{"gene":"PPP2CA","run_date":"2026-06-10T06:43:35","timeline":{"discoveries":[{"year":1999,"finding":"Human PP2Ac functionally replaced endogenous PP2Ac in S. cerevisiae and bound the yeast PR65/A subunit (Tpd3p) forming a dimer. The invariant C-terminal leucine residue (Leu-309) is dispensable for basic function but truncated or active-site mutant forms of PP2Ac exert dominant-negative effects by titrating regulatory subunits/substrates into non-productive complexes; the L199P active-site mutant was catalytically impaired despite binding Tpd3p.","method":"Yeast complementation assay, mutagenesis, in vitro phosphatase activity assay, computer modeling of active site","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution in yeast, mutagenesis, in vitro activity assay, multiple orthogonal methods in a single rigorous study","pmids":["10446173"],"is_preprint":false},{"year":2000,"finding":"The C-terminal Leu-377 residue of yeast Pph22p (equivalent to human PP2Ac Leu-309) is required for binding of the PR55/B subunit (Cdc55p) but not the PR61/B' subunit (Rts1p). Mutation of this leucine enhanced sensitivity to microtubule destabilization, phenocopying cdc55Δ cells with impaired spindle checkpoint function, linking PP2Ac C-terminal leucine to mitotic checkpoint regulation via B-subunit binding.","method":"Yeast genetics, in vitro phosphatase activity assay, yeast two-hybrid/binding assay, phenotypic analysis with microtubule-destabilizing drugs","journal":"Molecular & general genetics : MGG","confidence":"High","confidence_rationale":"Tier 1 / Strong — mutagenesis combined with functional assay and genetic epistasis, replicated mechanistic insight from companion paper","pmids":["11129046"],"is_preprint":false},{"year":2003,"finding":"Purified PP2Ac (but not PP4c or PP6c) bound to GST-alpha4 in pull-down assays and co-immunoprecipitated with endogenous and ectopic myc-tagged alpha4. The alpha4/PP2Ac interaction was disrupted by okadaic acid and microcystin-LR (phosphatase inhibitors) but was unaffected by rapamycin, implying the alpha4 binding site on PP2Ac includes the phosphatase catalytic domain.","method":"GST pull-down, co-immunoprecipitation, microcystin-Sepharose affinity purification, pharmacological inhibition","journal":"Protein expression and purification","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP and pull-down with purified proteins, single lab, two orthogonal methods","pmids":["12963337"],"is_preprint":false},{"year":2004,"finding":"Missense mutations and deletions within the PP2A-Aβ subunit (PPP2R1B) at amino acids 412–601 (the PP2Ac-binding region) inhibited co-immunoprecipitation of PP2A-Aβ and PP2Ac, demonstrating that this region of the A subunit is required for physical interaction with the catalytic subunit.","method":"Co-immunoprecipitation from colorectal cancer tissues, sequence analysis","journal":"Oncology reports","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP from patient tissues, replicated across multiple cancer samples but single lab, single method","pmids":["14767517"],"is_preprint":false},{"year":2005,"finding":"Alpha4 interacts with S6K1 through PP2Ac; pull-down assays defined that S6K1 binds the region from aa 88–309 of PP2Ac, while alpha4 binds two separated regions (aa 19–22 and aa 150–164) of PP2Ac. Stimulation of B cells with LPS induced the interaction of alpha4/PP2Ac with S6K1, suggesting alpha4 regulates S6K1 activity through PP2Ac.","method":"Co-immunoprecipitation, pull-down assay with deletion constructs, LPS stimulation of primary B cells","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain mapping by pull-down and Co-IP, single lab, two orthogonal methods","pmids":["15796902"],"is_preprint":false},{"year":2006,"finding":"CaMKIIα associates with PP2Ac and alpha4 in brain neurons. Neuron-specific alpha4 knockout mice showed increased CaMKIIα activity in hippocampus and impaired learning/memory. Alpha4 and PP2Ac were localized in the cytoplasm (not in the PSD), indicating cytoplasmic dephosphorylation of CaMKIIα by the alpha4/PP2Ac complex.","method":"Co-immunoprecipitation, neuron-specific conditional knockout mouse, behavioral assays (water maze, shuttle-box), subcellular fractionation/localization","journal":"Brain research","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional KO with defined behavioral phenotype, Co-IP, subcellular localization, multiple orthogonal approaches","pmids":["16516168"],"is_preprint":false},{"year":2007,"finding":"siRNA-mediated depletion of PP2Ac in pancreatic INS-1 832/13 β-cells markedly attenuated PP2A activity and glucose-stimulated