamma in astrocyte.","date":"2009","source":"Journal of neurochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/19393026","citation_count":39,"is_preprint":false},{"pmid":"17394238","id":"PMC_17394238","title":"Overexpression of 14-3-3gamma causes polyploidization in H322 lung cancer cells.","date":"2007","source":"Molecular carcinogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/17394238","citation_count":38,"is_preprint":false},{"pmid":"19574997","id":"PMC_19574997","title":"Silencing neuroglobin enhances neuronal vulnerability to oxidative injury by down-regulating 14-3-3gamma.","date":"2009","source":"Acta pharmacologica Sinica","url":"https://pubmed.ncbi.nlm.nih.gov/19574997","citation_count":38,"is_preprint":false},{"pmid":"34168120","id":"PMC_34168120","title":"LncRNA CERS6-AS1 promotes proliferation and metastasis through the upregulation of YWHAG and activation of ERK signaling in pancreatic cancer.","date":"2021","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/34168120","citation_count":38,"is_preprint":false},{"pmid":"24957688","id":"PMC_24957688","title":"14-3-3γ protein attenuates lipopolysaccharide-induced cardiomyocytes injury through the Bcl-2 family/mitochondria pathway.","date":"2014","source":"International immunopharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/24957688","citation_count":36,"is_preprint":false},{"pmid":"28745316","id":"PMC_28745316","title":"Elevation of adenylate energy charge by angiopoietin-like 4 enhances epithelial-mesenchymal transition by inducing 14-3-3γ expression.","date":"2017","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/28745316","citation_count":36,"is_preprint":false},{"pmid":"33986684","id":"PMC_33986684","title":"Capsaicin protects cardiomyocytes against lipopolysaccharide-induced damage via 14-3-3γ-mediated autophagy augmentation.","date":"2021","source":"Frontiers in pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/33986684","citation_count":35,"is_preprint":false},{"pmid":"22427666","id":"PMC_22427666","title":"Hormone-induced 14-3-3γ adaptor protein regulates steroidogenic acute regulatory protein activity and steroid biosynthesis in MA-10 Leydig cells.","date":"2012","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/22427666","citation_count":35,"is_preprint":false},{"pmid":"12176032","id":"PMC_12176032","title":"The association of 14-3-3gamma and actin plays a role in cell division and apoptosis in astrocytes.","date":"2002","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/12176032","citation_count":34,"is_preprint":false},{"pmid":"32958119","id":"PMC_32958119","title":"Emerging roles of 14-3-3γ in the brain disorder.","date":"2020","source":"BMB reports","url":"https://pubmed.ncbi.nlm.nih.gov/32958119","citation_count":33,"is_preprint":false},{"pmid":"36076529","id":"PMC_36076529","title":"Puerarin activates adaptive autophagy and protects the myocardium against doxorubicin-induced cardiotoxicity via the 14-3-3γ/PKCε pathway.","date":"2022","source":"Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie","url":"https://pubmed.ncbi.nlm.nih.gov/36076529","citation_count":33,"is_preprint":false},{"pmid":"20146355","id":"PMC_20146355","title":"Zebrafish gene knockdowns imply roles for human YWHAG in infantile spasms and cardiomegaly.","date":"2010","source":"Genesis (New York, N.Y. : 2000)","url":"https://pubmed.ncbi.nlm.nih.gov/20146355","citation_count":33,"is_preprint":false},{"pmid":"34709972","id":"PMC_34709972","title":"Long non-coding RNA NORAD protects against cerebral ischemia/reperfusion injury induced brain damage, cell apoptosis, oxidative stress and inflammation by regulating miR-30a-5p/YWHAG.","date":"2021","source":"Bioengineered","url":"https://pubmed.ncbi.nlm.nih.gov/34709972","citation_count":32,"is_preprint":false},{"pmid":"28126486","id":"PMC_28126486","title":"MiR-217 promoted the proliferation and invasion of glioblastoma by repressing YWHAG.","date":"2017","source":"Cytokine","url":"https://pubmed.ncbi.nlm.nih.gov/28126486","citation_count":31,"is_preprint":false},{"pmid":"29782860","id":"PMC_29782860","title":"Tetramethylpyrazine attenuates lipopolysaccharide-induced cardiomyocyte injury via improving mitochondrial function mediated by 14-3-3γ.","date":"2018","source":"European journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/29782860","citation_count":31,"is_preprint":false},{"pmid":"10433554","id":"PMC_10433554","title":"14-3-3Gamma interacts with and is phosphorylated by multiple protein kinase C isoforms in PDGF-stimulated human vascular smooth muscle cells.","date":"1999","source":"DNA and cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/10433554","citation_count":31,"is_preprint":false},{"pmid":"30478609","id":"PMC_30478609","title":"Preconditioning exercise reduces brain damage and neuronal apoptosis through enhanced endogenous 14-3-3γ after focal brain ischemia in rats.","date":"2018","source":"Brain structure & function","url":"https://pubmed.ncbi.nlm.nih.gov/30478609","citation_count":31,"is_preprint":false},{"pmid":"27253419","id":"PMC_27253419","title":"14-3-3γ Prevents Centrosome Amplification and Neoplastic Progression.","date":"2016","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/27253419","citation_count":29,"is_preprint":false},{"pmid":"25047048","id":"PMC_25047048","title":"Depletion of 14-3-3γ reduces the surface expression of Transient Receptor Potential Melastatin 4b (TRPM4b) channels and attenuates TRPM4b-mediated glutamate-induced neuronal cell death.","date":"2014","source":"Molecular brain","url":"https://pubmed.ncbi.nlm.nih.gov/25047048","citation_count":29,"is_preprint":false},{"pmid":"20467836","id":"PMC_20467836","title":"14-3-3gamma and neuroglobin are new intrinsic protective factors for cerebral ischemia.","date":"2010","source":"Molecular neurobiology","url":"https://pubmed.ncbi.nlm.nih.gov/20467836","citation_count":28,"is_preprint":false},{"pmid":"31885804","id":"PMC_31885804","title":"Tetramethylpyrazine Attenuates the Endotheliotoxicity and the Mitochondrial Dysfunction by Doxorubicin via 14-3-3γ/Bcl-2.","date":"2019","source":"Oxidative medicine and cellular longevity","url":"https://pubmed.ncbi.nlm.nih.gov/31885804","citation_count":27,"is_preprint":false},{"pmid":"24743739","id":"PMC_24743739","title":"Selective 14-3-3γ induction quenches p-β-catenin Ser37/Bax-enhanced cell death in cerebral cortical neurons during ischemia.","date":"2014","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/24743739","citation_count":27,"is_preprint":false},{"pmid":"26204835","id":"PMC_26204835","title":"14-3-3γ Regulates Lipopolysaccharide-Induced Inflammatory Responses and Lactation in Dairy Cow Mammary Epithelial Cells by Inhibiting NF-κB and MAPKs and Up-Regulating mTOR Signaling.","date":"2015","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/26204835","citation_count":27,"is_preprint":false},{"pmid":"10486217","id":"PMC_10486217","title":"Cloning, expression, and chromosomal mapping of the human 14-3-3gamma gene (YWHAG) to 7q11.23.","date":"1999","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/10486217","citation_count":26,"is_preprint":false},{"pmid":"26427597","id":"PMC_26427597","title":"Deubiquitinating enzyme USP37 regulating oncogenic function of 14-3-3γ.","date":"2015","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/26427597","citation_count":26,"is_preprint":false},{"pmid":"26297819","id":"PMC_26297819","title":"Ablation of the 14-3-3gamma Protein Results in Neuronal Migration Delay and Morphological Defects in the Developing Cerebral Cortex.","date":"2015","source":"Developmental neurobiology","url":"https://pubmed.ncbi.nlm.nih.gov/26297819","citation_count":26,"is_preprint":false},{"pmid":"20870266","id":"PMC_20870266","title":"Involvement of 14-3-3γ overexpression in extrahepatic metastasis of hepatocellular carcinoma.","date":"2010","source":"Human pathology","url":"https://pubmed.ncbi.nlm.nih.gov/20870266","citation_count":26,"is_preprint":false},{"pmid":"35342350","id":"PMC_35342350","title":"miR-200c suppression increases tau hyperphosphorylation by targeting 14-3-3γ in early stage of 5xFAD mouse model of Alzheimer's disease.","date":"2022","source":"International journal of biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/35342350","citation_count":25,"is_preprint":false},{"pmid":"33767733","id":"PMC_33767733","title":"YWHAG Mutations Cause Childhood Myoclonic Epilepsy and Febrile Seizures: Molecular Sub-regional Effect and Mechanism.","date":"2021","source":"Frontiers in genetics","url":"https://pubmed.ncbi.nlm.nih.gov/33767733","citation_count":24,"is_preprint":false},{"pmid":"25281768","id":"PMC_25281768","title":"Molecular network including eIF1AX, RPS7, and 14-3-3γ regulates protein translation and cell proliferation in bovine mammary epithelial cells.","date":"2014","source":"Archives of biochemistry and biophysics","url":"https://pubmed.ncbi.nlm.nih.gov/25281768","citation_count":23,"is_preprint":false},{"pmid":"26940479","id":"PMC_26940479","title":"Sporadic Creutzfeldt-Jakob disease diagnostic accuracy is improved by a new CSF ELISA 14-3-3γ assay.","date":"2016","source":"Neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/26940479","citation_count":22,"is_preprint":false},{"pmid":"22556425","id":"PMC_22556425","title":"Hypoxia activates tumor suppressor p53 by inducing ATR-Chk1 kinase cascade-mediated phosphorylation and consequent 14-3-3γ inactivation of MDMX protein.","date":"2012","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/22556425","citation_count":21,"is_preprint":false},{"pmid":"37759388","id":"PMC_37759388","title":"YWHAG Deficiency Disrupts the EMT-Associated Network to Induce Oxidative Cell Death and Prevent Metastasis.","date":"2023","source":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/37759388","citation_count":20,"is_preprint":false},{"pmid":"32535685","id":"PMC_32535685","title":"Functional genetic variants in centrosome-related genes CEP72 and YWHAG confer susceptibility to gastric cancer.","date":"2020","source":"Archives of toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/32535685","citation_count":20,"is_preprint":false},{"pmid":"24947669","id":"PMC_24947669","title":"Phosphorylation dependence and stoichiometry of the complex formed by tyrosine hydroxylase and 14-3-3γ.","date":"2014","source":"Molecular & cellular proteomics : MCP","url":"https://pubmed.ncbi.nlm.nih.gov/24947669","citation_count":20,"is_preprint":false},{"pmid":"25086053","id":"PMC_25086053","title":"Protein modifications regulate the role of 14-3-3γ adaptor protein in cAMP-induced steroidogenesis in MA-10 Leydig cells.","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/25086053","citation_count":20,"is_preprint":false},{"pmid":"26459638","id":"PMC_26459638","title":"Mitotic Golgi translocation of ERK1c is mediated by a PI4KIIIβ-14-3-3γ shuttling complex.","date":"2015","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/26459638","citation_count":19,"is_preprint":false},{"pmid":"24610948","id":"PMC_24610948","title":"14-3-3γ-Mediated transport of plakoglobin to the cell border is required for the initiation of desmosome assembly in vitro and in vivo.","date":"2014","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/24610948","citation_count":19,"is_preprint":false},{"pmid":"27437776","id":"PMC_27437776","title":"YAP and 14-3-3γ are involved in HS-OA-induced growth inhibition of hepatocellular carcinoma cells: A novel mechanism for hydrogen sulfide releasing oleanolic acid.","date":"2016","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/27437776","citation_count":18,"is_preprint":false},{"pmid":"21148311","id":"PMC_21148311","title":"14-3-3Gamma inhibition of MDMX-mediated p21 turnover independent of p53.","date":"2010","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/21148311","citation_count":17,"is_preprint":false},{"pmid":"23189152","id":"PMC_23189152","title":"The peripheral binding of 14-3-3γ to membranes involves isoform-specific histidine residues.","date":"2012","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/23189152","citation_count":17,"is_preprint":false},{"pmid":"30830684","id":"PMC_30830684","title":"Breast cancer cell motility is promoted by 14-3-3γ.","date":"2019","source":"Breast cancer (Tokyo, Japan)","url":"https://pubmed.ncbi.nlm.nih.gov/30830684","citation_count":16,"is_preprint":false},{"pmid":"27288018","id":"PMC_27288018","title":"Overexpression of the 14-3-3gamma protein in embryonic mice results in neuronal migration delay in the developing cerebral cortex.","date":"2016","source":"Neuroscience letters","url":"https://pubmed.ncbi.nlm.nih.gov/27288018","citation_count":16,"is_preprint":false},{"pmid":"26198629","id":"PMC_26198629","title":"Protein kinase CK2 interacts at the neuromuscular synapse with Rapsyn, Rac1, 14-3-3γ, and Dok-7 proteins and phosphorylates the latter two.","date":"2015","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/26198629","citation_count":16,"is_preprint":false},{"pmid":"30093406","id":"PMC_30093406","title":"14-3-3γ binds regulator of G protein signaling 14 (RGS14) at distinct sites to inhibit the RGS14:Gαi-AlF4- signaling complex and RGS14 nuclear localization.","date":"2018","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/30093406","citation_count":15,"is_preprint":false},{"pmid":"15010287","id":"PMC_15010287","title":"Inducible expression of the signal transduction protein 14-3-3gamma in injured arteries and stimulated human vascular smooth muscle cells.","date":"2004","source":"Experimental and molecular pathology","url":"https://pubmed.ncbi.nlm.nih.gov/15010287","citation_count":15,"is_preprint":false},{"pmid":"35795276","id":"PMC_35795276","title":"Curcumenol Targeting YWHAG Inhibits the Pentose Phosphate Pathway and Enhances Antitumor Effects of Cisplatin.","date":"2022","source":"Evidence-based complementary and alternative medicine : eCAM","url":"https://pubmed.ncbi.nlm.nih.gov/35795276","citation_count":14,"is_preprint":false},{"pmid":"29121962","id":"PMC_29121962","title":"Direct interaction with 14-3-3γ promotes surface expression of Best1 channel in astrocyte.","date":"2017","source":"Molecular brain","url":"https://pubmed.ncbi.nlm.nih.gov/29121962","citation_count":14,"is_preprint":false},{"pmid":"29848704","id":"PMC_29848704","title":"Knockdown of 14-3-3γ Suppresses Epithelial-Mesenchymal Transition and Reduces Metastatic Potential of Human Non-small Cell Lung Cancer Cells.","date":"2018","source":"Anticancer research","url":"https://pubmed.ncbi.nlm.nih.gov/29848704","citation_count":14,"is_preprint":false},{"pmid":"23851690","id":"PMC_23851690","title":"Induction of activation-induced cytidine deaminase-targeting adaptor 14-3-3γ is mediated by NF-κB-dependent recruitment of CFP1 to the 5'-CpG-3'-rich 14-3-3γ promoter and is sustained by E2A.","date":"2013","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/23851690","citation_count":14,"is_preprint":false},{"pmid":"23500129","id":"PMC_23500129","title":"Quantitative proteome analysis of overexpressed Cripto-1 tumor cell reveals 14-3-3γ as a novel biomarker in nasopharyngeal carcinoma.","date":"2013","source":"Journal of proteomics","url":"https://pubmed.ncbi.nlm.nih.gov/23500129","citation_count":14,"is_preprint":false},{"pmid":"27073437","id":"PMC_27073437","title":"14-3-3γ regulates cell viability and milk fat synthesis in lipopolysaccharide-induced dairy cow mammary epithelial cells.","date":"2016","source":"Experimental and therapeutic medicine","url":"https://pubmed.ncbi.nlm.nih.gov/27073437","citation_count":13,"is_preprint":false},{"pmid":"28412242","id":"PMC_28412242","title":"14-3-3γ regulates Copine1-mediated neuronal differentiation in HiB5 hippocampal progenitor cells.","date":"2017","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/28412242","citation_count":12,"is_preprint":false},{"pmid":"26183263","id":"PMC_26183263","title":"14-3-3γ affects mTOR pathway and regulates lactogenesis in dairy cow mammary epithelial cells.","date":"2015","source":"In vitro cellular & developmental biology. Animal","url":"https://pubmed.ncbi.nlm.nih.gov/26183263","citation_count":12,"is_preprint":false},{"pmid":"35729836","id":"PMC_35729836","title":"Puerarin attenuates lipopolysaccharide-induced myocardial injury via the 14-3-3γ/PKCε pathway activating adaptive autophagy.","date":"2022","source":"International immunopharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/35729836","citation_count":11,"is_preprint":false},{"pmid":"22532927","id":"PMC_22532927","title":"Identification of 14-3-3γ as a Mieap-interacting protein and its role in mitochondrial quality control.","date":"2012","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/22532927","citation_count":11,"is_preprint":false},{"pmid":"25711139","id":"PMC_25711139","title":"Ischemia preconditioning protects astrocytes from ischemic injury through 14-3-3γ.","date":"2015","source":"Journal of neuroscience research","url":"https://pubmed.ncbi.nlm.nih.gov/25711139","citation_count":11,"is_preprint":false},{"pmid":"31721359","id":"PMC_31721359","title":"A histone deacetylase 7-derived peptide promotes vascular regeneration via facilitating 14-3-3γ phosphorylation.","date":"2020","source":"Stem cells (Dayton, Ohio)","url":"https://pubmed.ncbi.nlm.nih.gov/31721359","citation_count":11,"is_preprint":false},{"pmid":"30635843","id":"PMC_30635843","title":"Selective 14-3-3γ Upregulation Promotes Beclin-1-LC3-Autophagic Influx via β-Catenin Interaction in Starved Neurons In Vitro and In Vivo.","date":"2019","source":"Neurochemical research","url":"https://pubmed.ncbi.nlm.nih.gov/30635843","citation_count":11,"is_preprint":false},{"pmid":"23115241","id":"PMC_23115241","title":"Activation of phosphatidylinositol 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) signaling and the consequent induction of transformation by overexpressed 14-3-3γ protein require specific amino acids within 14-3-3γ N-terminal variable region II.","date":"2012","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/23115241","citation_count":11,"is_preprint":false},{"pmid":"29771442","id":"PMC_29771442","title":"MicroRNA-222 contributed to cell proliferation, invasion and migration via regulating YWHAG in osteosarcoma.","date":"2018","source":"European review for medical and pharmacological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/29771442","citation_count":10,"is_preprint":false},{"pmid":"32190292","id":"PMC_32190292","title":"miR-140-5p mediates bevacizumab-induced cytotoxicity to cardiomyocytes by targeting the VEGFA/14-3-3γ signal pathway.","date":"2019","source":"Toxicology research","url":"https://pubmed.ncbi.nlm.nih.gov/32190292","citation_count":10,"is_preprint":false},{"pmid":"36604743","id":"PMC_36604743","title":"14-3-3γ haploinsufficiency leads to altered dopamine pathway and Parkinson's disease-like motor incoordination in mice.","date":"2023","source":"Molecular brain","url":"https://pubmed.ncbi.nlm.nih.gov/36604743","citation_count":8,"is_preprint":false},{"pmid":"33393734","id":"PMC_33393734","title":"Epilepsy and electroencephalogram evolution in YWHAG gene mutation: A new phenotype and review of the literature.","date":"2021","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/33393734","citation_count":8,"is_preprint":false},{"pmid":"25384678","id":"PMC_25384678","title":"p53 