insulin secretion (GSIS). PP2Ac was detected in all subcellular fractions (cytosol > microsomes > secretory granules = nucleus > mitochondria), and its catalytic activity (regulated by carboxylmethylation) is required for GSIS signaling.","method":"siRNA knockdown, phosphatase activity assay, GSIS measurement, subcellular fractionation","journal":"Endocrine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA KD with defined functional readout and subcellular localization, single lab","pmids":["17906371"],"is_preprint":false},{"year":2013,"finding":"PP2Ac suppression (chemical or siRNA) in T-cells resulted in sustained phosphorylation of MEK and ERK after stimulation, increased DNMT enzyme activity, and DNA hypermethylation. Conversely, elevated PP2Ac dephosphorylates MEK/ERK to reduce DNMT1 expression and promote DNA hypomethylation, placing PP2Ac upstream of the MEK/ERK/DNMT1 axis in T-cell epigenetic regulation.","method":"siRNA knockdown, pharmacological inhibition, MEK/ERK phosphorylation assays, DNMT enzyme activity assay, gene expression analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — siRNA KD plus chemical inhibition with multiple functional readouts in both normal and SLE T-cells, multiple orthogonal methods","pmids":["23775084"],"is_preprint":false},{"year":2013,"finding":"MID1 E3 ubiquitin ligase catalyzes in vitro ubiquitination of PP2Ac in the absence of alpha4. In the presence of alpha4, the level of PP2Ac ubiquitination is reduced. Bbox1 domain mutations found in X-linked Opitz syndrome abolish alpha4 polyubiquitination but not PP2Ac ubiquitination, indicating distinct MID1 substrate-binding mechanisms for PP2Ac versus alpha4.","method":"In vitro ubiquitination assay, domain deletion/mutation constructs","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of ubiquitination with mutagenesis, single lab but rigorous biochemical assay","pmids":["25207814"],"is_preprint":false},{"year":2013,"finding":"EDD E3 ubiquitin ligase binds the C-terminus of alpha4 and promotes polyubiquitination and proteasomal degradation of PP2Ac; siEDD knockdown reduced polyubiquitinated PP2Ac species, and progesterone induction of EDD correlated with decreased PP2Ac levels.","method":"Co-immunoprecipitation with alpha4 deletion constructs, siRNA knockdown, proteasomal inhibitor (MG132) treatment, Western blot","journal":"Molecular and cellular endocrinology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP domain mapping and siRNA KD with proteasomal inhibitor confirmation, single lab, two orthogonal methods","pmids":["24145130"],"is_preprint":false},{"year":2014,"finding":"PPP2CA restoration in prostate cancer cells decreased nuclear accumulation and transcriptional activity of β-catenin and NF-κB. Akt mediated PPP2CA loss-induced nuclear accumulation of β-catenin/NF-κB through inactivation of Gsk3-β and IκB-α, respectively. Restitution of β-catenin/NF-κB activity abrogated PPP2CA-induced reversal of EMT.","method":"Stable overexpression/silencing, luciferase promoter-reporter assay, immunoblot, immunofluorescence, orthotopic mouse model","journal":"British journal of cancer","confidence":"High","confidence_rationale":"Tier 2 / Strong — loss- and gain-of-function with epistasis rescue experiments and in vivo validation, multiple orthogonal methods","pmids":["24642616"],"is_preprint":false},{"year":2014,"finding":"Gi-coupled receptor stimulation (A1R, M2, AT2) induces PP2Ac carboxylmethylation at Leu309 in adult rat ventricular myocytes via a Gβγ-PI3K signaling pathway. A1R stimulation increased PP2Ac association with its methyltransferase LCMT-1 and promoted PP2Ac translocation to the particulate fraction; these effects were blocked by PI3K inhibition or Gαt1 expression.","method":"Carboxylmethylation assay, pharmacological receptor stimulation, adenoviral Gαt1 expression, subcellular fractionation, co-immunoprecipitation","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple receptor agonists, genetic (adenoviral) and pharmacological epistasis, subcellular fractionation with functional consequence, multiple orthogonal methods","pmids":["24475092"],"is_preprint":false},{"year":2016,"finding":"TIPRL crystal structure solved at 2.15 Å reveals a conserved cleft that binds the C-terminal tail of PP2Ac. Mutagenesis, pull-down, and hydrogen/deuterium exchange mass spectrometry confirmed TIPRL preferentially binds the unmodified (unmethylated) PP2Ac C-terminal peptide (DYFL) over the tyrosine-phosphorylated version. A docking model suggests TIPRL blocks the phosphatase active site.","method":"Crystal structure determination, mutagenesis, GST