suppresses 14-3-3γ by stimulating proteasome-mediated 14-3-3γ protein degradation.","date":"2014","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/25384678","citation_count":8,"is_preprint":false},{"pmid":"39741303","id":"PMC_39741303","title":"YWHAG promotes bladder cancer metastasis by regulating TMOD3 to activate ERK1/2 and JNK phosphorylation in the MAPK pathway.","date":"2024","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/39741303","citation_count":7,"is_preprint":false},{"pmid":"30570871","id":"PMC_30570871","title":"MicroRNA-182 promoted esophageal squamous cell carcinoma cell growth and metastasis via targeting YWHAG.","date":"2018","source":"Journal of B.U.ON. : official journal of the Balkan Union of Oncology","url":"https://pubmed.ncbi.nlm.nih.gov/30570871","citation_count":7,"is_preprint":false},{"pmid":"29382566","id":"PMC_29382566","title":"Overexpression of the 14-3-3γ protein in uterine leiomyoma cells results in growth retardation and increased apoptosis.","date":"2018","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/29382566","citation_count":7,"is_preprint":false},{"pmid":"31875919","id":"PMC_31875919","title":"Identification of Sec23ip, Part of 14-3-3γ Protein Network, as a Regulator of Acute Steroidogenesis in MA-10 Leydig Cells.","date":"2020","source":"Endocrinology","url":"https://pubmed.ncbi.nlm.nih.gov/31875919","citation_count":7,"is_preprint":false},{"pmid":"22856523","id":"PMC_22856523","title":"Superhelical DNA as a preferential binding target of 14-3-3γ protein.","date":"2012","source":"Journal of biomolecular structure & dynamics","url":"https://pubmed.ncbi.nlm.nih.gov/22856523","citation_count":7,"is_preprint":false},{"pmid":"38538804","id":"PMC_38538804","title":"YWHAG promotes colorectal cancer progression by regulating the CTTN-Wnt/β-catenin signaling axis.","date":"2024","source":"Medical oncology (Northwood, London, England)","url":"https://pubmed.ncbi.nlm.nih.gov/38538804","citation_count":6,"is_preprint":false},{"pmid":"38286872","id":"PMC_38286872","title":"Serum level of YWHAG as a diagnostic marker of cognitive impairment in Parkinson's disease patients.","date":"2024","source":"Acta neurologica Belgica","url":"https://pubmed.ncbi.nlm.nih.gov/38286872","citation_count":6,"is_preprint":false},{"pmid":"35485284","id":"PMC_35485284","title":"TXNDC9 knockdown inhibits lung adenocarcinoma progression by targeting YWHAG.","date":"2022","source":"Molecular medicine reports","url":"https://pubmed.ncbi.nlm.nih.gov/35485284","citation_count":6,"is_preprint":false},{"pmid":"36243722","id":"PMC_36243722","title":"A heterozygous missense variant in the YWHAG gene causing developmental and epileptic encephalopathy 56 in a Chinese family.","date":"2022","source":"BMC medical genomics","url":"https://pubmed.ncbi.nlm.nih.gov/36243722","citation_count":6,"is_preprint":false},{"pmid":"29253567","id":"PMC_29253567","title":"Plakoglobin localization to the cell border restores desmosome function in cells lacking 14-3-3γ.","date":"2017","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/29253567","citation_count":6,"is_preprint":false},{"pmid":"38640169","id":"PMC_38640169","title":"Prospective Role of PAK6 and 14-3-3γ as Biomarkers for Parkinson's Disease.","date":"2024","source":"Journal of Parkinson's disease","url":"https://pubmed.ncbi.nlm.nih.gov/38640169","citation_count":5,"is_preprint":false},{"pmid":"39918010","id":"PMC_39918010","title":"Thymoquinone mitigates cardiac hypertrophy by activating adaptive autophagy via the PPAR‑γ/14‑3‑3γ pathway.","date":"2025","source":"International journal of molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/39918010","citation_count":5,"is_preprint":false},{"pmid":"34626438","id":"PMC_34626438","title":"Disruption of desmosome function leads to increased centrosome clustering in 14-3-3γ-knockout cells with supernumerary centrosomes.","date":"2021","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/34626438","citation_count":5,"is_preprint":false},{"pmid":"34876904","id":"PMC_34876904","title":"LncRNA PTPRG-AS1 Promotes the Metastasis of Hepatocellular Carcinoma by Enhancing YWHAG.","date":"2021","source":"Journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/34876904","citation_count":5,"is_preprint":false},{"pmid":"36549645","id":"PMC_36549645","title":"The 14-3-3γ isoform binds to and regulates the localization of endoplasmic reticulum (ER) membrane protein TMCC3 for the reticular network of the ER.","date":"2022","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/36549645","citation_count":4,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":54549,"output_tokens":13992,"usd":0.186763,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":26256,"output_tokens":5002,"usd":0.128165,"stage2_stop_reason":"end_turn"},"total_usd":0.314928,"stage1_batch_id":"msgbatch_01UT1AahFPCAyn2uVweS7A9z","stage2_batch_id":"msgbatch_01HuKWtKNT4Pqj1bc8eEP5RV","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2006,\n      \"finding\": \"14-3-3γ binds directly to MDMX phosphorylated at Ser367 by Chk1 in response to UV irradiation, causing cytoplasmic retention of MDMX and suppression of MDMX-enhanced p53 ubiquitination, leading to p53 stabilization. The interaction requires phosphorylation of MDMX; the K50E mutant of 14-3-3γ that cannot bind MDMX fails to stabilize p53. siRNA ablation of 14-3-3γ reduces UV-induced p53 levels and G1 arrest.\",\n      \"method\": \"Immuno-affinity purification coupled with mass spectrometry, in vitro binding assay, co-immunoprecipitation, kinase-dead Chk1 mutant, Chk1 inhibitor UCN-01, siRNA knockdown, p53 ubiquitination assay\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, in vitro direct binding, active-site/mutant controls, kinase inhibitor, siRNA, multiple orthogonal methods in a single rigorous study\",\n      \"pmids\": [\"16511572\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"14-3-3γ forms a ternary complex with Chk1 (phosphorylated at Ser296 by autophosphorylation after ATR activation) and Cdc25A, mediating Cdc25A phosphorylation and proteasomal degradation to block premature mitotic entry after UV-induced DNA damage. Replacement of endogenous Chk1 with a Ser296Ala mutant causes premature mitotic entry after UV irradiation.\",\n      \"method\": \"Co-immunoprecipitation, phospho-specific antibodies, Chk1 Ser296Ala mutant replacement, UV irradiation, cell cycle analysis\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, phospho-site mutant rescue experiment, multiple orthogonal methods, clear functional readout\",\n      \"pmids\": [\"20639859\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Plk1 phosphorylated at Ser99 (by a PI3K/Akt-dependent mechanism) creates a docking site for 14-3-3γ; this interaction stimulates Plk1 catalytic activity. Knockdown of 14-3-3γ or expression of a Ser99-phospho-blocking Plk1 mutant causes prometaphase/metaphase arrest by activating the spindle assembly checkpoint, demonstrating that this interaction is required for proper metaphase-to-anaphase transition.\",\n      \"method\": \"Co-immunoprecipitation, phospho-specific antibodies, Plk1 Ser99Ala mutant, siRNA knockdown of 14-3-3γ, PI3K/Akt inhibitors, kinase activity assay, mitotic arrest analysis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, phospho-site mutant, kinase activity assay, siRNA, multiple orthogonal methods with clear functional readout\",\n      \"pmids\": [\"23695676\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"14-3-3γ dimers act as a scaffold that bridges CtBP1-S/BARS to PI(4)KIIIβ at the Golgi complex, coupling carrier budding and fission processes. The complex is stabilized by PKD- and PAK-mediated phosphorylation. Disrupting the association of these proteins inhibits fission of elongating post-Golgi carrier precursors.\",\n      \"method\": \"Co-immunoprecipitation, protein complex reconstitution, phosphorylation assays with PKD and PAK, dominant-negative disruption, live-cell imaging of carrier fission\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, functional disruption of complex with defined cellular phenotype, phosphorylation-dependent complex stabilization, multiple orthogonal methods\",\n      \"pmids\": [\"22366688\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"14-3-3γ negatively regulates steroidogenesis in MA-10 Leydig cells by binding to phospho-Ser194 in the START domain of StAR protein in a hormone-dependent manner, keeping StAR in an unfolded, inactive state. Over time, 14-3-3γ homodimerizes and dissociates from StAR, permitting maximal mitochondrial steroid formation. Silencing 14-3-3γ potentiates steroidogenesis.\",\n      \"method\": \"Mass spectrometry identification in native mitochondrial complexes, immunoprecipitation, site-directed mutagenesis (Ser194 binding site), siRNA knockdown, steroid output assay, cAMP stimulation\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — MS identification, Co-IP, phospho-site mutagenesis, siRNA, functional steroid output assay — multiple orthogonal methods in single study\",\n      \"pmids\": [\"22427666\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Ser58 phosphorylation and Lys49 acetylation of 14-3-3γ regulate its homodimerization and interaction with StAR in a coordinated, time-dependent manner during cAMP-induced steroidogenesis. Blocking either modification further induces steroidogenesis and reduces lipid storage.\",\n      \"method\": \"TAT-peptide blocking of phosphorylation (Ser58) and acetylation (Lys49) sites, co-immunoprecipitation, steroid output and lipid storage assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — peptide-blocking approach, Co-IP, functional readout; single lab, two orthogonal methods\",\n      \"pmids\": [\"25086053\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Phosphorylated tyrosine hydroxylase (TH) at Ser19 binds 14-3-3γ with high affinity (Kd ~3.2 nM), forming complexes of one TH tetramer with one or two 14-3-3γ dimers. 14-3-3γ binding inhibits PKA-mediated phosphorylation of TH at Ser40 (3.5-fold reduction), suggesting Ser40 has reduced accessibility in the complex. TH-pS40 alone does not detectably bind 14-3-3γ.\",\n      \"method\": \"Native mass spectrometry, surface plasmon resonance, electron microscopy, phosphatase kinetics, in vitro kinase assay\",\n      \"journal\": \"Molecular & cellular proteomics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — native MS stoichiometry, SPR binding kinetics, EM structural analysis, in vitro enzymatic assay; multiple orthogonal methods in one rigorous study\",\n      \"pmids\": [\"24947669\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"PAK6 phosphorylates 14-3-3γ at Ser59, and this phosphorylation acts as a switch that dissociates 14-3-3γ from client proteins including LRRK2 (at phospho-Ser935), causing LRRK2 dephosphorylation. A constitutively active PAK6 rescues G2019S LRRK2-associated neurite shortening through this phosphorylation of 14-3-3γ.\",\n      \"method\": \"Co-immunoprecipitation of PAK6 interactome, in vitro kinase assay with phospho-site identification (Ser59), neurite length measurement in neurons, LRRK2 phospho-Ser935 western blot, PAK6 constitutively active mutant\",\n      \"journal\": \"Frontiers in molecular neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP interactome, in vitro kinase assay with phospho-site identification, loss-of-function (14-3-3γ dissociation), neuronal rescue experiment; multiple orthogonal methods\",\n      \"pmids\": [\"29311810\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Phosphorylated Hsp20 (HspB6) forms a tight complex with 14-3-3γ in which a dimer of 14-3-3γ binds a dimer of Hsp20. 14-3-3γ increases the chaperone activity of phosphorylated Hsp20 when insulin is used as a model substrate. Unphosphorylated Hsp20 and its S16D phosphomimetic mutant do not interact with 14-3-3γ.\",\n      \"method\": \"Size-exclusion chromatography, chemical crosslinking, in vitro chaperone activity assay with insulin substrate\",\n      \"journal\": \"Molecular and cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro reconstitution of complex, functional chaperone assay; single lab, two orthogonal methods\",\n      \"pmids\": [\"17109079\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"14-3-3γ is poly-ADP-ribosylated in the nucleus after traumatic brain injury, and nuclear poly-ADP-ribosylation of 14-3-3γ is completely inhibited by the PARP-1 inhibitor dose that produces profound memory disturbances, linking this modification to spatial memory acquisition.\",\n      \"method\": \"Proteomics identification of poly-ADP-ribosylated peptides from brain, PARP-1 inhibitor treatment in vivo (INH2BP), Morris water maze behavioral testing\",\n      \"journal\": \"Journal of neurochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proteomics-based identification of ribosylation site, in vivo pharmacological intervention with functional behavioral readout; single lab\",\n      \"pmids\": [\"12694396\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Dimeric 14-3-3γ simultaneously binds both AICD (amyloid beta-protein precursor intracellular domain) and FE65, facilitating FE65-dependent gene transactivation by enhancing AICD-FE65 association. The interaction requires the VTPEER motif (residues 667-672) of AICD; phosphorylation of AICD at Thr668 within this motif inhibits 14-3-3γ binding and blocks gene transactivation. 14-3-3γ binds to a region between the WW domain and first PTB domain of FE65.\",\n      \"method\": \"Co-immunoprecipitation, in vitro binding assay, deletion/point mutants of AICD and FE65, gene transactivation reporter assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP, in vitro direct binding, multiple domain-mapping mutants, functional reporter assay; multiple orthogonal methods in one study\",\n      \"pmids\": [\"16223726\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Endogenous 14-3-3γ co-immunoprecipitates with c-Raf-1 and p-Raf-259 in primary astrocyte cultures, suggesting 14-3-3γ links Raf to signaling pathways controlling cell growth and ischemia-induced apoptosis. The induction of 14-3-3γ in ischemic astrocytes was not suppressed by PI3K or MAP kinase inhibitors.\",\n      \"method\": \"Co-immunoprecipitation, Northern/Western blot, pharmacological inhibitors (U0126, LY294002), anaerobic ischemia model in cultured astrocytes\",\n      \"journal\": \"Glia\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — single Co-IP with Raf, replicated in two Raf forms, inhibitor experiments showing pathway independence; single lab\",\n      \"pmids\": [\"12730952\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"14-3-3γ associates with phosphorylated GFAP (specifically at Ser8 in the head domain) in a phosphorylation- and cell-cycle-dependent manner, with increased association during G2/M phase. Overexpression of 14-3-3γ destroys the integrity and affects the movement of GFAP intermediate filaments in astrocytes.\",\n      \"method\": \"Co-immunoprecipitation, domain deletion and Ser-to-Ala substitution mutants of GFAP, cell-cycle synchronization, live imaging of intermediate filament dynamics, immunofluorescence\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP with phospho-site mutagenesis, cell-cycle synchronization, live imaging of functional consequence; multiple orthogonal methods\",\n      \"pmids\": [\"17032734\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Under ischemia, 14-3-3γ (the sole 14-3-3 isoform inducible by ischemia in astrocytes) binds phospho-Bad, preventing Bad translocation to mitochondria and inhibiting apoptosis. Overexpression of 14-3-3γ promotes astrocyte survival; antisense suppression enhances apoptosis under ischemia.\",\n      \"method\": \"Co-immunoprecipitation of endogenous 14-3-3γ with p-Bad, overexpression, antisense knockdown, cell death quantification, isoform-specific western blots\",\n      \"journal\": \"Journal of cerebral blood flow and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP of endogenous complex, gain- and loss-of-function with survival readout; single lab\",\n      \"pmids\": [\"15660102\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Endogenous 14-3-3γ co-immunoprecipitates with detergent-soluble actin in astrocytes, and this association increases after 4 h of ischemia. 14-3-3γ co-localizes with F-actin during cell division (forming a ring-like structure around daughter nuclei) and in surviving ischemic astrocytes, but dissociates from actin filaments in apoptotic astrocytes.\",\n      \"method\": \"Reciprocal co-immunoprecipitation, immunofluorescence co-localization, ischemia model\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — reciprocal Co-IP, immunofluorescence co-localization across multiple functional states; single lab\",\n      \"pmids\": [\"12176032\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"14-3-3γ is phosphorylated by multiple PKC isoforms (alpha, beta, gamma, theta, delta) in a PDGF-dependent manner in vascular smooth muscle cells. 14-3-3γ also interacts with the signal transduction protein Raf-1, suggesting it links Raf to PKC signaling.\",\n      \"method\": \"Co-immunoprecipitation with PKC isoforms, PKC inhibitor treatment, PKC activator stimulation, phosphorylation assay\",\n      \"journal\": \"DNA and cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP with multiple PKC isoforms, pharmacological activation/inhibition; single lab, two orthogonal approaches\",\n      \"pmids\": [\"10433554\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"14-3-3γ interaction with phospholipid bilayers is stimulated when complexed with a Ser19-phosphorylated tyrosine hydroxylase peptide. Membrane binding is isoform-specific and depends on histidine residues His158 and His195 (unique to the γ isoform) at the convex lateral side, as shown by site-directed mutagenesis. Electrostatic analysis and molecular dynamics indicate that phosphopeptide-bound 14-3-3γ has an optimal electrostatic potential for membrane interaction through N-terminal amphipathic helices.\",\n      \"method\": \"Surface plasmon resonance (membrane binding assay), site-directed mutagenesis (His158, His195), molecular dynamics simulation, electrostatic analysis of crystal structures\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — SPR biophysical assay, site-directed mutagenesis with functional validation, MD simulation; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"23189152\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"14-3-3γ identified as a binding partner of ANO1 (anoctamin-1) by yeast two-hybrid screening; the Thr9 residue of ANO1 is critical for the interaction. 14-3-3γ enhances surface expression of ANO1 (anterograde trafficking). Gene silencing of 14-3-3γ and/or ANO1 inhibits migration and invasion of glioblastoma cells.