pull-down, hydrogen/deuterium exchange mass spectrometry, docking modeling","journal":"Scientific reports","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure at 2.15 Å combined with mutagenesis and HDX-MS, multiple orthogonal methods in one study","pmids":["27489114"],"is_preprint":false},{"year":2016,"finding":"PP2Ac (alpha4/PP2Ac complex) dephosphorylates CaMKIIα in the cytoplasm, regulating its activity and influencing learning/memory. PP2Ac and alpha4 are localized in the cytoplasm and not in the post-synaptic density, indicating compartment-specific regulation of CaMKIIα.","method":"Conditional neuronal knockout, co-immunoprecipitation, subcellular fractionation, overexpression in neuronal cell lines","journal":"Brain research","confidence":"High","confidence_rationale":"Tier 2 / Strong — replicated and extended (2006 paper same group), conditional KO plus localization plus in vitro functional rescue","pmids":["16516168"],"is_preprint":false},{"year":2016,"finding":"PP2Ac promotes Raf-MEK-ERK signaling in endothelial cells; inhibition of PP2Ac by okadaic acid or siRNA depletion led to significant inactivation of Raf-MEK-ERK and reduced glutaminolysis (measured by cellular glutamate levels and KGA expression). TGF-β1-induced glutaminolysis required PP2Ac-dependent Raf-MEK-ERK activation.","method":"Pharmacological inhibition (okadaic acid), siRNA knockdown, MEK inhibitor (U0126), Western blot, glutamate measurement","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA KD and pharmacological inhibition with defined metabolic readout, single lab, two complementary approaches","pmids":["27612201"],"is_preprint":false},{"year":2017,"finding":"DDX3 forms a complex with PP2A-C and IKK-β (identified by co-immunoprecipitation and mass spectrometry). DDX3 modulates PP2A-C activity by controlling phosphorylation of PP2A-C, enabling PP2A-C to dephosphorylate IKK-β and regulate NF-κB signaling; this regulatory effect was independent of DDX3's ATPase or helicase activity.","method":"Co-immunoprecipitation, mass spectrometry, phosphorylation assays, siRNA knockdown","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP/MS identification of complex plus functional phosphorylation assay, single lab","pmids":["28402257"],"is_preprint":false},{"year":2018,"finding":"PP2Ac promotes DNA hypomethylation by dephosphorylating MEK/ERK, reducing DNMT1 expression. De novo PPP2CA mutations in patients with intellectual disability showed mutation-specific biochemical distortions including poor expression, altered A-subunit and B-subunit binding, impaired phosphatase activity, and impaired C-terminal methylation. Four variants caused complete PP2A dysfunction consistent with haploinsufficiency; four others had dominant-negative mechanisms correlating with severe ID. All pathogenic variants impair PP2A-B56δ functionality.","method":"Functional expression assays, phosphatase activity measurement, co-immunoprecipitation (A-subunit and B-subunit binding), methylation assays","journal":"American journal of human genetics","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal biochemical assays on 16 patient variants, phosphatase activity, binding assays, methylation, large cohort","pmids":["30595372"],"is_preprint":false},{"year":2018,"finding":"Glucose deprivation triggers rapid plasma membrane depolarization and Ca2+ influx through Cav1.3, activating CAMK1, which together with PPME1 demethylates and inactivates PP2Ac. PP2Ac demethylation leads to RIPK1 phosphorylation and cell death. Glucose (but not its metabolizable function, since 2-DG also prevented it) prevents PP2Ac demethylation and cell death, identifying glucose as a signaling molecule protecting cells from depolarization-induced PP2Ac inactivation.","method":"Calcium channel pharmacology, CAMK1/PPME1 knockdown, PP2Ac demethylation assay, cell death assays (RIPK1 phosphorylation)","journal":"Science signaling","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple genetic (KD) and pharmacological approaches with orthogonal cell death readouts, mechanistic pathway defined","pmids":["29317521"],"is_preprint":false},{"year":2019,"finding":"LB-100, an experimental antitumor drug, is a catalytic inhibitor of both PP2Ac (PPP2CA) and PPP5C in vitro using purified enzymes. The crystal structure of PPP5C co-crystallized with LB-100 at 1.65 Å resolution revealed that the 7-oxabicyclo[2.2.1]heptane-2,3-dicarbonyl moiety coordinates with metal ions and conserved active-site residues shared by both PP2Ac and PPP5C.","method":"In vitro inhibition assay with purified enzymes, crystal structure determination (1.65 