\",\n      \"method\": \"Yeast two-hybrid screening, co-immunoprecipitation, site-directed mutagenesis (Thr9), cell surface expression assay, siRNA knockdown, migration/invasion assays\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast two-hybrid + Co-IP + mutagenesis + functional cell assay; single lab\",\n      \"pmids\": [\"27212225\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"14-3-3γ binds TRPM4b at its N-terminus via a Ser88-dependent interaction (GST pull-down and Co-IP). Overexpression of 14-3-3γ increases TRPM4b plasma membrane expression measured by whole-cell electrophysiology and surface biotinylation; shRNA against 14-3-3γ greatly reduces TRPM4b surface expression and attenuates glutamate-induced TRPM4b currents and neuronal cell death.\",\n      \"method\": \"Yeast two-hybrid, GST pull-down, co-immunoprecipitation, whole-cell patch clamp, cell surface biotinylation, shRNA knockdown, glutamate-induced cell death assay\",\n      \"journal\": \"Molecular brain\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — yeast two-hybrid + GST pull-down + Co-IP + electrophysiology + surface biotinylation + shRNA functional rescue; multiple orthogonal methods\",\n      \"pmids\": [\"25047048\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"14-3-3γ binds plakoglobin in a PKCμ-dependent (PKD-dependent) manner and mediates microtubule-dependent (KIF5B-KLC1 motor complex-dependent) transport of plakoglobin to cell borders, initiating desmosome assembly. Loss of 14-3-3γ reduces cell-cell adhesion and desmosome formation both in vitro (HCT116 cells) and in vivo (mouse testis), leading to defects in spermatogenesis.\",\n      \"method\": \"Co-immunoprecipitation, PKCμ inhibitor, microtubule disruption, KIF5B knockdown, immunofluorescence, in vivo testis phenotype analysis\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP with kinase inhibitor, motor protein knockdown, in vitro and in vivo loss-of-function with defined cellular/tissue phenotypes; multiple orthogonal methods\",\n      \"pmids\": [\"24610948\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Loss of 14-3-3γ leads to centrosome amplification via phosphorylation of NPM1 at Thr199, causing early centriole disjunction and centrosome hyper-duplication, which results in aneuploidy and increased tumor formation. 14-3-3γ localizes to the centrosome. Expression of a constitutively active 14-3-3-binding-defective Cdc25C (S216A) mutant in 14-3-3γ-knockdown cells increases multipolar spindle formation.\",\n      \"method\": \"14-3-3γ knockdown, NPM1 phospho-Thr199 detection, centriole duplication assay, centrosome immunofluorescence, in vivo tumor formation, Cdc25C S216A mutant expression, spindle analysis\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with defined molecular (NPM1 phosphorylation) and cellular (centrosome amplification) phenotypes; single lab, multiple readouts\",\n      \"pmids\": [\"27253419\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Ischemia-induced upregulation of 14-3-3γ in astrocytes is mediated by activation of the JNK/c-Jun/AP-1 pathway. Only the JNK inhibitor SP600125 (not ERK, p38, or PI3K inhibitors) blocked ischemia-induced 14-3-3γ upregulation. Nuclear translocation of p-c-Jun under ischemia and AP-1 inhibition by curcumin also suppressed 14-3-3γ induction.\",\n      \"method\": \"Selective kinase inhibitors (SP600125, U0126, SB203580, LY294002), curcumin AP-1 inhibition, nuclear translocation of p-c-Jun by immunofluorescence, Western blot for 14-3-3γ, in vitro ischemia model\",\n      \"journal\": \"Journal of neurochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological pathway dissection with multiple inhibitors, nuclear translocation imaging; single lab\",\n      \"pmids\": [\"19393026\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Overexpression of 14-3-3γ in NIH3T3 cells induces oncogenic transformation (focus formation and tumor formation in SCID mice). Transformation requires activation of both MAPK and PI3K signaling pathways. 14-3-3γ co-immunoprecipitates with PI3K and TSC2, indicating it can stimulate PI3K signaling at two points.\",\n      \"method\": \"Focus formation assay, soft agar colony formation, tumor formation in SCID mice, PI3K/MAPK inhibitor treatment, co-immunoprecipitation with PI3K and TSC2\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo tumor formation, Co-IP with signaling proteins, pharmacological pathway inhibition; single lab, multiple methods\",\n      \"pmids\": [\"20628654\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The N-terminal variable region II (VRII) of 14-3-3γ (within the first 40 amino acids) is required for its oncogenic activity (activation of PI3K and MAPK signaling and cellular transformation). Two residues within VRII are required and two contribute to the γ-specific phenotypes; swapping this region with the equivalent region from the tumor suppressor 14-3-3σ switches the functional outcome.\",\n      \"method\": \"14-3-3γ/σ chimeric protein constructs, focus formation assay, soft agar growth, PI3K/MAPK activation assays, individual amino acid substitutions\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain-swap chimeras and individual amino acid substitutions with transformation functional assay; single lab\",\n      \"pmids\": [\"23115241\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"14-3-3γ protects p21 from MDMX-mediated proteasomal degradation (independent of p53) by competing with p21 for MDMX binding in a dose-dependent manner. Overexpression of 14-3-3γ extends p21 half-life and causes p21-dependent G1 arrest in p53-null cells. In response to DNA damage, the 14-3-3γ-MDMX complex increases while the MDMX-p21 complex decreases.\",\n      \"method\": \"Co-immunoprecipitation in vitro with purified proteins and in cells, p21 half-life measurement, G1 arrest assay in p53-null cells, siRNA knockdown of 14-3-3γ\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vitro Co-IP with purified proteins + in-cell Co-IP + half-life measurement + loss-of-function siRNA + functional cell cycle readout; multiple orthogonal methods\",\n      \"pmids\": [\"21148311\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"14-3-3γ binds to and undergoes poly-ADP-ribosylation; in the context of the MALM (Mieap-induced accumulation of lysosome-like organelles within mitochondria) process, 14-3-3γ localizes within mitochondria and interacts with Mieap. Deficiency of 14-3-3γ does not affect accumulation of Mieap or lysosomal proteins within mitochondria but dramatically inhibits elimination of oxidized mitochondrial proteins.\",\n      \"method\": \"2DICAL (two-dimensional image-converted LC-MS) proteomics of immunoprecipitated Mieap complex, co-immunoprecipitation of exogenous and endogenous proteins, subcellular fractionation, immunofluorescence, 14-3-3γ knockdown with functional readout\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS-based identification + Co-IP + subcellular localization + functional knockdown; single lab\",\n      \"pmids\": [\"22532927\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"14-3-3γ interacts with ERK1c as part of a PI4KIIIβ-14-3-3γ complex that mediates Golgi translocation of ERK1c during prophase/prometaphase. CDK1 phosphorylates ERK1c at Ser343, enabling complex assembly. PKD-mediated phosphorylation of PI4KIIIβ stabilizes the complex. The complex assembly induces ERK1c Golgi translocation where it is activated by MEK1b to induce Golgi fragmentation.\",\n      \"method\": \"Co-immunoprecipitation, CDK1 phosphorylation assay, PKD inhibitor, dominant-negative constructs, time-lapse imaging to determine translocation timing, kinase assays\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, kinase assays, pharmacological inhibitors, live imaging; single lab\",\n      \"pmids\": [\"26459638\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"14-3-3γ co-immunoprecipitates with phospho-Bad(S112) in LPS-treated cardiomyocytes, and overexpressed 14-3-3γ promotes Bad(S112) phosphorylation and sequesters p-Bad, causing Bcl-2 dissociation from the Bad/Bcl-2 complex and Bcl-2 translocation to mitochondria, preventing mPTP opening and cytochrome c release.\",\n      \"method\": \"Co-immunoprecipitation, subcellular fractionation (cytosolic/mitochondrial), flow cytometry (apoptosis, MMP), mitochondrial swelling assay (mPTP), Western blot for Bcl-2 family members\",\n      \"journal\": \"International immunopharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, subcellular fractionation with functional readout, multiple downstream markers; single lab\",\n      \"pmids\": [\"24957688\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"14-3-3γ binds directly to p-β-catenin Ser37 (but not p-Bad, p-Ask-1, p-p53, or Bax) in cortical neurons as shown by Co-IP and FRET. During oxygen-glucose deprivation, 14-3-3γ translocates to the nucleus correlating with increased nuclear p-β-catenin Ser37. 14-3-3γ overexpression reduces Bax expression and prevents p-β-catenin-Ser37-dependent Bax upregulation and cell death.\",\n      \"method\": \"Co-immunoprecipitation, FRET, nuclear fractionation, OGD model, 14-3-3γ overexpression/knockdown, β-catenin S37A mutant, Bax measurement, cell viability\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — FRET (direct interaction), Co-IP, phospho-site mutant S37A, loss- and gain-of-function with multiple functional readouts; multiple orthogonal methods\",\n      \"pmids\": [\"24743739\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"14-3-3γ specifically binds to 5'-AGCT-3' repeats in IgH switch regions and interacts directly with the C-terminal region of activation-induced cytidine deaminase (AID), targeting AID to switch regions to mediate class switch recombination. Induction of 14-3-3γ expression in B cells requires NF-κB recruitment to the 14-3-3γ promoter, which promotes CFP1-mediated H3K4me3 enrichment, followed by E2A binding for sustained expression.\",\n      \"method\": \"ChIP assays, B cell stimulation with CSR-inducing stimuli, NF-κB inhibition, promoter analysis, 14-3-3γ expression kinetics\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP, pharmacological NF-κB inhibition, B cell stimulation kinetics; single lab; mechanistic detail on transcriptional regulation\",\n      \"pmids\": [\"23851690\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Protein kinase CK2 interacts strongly with 14-3-3γ at the neuromuscular junction and phosphorylates 14-3-3γ at serine residue 235.\",\n      \"method\": \"Co-immunoprecipitation, in vitro kinase assay with phospho-site mapping\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP + in vitro kinase assay with specific phospho-site identification; single lab\",\n      \"pmids\": [\"26198629\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"14-3-3γ binds deubiquitinase USP37, which stabilizes 14-3-3γ by preventing its ubiquitin-dependent proteasomal degradation through its catalytic deubiquitinase activity.\",\n      \"method\": \"Co-immunoprecipitation (binding partner screen), ubiquitination assay, USP37 catalytic mutant, protein stability assay\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP, ubiquitination assay, catalytic mutant; single lab\",\n      \"pmids\": [\"26427597\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"p53 interacts with the C-terminal domain of 14-3-3γ, induces 14-3-3γ ubiquitination, and promotes proteasome-mediated degradation of 14-3-3γ. MG132 (26S proteasome inhibitor) blocks this effect. Wild-type but not mutant p53 (R175H) suppresses 14-3-3γ protein levels.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, proteasome inhibitor (MG132), wild-type vs. mutant p53 expression, protein half-life analysis\",\n      \"journal\": \"International journal of oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP, ubiquitination assay, proteasome inhibitor, mutant controls; single lab\",\n      \"pmids\": [\"25384678\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"14-3-3γ binds preferentially to supercoiled DNA over linear DNA, with particular affinity for cruciform DNA structures. In HCT-116 cells, 14-3-3γ co-localizes with DNA cruciforms by confocal microscopy.\",\n      \"method\": \"Electrophoretic mobility shift assay, competition with magnetic beads (linear vs. supercoiled DNA), confocal microscopy with cruciform-detecting probes\",\n      \"journal\": \"Journal of biomolecular structure & dynamics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — EMSA, magnetic bead competition assay, confocal co-localization; single lab, in vitro binding confirmed in cells\",\n      \"pmids\": [\"22856523\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"14-3-3γ interacts with eukaryotic translation initiation factor eIF1AX and ribosomal protein RPS7, as confirmed by Co-IP with mass spectrometry and FRET/co-localization. 14-3-3γ positively regulates protein synthesis by affecting eIF1AX and RPS7 expression levels and mTOR pathway activity in bovine mammary epithelial cells.\",\n      \"method\": \"Co-immunoprecipitation with MALDI-TOF/TOF mass spectrometry, FRET, co-localization, 14-3-3γ overexpression and silencing with protein synthesis readout\",\n      \"journal\": \"Archives of biochemistry and biophysics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP+MS identification confirmed by FRET, gain- and loss-of-function; single lab\",\n      \"pmids\": [\"25281768\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"14-3-3γ directly interacts with RGS14 at two distinct sites: a phosphorylation-independent site and a phosphorylation-dependent site at Ser218 (potentiated by active H-Ras signaling). The pSer218-dependent interaction inhibits active Gαi1-AlF4- binding to the RGS domain of RGS14 (measured by BRET), while the phosphorylation-independent interaction inhibits RGS14 nuclear import and nucleocytoplasmic shuttling.\",\n      \"method\": \"Bioluminescence resonance energy transfer (BRET), co-immunoprecipitation, site-directed mutagenesis (Ser218), active H-Ras stimulation, nuclear import assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — BRET (quantitative direct interaction), Co-IP, phospho-site mutagenesis, distinct functional readouts for each interaction site; multiple orthogonal methods\",\n      \"pmids\": [\"30093406\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"14-3-3γ directly interacts with bestrophin-1 (Best1) anion channel in astrocytes (identified by yeast two-hybrid; confirmed by BiFC). The interaction is mediated by phosphorylation of Ser358 in the C-terminus of Best1. 14-3-3γ silencing reduces Best1 surface expression and Best1-mediated whole-cell currents, and decreases Best1-mediated glutamate release from hippocampal astrocytes (recorded as reduced NMDA receptor current in CA1 neurons).\",\n      \"method\": \"Yeast two-hybrid, bimolecular fluorescence complementation (BiFC), whole-cell patch clamp, shRNA knockdown, hippocampal slice electrophysiology\",\n      \"journal\": \"Molecular brain\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — yeast two-hybrid + BiFC + electrophysiology + in vivo slice recording; multiple orthogonal methods, clear functional consequence at synapse level\",\n      \"pmids\": [\"29121962\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"14-3-3γ binds to TMCC3 (an ER membrane protein at three-way junctions) through phosphorylation of serine residues in deduced 14-3-3 binding motifs in the N-terminus of TMCC3. Overexpression of 14-3-3γ reduces TMCC3 localization to three-way junctions and decreases their number. A TMCC3 alanine-substitution mutant at the phosphorylatable serine shows reduced 14-3-3γ binding and is more resistant to 14-3-3γ-driven mislocalization, demonstrating that 14-3-3γ negatively regulates the reticular ER network by controlling TMCC3 distribution.\",\n      \"method\": \"Co-immunoprecipitation, site-directed mutagenesis (Ser-to-Ala in binding motif), overexpression, three-way junction quantification by fluorescence microscopy, ER morphology assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP, phospho-site mutagenesis, overexpression with quantified organelle morphology readout; multiple orthogonal methods in one study\",\n      \"pmids\": [\"36549645\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"14-3-3γ directly interacts with Copine1 (CPNE1) in hippocampal progenitor cells; among all seven 14-3-3 isoforms, only 14-3-3γ binds CPNE1. The interaction requires the Ser54 residue of the C2A domain of CPNE1; Ser54 mutation reduces 14-3-3γ binding and CPNE1-dependent AKT phosphorylation and neuronal differentiation.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation (in vitro and in vivo), Ser54 mutation, isoform selectivity assay, neurite outgrowth and neuronal marker expression\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast two-hybrid + Co-IP + phospho-site mutagenesis + neuronal differentiation readout; single lab\",\n      \"pmids\": [\"28412242\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"14-3-3γ hypoxia-mediated p53 activation occurs through the ATR-Chk1-MDMX-14-3-3γ pathway: hypoxia induces ATR-Chk1-dependent phosphorylation of MDMX at Ser367, enhancing MDMX binding to 14-3-3γ, which leads to p53 activation. In MEFs with MDMX containing the S367A mutation, hypoxia fails to induce MDMX-14-3-3γ binding or p53 activation.\",\n      \"method\": \"Co-immunoprecipitation, Chk1/ATR inhibitor and siRNA knockdown, phospho-specific antibody (pSer367-MDMX), MEFs with knock-in MDMX S367A mutant, p53 target gene induction\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP, knock-in phospho-site mutant in primary MEFs, ATR/Chk1 inhibitors, multiple functional readouts; replicates and extends prior UV-damage findings\",\n      \"pmids\": [\"22556425\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Loss of 14-3-3γ in mice leads to delayed neuronal migration and morphological defects (thicker leading process stem, failure to reach cortical plate-marginal zone boundary, increased multipolar neurons) in the developing cerebral cortex as shown by in utero electroporation knockdown and time-lapse live imaging of brain slices.\",\n      \"method\": \"In utero electroporation (shRNA knockdown), time-lapse live imaging of brain slices, immunofluorescence for cortical layer markers\",\n      \"journal\": \"Developmental neurobiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in utero loss-of-function with live imaging and morphological quantification; single lab\",\n      \"pmids\": [\"26297819\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Overexpression of 14-3-3γ in utero also causes delayed pyramidal neuron migration in the developing mouse cortex, similar to knockdown, indicating that a precise balance of 14-3-3γ expression is required for proper cortical development.\",\n      \"method\": \"In utero electroporation (overexpression), cortical layer marker immunofluorescence, neuronal position quantification\",\n      \"journal\": \"Neuroscience letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in utero gain-of-function with defined cortical migration phenotype; single lab, replicates the loss-of-function findings with the opposite manipulation\",\n      \"pmids\": [\"27288018\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"An LC-MS proteomics screen of 14-3-3γ-interacting proteins in MA-10 Leydig cells identified 688 interactors including Sec23ip (a vesicle trafficking protein). Silencing Sec23ip decreased steroidogenesis and impaired cholesterol mobilization from cytoplasmic membrane to mitochondria, placing Sec23ip in the 14-3-3γ-regulated steroidogenic network.\",\n      \"method\": \"Liquid chromatography–mass spectrometry of immunoprecipitated 14-3-3γ complex, siRNA knockdown of Sec23ip, steroidogenesis assay, cholesterol trafficking assay\",\n      \"journal\": \"Endocrinology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — LC-MS interactome, siRNA functional validation with steroid output and cholesterol trafficking; single lab\",\n      \"pmids\": [\"31875919\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"14-3-3γ knockout cells with supernumerary centrosomes show increased centrosome clustering and pseudo-bipolar mitoses compared to 14-3-3ε knockout cells that show multipolar mitoses. Loss of 14-3-3γ compromises desmosome function and decreases keratin levels, reducing cell stiffness and promoting centrosome clustering. Restoration of desmosome function increased multipolar mitoses; knockdown of plakoglobin or keratin 5 reduced cell stiffness and increased pseudo-bipolar mitoses.\",\n      \"method\": \"14-3-3γ/ε knockout cell lines, centrosome counting and spindle analysis, desmosome rescue experiments, plakoglobin/keratin 5 knockdown, atomic force microscopy or equivalent cell stiffness assay\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knockout comparison, desmosome rescue, cytoskeletal knockdown with mechanical and mitotic readouts; single lab\",\n      \"pmids\": [\"34626438\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"14-3-3γ directly interacts with CPNE1 and a yeast two-hybrid screen confirmed isoform specificity (only γ among seven isoforms). Also, YWHAG knockdown in breast cancer cells reduces Snail protein stability, EMT markers, and invasion; ectopic YWHAG overexpression abrogates miR-181b-3p-induced Snail stabilization and EMT.\",\n      \"method\": \"Luciferase reporter assay (miR-181b-3p target validation), siRNA/miRNA transfection, Western blot for Snail stability, invasion and migration assays, in vivo lung metastasis assay in mice\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — miRNA target validation by luciferase, protein stability by siRNA knockdown/overexpression with invasion readout and in vivo metastasis; single lab\",\n      \"pmids\": [\"27102539\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"14-3-3γ deficiency causes rapid ROS accumulation and delayed EMT in cancer cells, revealing a YWHAG-dependent cytoprotective mechanism embedded in EMT-associated networks that protects cancer cells from oxidative catastrophe through enhanced autophagy. Tumor allografts show that metastasis potential correlates with YWHAG expression level, and YWHAG silencing diminishes primary tumor volumes and prevents metastasis.\",\n      \"method\": \"Cellular kinome and transcriptome analysis (regulome construction), ROS assay, EMT markers, autophagy flux assay, 14-3-3γ knockdown, in vivo tumor allograft and metastasis model\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — transcriptome/kinome regulome, ROS + autophagy assays, in vivo metastasis model with YWHAG knockdown; single lab, multiple readouts\",\n      \"pmids\": [\"37759388\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"YWHAG promotes bladder cancer cell invasion and metastasis by interacting with and upregulating TMOD3, which then activates ERK1/2 and JNK phosphorylation in the MAPK pathway. The YWHAG-TMOD3 interaction was confirmed by pull-down with mass spectrometry and co-immunoprecipitation. TMOD3 knockdown reversed the pro-metastatic effects of YWHAG.\",\n      \"method\": \"Pull-down assay with mass spectrometry, co-immunoprecipitation, transcriptome sequencing, TMOD3 knockdown, in vitro invasion/metastasis assays, in vivo metastasis model\",\n      \"journal\": \"Journal of translational medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pull-down+MS, Co-IP, transcriptome, epistasis (TMOD3 knockdown reverses YWHAG effect); single lab\",\n      \"pmids\": [\"39741303\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"14-3-3γ is localized to pseudopodia of MDA-MB-231 breast cancer cells and co-localizes with F-actin there. Knockdown of 14-3-3γ decreases pseudopodial formation/elongation and cell migration; forced expression has the opposite effect.\",\n      \"method\": \"Excimer laser cell etching to isolate pseudopodial proteins, confocal imaging, siRNA knockdown and overexpression, pseudopodia counting, wound healing and Transwell migration assays\",\n      \"journal\": \"Breast cancer (Tokyo, Japan)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — subcellular fractionation (pseudopodia isolation), confocal co-localization, gain- and loss-of-function with motility readout; single lab\",\n      \"pmids\": [\"30830684\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"YWHAG promotes CRC cell proliferation, migration, and invasion by interacting with cortactin (CTTN), which activates Wnt/β-catenin signaling. CTTN was identified as a YWHAG-associated protein mediating YWHAG's tumor-promoting functions.\",\n      \"method\": \"Co-immunoprecipitation (CTTN-YWHAG interaction), RNA-seq pathway analysis, CTTN/YWHAG knockdown with proliferation/invasion readouts, Wnt/β-catenin reporter\",\n      \"journal\": \"Medical oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP, RNA-seq pathway analysis, functional knockdown; single lab\",\n      \"pmids\": [\"38538804\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"14-3-3γ haploinsufficiency in aged mice leads to decreased dopamine levels and altered dopamine metabolism in the brain, along with changes in phosphorylation of proteins implicated in PD pathology, and PD-like motor coordination deficits.\",\n      \"method\": \"Heterozygous 14-3-3γ knockout mice, dopamine HPLC quantification, phospho-Western blot of PD-relevant proteins, behavioral testing (rotarod, nest-building)\",\n      \"journal\": \"Molecular brain\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic loss-of-function model with biochemical (dopamine, phospho-proteins) and behavioral (motor) readouts; single lab\",\n      \"pmids\": [\"36604743\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"14-3-3γ (YWHAG) is a phosphoserine/phosphothreonine-binding dimeric scaffold protein that regulates diverse cellular processes by sequestering or co-localizing phosphorylated client proteins: it binds Chk1-phosphorylated MDMX to retain it in the cytoplasm and activate p53, forms a ternary complex with Chk1 and Cdc25A to trigger Cdc25A degradation and prevent premature mitosis, docks to Plk1-pSer99 to stimulate its kinase activity and drive metaphase-to-anaphase transition, bridges CtBP1-S/BARS with PI4KIIIβ at the Golgi to couple carrier budding and fission, protects p21 from MDMX-dependent proteasomal degradation, sequesters phospho-Bad to prevent apoptosis, binds phospho-GFAP to regulate intermediate filament dynamics, promotes surface expression of ion channels (TRPM4b, ANO1, Best1) by binding phosphorylated N/C-terminal motifs, negatively regulates StAR-dependent steroidogenesis via phospho-Ser194 binding (controlled by its own Ser58 phosphorylation and Lys49 acetylation), transports plakoglobin to cell borders (via PKD/KIF5B) to initiate desmosome assembly, controls the reticular ER network by displacing TMCC3 from three-way junctions, binds tyrosine hydroxylase phosphorylated at pSer19 with nanomolar affinity to stabilize and regulate it, is itself phosphorylated by PAK6 at Ser59 (switching it off LRRK2), by CK2 at Ser235, and by multiple PKC isoforms in response to PDGF; its stability is maintained by the deubiquitinase USP37 and suppressed by p53-driven proteasomal degradation; and its precise expression level is critical for proper neuronal migration in cortical development.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"14-3-3γ (YWHAG) is a dimeric phosphoserine/phosphothreonine-binding scaffold that governs the localization, stability, and activity of phosphorylated client proteins across cell-cycle control, membrane trafficking, cytoskeletal dynamics, and survival signaling [#0, #6, #19]. In the DNA-damage and hypoxia response it acts as a node of the ATR–Chk1 axis: it binds Chk1-phosphorylated MDMX (pSer367) to enforce cytoplasmic retention of MDMX and stabilize p53 [#0, #39], forms a ternary complex with Chk1-pSer296 and Cdc25A to drive Cdc25A degradation and block premature mitosis [#1], and protects p21 from MDMX-dependent proteasomal turnover independently of p53 [#24]. During mitosis it docks onto Plk1-pSer99 to stimulate Plk1 activity and enable the metaphase-to-anaphase transition [#2], and its loss produces NPM1-pThr199–dependent centrosome amplification and aneuploidy [#20]. As a phosphorylation-gated scaffold it bridges CtBP1-S/BARS to PI4KIIIβ at the Golgi to couple post-Golgi carrier budding and fission [#3], displaces TMCC3 from ER three-way junctions to remodel the reticular ER [#37], directs PKD/KIF5B-dependent transport of plakoglobin to initiate desmosome assembly [#19], and promotes surface expression of the ion channels TRPM4b, ANO1, and Best1 through binding their phosphorylated N/C-terminal motifs [#17, #18, #36]. It binds phospho-Bad and p-β-catenin(Ser37) to suppress apoptosis [#27, #28], engages phospho-GFAP to control intermediate-filament integrity [#12], and binds tyrosine hydroxylase pSer19 with nanomolar affinity while restricting its PKA-mediated Ser40 phosphorylation [#6]. Its own activity is set by post-translational marks—PAK6 phosphorylates Ser59 to release clients including LRRK2-pSer935 [#7], CK2 phosphorylates Ser235 [#30]—and its abundance is balanced by USP37-mediated deubiquitination and p53-driven degradation [#31, #32]. In the brain, precise 14-3-3γ dosage is required for cortical neuronal migration, since both knockdown and overexpression delay migration [#40, #41], and haploinsufficiency lowers dopamine and produces PD-like motor deficits [#49]. Through oncogenic PI3K/MAPK activation and interactions with effectors such as TMOD3 and cortactin, it also drives transformation, EMT, and metastasis [#22, #46, #48].\"\n,\n  \"teleology\": [\n    {\n      \"year\": 2003,\n      \"claim\": \"Early work asked whether 14-3-3γ couples stress signaling to survival in glia, establishing it as a Raf-associated, ischemia-induced scaffold rather than a passive housekeeping protein.\",\n      \"evidence\": \"Co-IP of endogenous 14-3-3γ with c-Raf-1/p-Raf and actin in ischemic astrocyte cultures, with inhibitor-resistant induction\",\n      \"pmids\": [\"12730952\", \"12176032\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct binding to Raf not demonstrated with purified proteins\", \"Functional consequence of the Raf association not defined\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Defining 14-3-3γ as a phospho-dependent client binder, it was shown to retain Chk1-phosphorylated MDMX in the cytoplasm to stabilize p53, placing it directly in the DNA-damage checkpoint.\",\n      \"evidence\": \"Affinity-MS, in vitro binding, reciprocal Co-IP, K50E binding-dead mutant, Chk1-KD/UCN-01 and siRNA, with p53/G1-arrest readouts\",\n      \"pmids\": [\"16511572\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether the same module operates outside UV damage was not yet tested\", \"Structural basis of pSer367 recognition not resolved\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"The checkpoint role was extended to mitotic timing and to p53-independent cell-cycle control, showing 14-3-3γ enforces both Cdc25A degradation and p21 protection.\",\n      \"evidence\": \"Ternary Co-IP with Chk1-pSer296/Cdc25A plus S296A rescue; in vitro and in-cell Co-IP with MDMX/p21, half-life and G1-arrest assays in p53-null cells\",\n      \"pmids\": [\"20639859\", \"21148311\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry of the ternary complex not defined\", \"How damage signals partition 14-3-3γ between MDMX, Cdc25A, and p21 clients unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Beyond the nucleus, 14-3-3γ was established as a phosphorylation-gated membrane-trafficking scaffold and shown to negatively regulate steroidogenesis, revealing a general client-sequestration mechanism.\",\n      \"evidence\": \"Reconstituted CtBP1-S/BARS–PI4KIIIβ Golgi complex with live-imaging fission readout; MS/Co-IP/Ser194-mutant and siRNA in MA-10 Leydig cells with steroid output\",\n      \"pmids\": [\"22366688\", \"22427666\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How PKD/PAK phosphorylation switches complex assembly mechanistically not fully resolved\", \"In vivo relevance of Golgi scaffolding not tested\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"14-3-3γ was shown to be an activator, not merely a tether, by docking Plk1-pSer99 to stimulate its kinase activity and license anaphase onset.\",\n      \"evidence\": \"Co-IP, Plk1 S99A mutant, PI3K/Akt inhibitors, in vitro kinase assay and SAC-dependent mitotic-arrest analysis after siRNA\",\n      \"pmids\": [\"23695676\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural mechanism by which docking increases catalytic activity unknown\", \"Whether other mitotic kinases are similarly activated not addressed\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Biophysical and channel studies defined isoform-specific, high-affinity, phospho-dependent binding modes—nanomolar TH-pSer19 capture and membrane association via γ-specific histidines—and a recurring role in trafficking phosphorylated ion channels to the surface.\",\n      \"evidence\": \"Native MS/SPR/EM with TH; SPR with His158/His195 mutants and MD; GST pull-down/Co-IP/patch-clamp/biotinylation for TRPM4b with shRNA rescue\",\n      \"pmids\": [\"24947669\", \"23189152\", \"25047048\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether membrane recruitment is required for channel trafficking in vivo not tested\", \"Generality of His-mediated membrane binding across clients unknown\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"14-3-3γ was placed at the start of desmosome assembly, coupling kinase signaling to motor-driven transport of an adhesion protein.\",\n      \"evidence\": \"Co-IP, PKCμ inhibitor, microtubule disruption, KIF5B knockdown, and in vitro/in vivo (mouse testis) loss-of-function with spermatogenesis defects\",\n      \"pmids\": [\"24610948\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct phospho-site on plakoglobin recognized by 14-3-3γ not mapped\", \"Whether cargo handoff to the KIF5B motor is direct unknown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"The protein's abundance was shown to be actively controlled, identifying opposing USP37-mediated stabilization and p53-driven degradation as a homeostatic loop nested within the same checkpoint it regulates.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, USP37 catalytic mutant, MG132, and WT vs R175H p53 with half-life analysis\",\n      \"pmids\": [\"26427597\", \"25384678\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"E3 ligase driving p53-dependent 14-3-3γ ubiquitination not identified\", \"Physiological setting where each arm dominates undefined\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"In vivo and dosage experiments established that 14-3-3γ levels must be tightly balanced for cortical neuronal migration, indicating a developmental requirement beyond its biochemical interactions.\",\n      \"evidence\": \"In utero electroporation knockdown and overexpression with time-lapse imaging and cortical-layer marker analysis\",\n      \"pmids\": [\"26297819\", \"27288018\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Migration-relevant client(s) not identified\", \"Cell-autonomous vs non-autonomous contribution not separated\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Phospho-regulation of 14-3-3γ itself was defined as a client-release switch, linking PAK6 phosphorylation at Ser59 to LRRK2 dephosphorylation and neurite rescue.\",\n      \"evidence\": \"PAK6 interactome Co-IP, in vitro kinase assay with Ser59 mapping, LRRK2-pSer935 blots, and constitutively active PAK6 rescue of G2019S neurite shortening\",\n      \"pmids\": [\"29311810\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full set of clients released by Ser59 phosphorylation unknown\", \"Whether CK2-pSer235 has an analogous regulatory effect not tested\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Expanding its organelle-shaping role, 14-3-3γ was shown to negatively regulate the reticular ER network by sequestering phosphorylated TMCC3 away from three-way junctions.\",\n      \"evidence\": \"Co-IP, Ser-to-Ala TMCC3 mutant, overexpression, and quantified ER three-way-junction morphology\",\n      \"pmids\": [\"36549645\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Kinase that phosphorylates TMCC3 binding motif not identified\", \"Physiological trigger for ER remodeling by 14-3-3γ unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"A body of cancer studies established 14-3-3γ as a pro-metastatic effector acting through PI3K/MAPK and adhesion/cytoskeletal partners, tying its scaffolding biochemistry to invasion phenotypes.\",\n      \"evidence\": \"Pull-down/MS and Co-IP identifying TMOD3 and cortactin, RNA-seq pathway analysis, knockdown epistasis, and in vivo metastasis models\",\n      \"pmids\": [\"39741303\", \"38538804\", \"37759388\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Phospho-dependence of TMOD3/cortactin binding not established\", \"Direct vs indirect activation of Wnt and MAPK pathways unresolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how a single dimeric scaffold prioritizes among its many phosphorylated clients in a given cell, and what defines γ-isoform-specific functions in vivo.\",\n      \"evidence\": null,\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural model of competitive client selection\", \"Isoform-specific in vivo client repertoire not mapped\", \"Quantitative rules linking 14-3-3γ dosage to migration/disease phenotypes undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 3, 19, 24, 35]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [2, 6, 4, 7]},\n      {\"term_id\": \"GO:0019904\", \"supporting_discovery_ids\": [0, 6, 18, 36, 37]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [0, 13, 27]},\n      {\"term_id\": \"GO:0005794\", \"supporting_discovery_ids\": [3, 26]},\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [37]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [9, 28, 33]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [20]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [25]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [12, 14, 47]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [1, 2, 20]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [0, 39]},\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [3, 19]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [13, 27, 28]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [7, 22, 35]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [40, 41]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [44, 45, 46, 48, 49]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"MDMX\", \"CHEK1\", \"CDC25A\", \"PLK1\", \"STAR\", \"TH\", \"LRRK2\", \"RGS14\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}