Å), cell-based genetic disruption of PPP5C","journal":"Molecular cancer therapeutics","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution with purified enzymes, crystal structure, and cell-based genetic validation","pmids":["30679389"],"is_preprint":false},{"year":2020,"finding":"CDK1 directly phosphorylates a threonine residue on the PP2A catalytic subunit (PP2Ac), disrupting its holoenzyme formation with the regulatory subunit B55. This consequent decrease in PP2A-B55 substrate dephosphorylation promotes mitotic entry, constituting an additional layer of CDK1-PP2A regulation beyond the previously known indirect mechanism.","method":"Mass spectrometry-based chemical proteomics (PPP holoenzyme enrichment and quantification), kinase profiling, phosphorylation site identification","journal":"Science signaling","confidence":"High","confidence_rationale":"Tier 1 / Strong — quantitative chemical proteomics with kinase profiling identifying direct phosphorylation site and functional consequence on holoenzyme assembly","pmids":["32900880"],"is_preprint":false},{"year":2020,"finding":"PP2Ac phospho-Tyr307 antibodies (clones E155 and F-8) are not specific for phosphorylated Tyr307 but are instead sensitive to other PP2Ac modifications including Leu309 methylation and Thr304 phosphorylation. pTyr307 was identified by targeted mass spectrometry only under overexpression conditions, and none of the tested antibodies showed exclusive pTyr307 specificity, requiring reinterpretation of over 180 prior studies.","method":"Targeted mass spectrometry, antibody specificity testing with phosphomimetic mutants, Western blot","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1 / Strong — replicated by two independent groups (PMIDs 32130916 and 32130915), mass spectrometry and multiple antibody validations","pmids":["32130916","32130915"],"is_preprint":false},{"year":2020,"finding":"Reduction in PP2Ac methylation (caused by ethanol-induced decrease in hepatic methylation capacity) led to increased degradation of the regulatory Bα subunit, promoting phosphorylation and nuclear exclusion of FOXO1, reducing FOXO1 transcriptional activity, and ultimately increasing TXNIP expression and hepatic lipid accumulation.","method":"Betaine supplementation (methyl donor), Western blot, phosphorylation analysis, subcellular fractionation, in vivo mouse model","journal":"Toxicology letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological manipulation of methylation with mechanistic pathway readouts in vitro and in vivo, single lab","pmids":["32492475"],"is_preprint":false},{"year":2021,"finding":"PDCD10 directly binds to PP2Ac and increases its enzymatic activity, leading to YAP dephosphorylation, YAP nuclear translocation and transcriptional activation. Knockdown of PP2Ac abolished PDCD10-mediated HCC cell migration, invasion and EMT; PP2Ac inhibitor LB100 restricted tumor growth and metastasis in HCC with high PDCD10 expression.","method":"Co-immunoprecipitation, phosphatase activity assay, siRNA knockdown, LB100 pharmacological inhibition, in vivo xenograft","journal":"Cell death & disease","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP, enzymatic activity assay, siRNA epistasis, and in vivo validation; multiple orthogonal methods","pmids":["34521817"],"is_preprint":false},{"year":2021,"finding":"Taurine supplementation reduces SAM availability, which is sensed by LCMT-1 and PME-1 to reduce PP2Ac methylation. PP2Ac methylation was found necessary for M1 macrophage polarization including positive regulation of VDAC1 and PINK1, and its activation promotes PINK1-mediated mitophagy for metabolic adaptation to glycolysis during M1 polarization.","method":"Metabolic assays, Western blot for PP2Ac methylation, PINK1/VDAC1 expression, mitophagy flux assays, pharmacological inhibition of PP2Ac methylation (ABL127)","journal":"Frontiers in immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological manipulation with mechanistic pathway readouts, single lab, multiple downstream markers assessed","pmids":["33912173"],"is_preprint":false},{"year":2021,"finding":"TRPC1-mediated Ca2+ signaling increases levels of alpha4 and PP2Ac proteins and promotes alpha4/PP2Ac association in intestinal epithelial cells, enhancing cell migration after wounding. siRNA silencing of either alpha4 or PP2Ac destabilized alpha4/PP2Ac complexes and repressed migration; cellular polyamines regulated this pathway by controlling alpha4/PP2Ac expression and association.","method":"Stable TRPC1 overexpression, siRNA knockdown, co-immunoprecipitation, wound migration assay, polyamine manipulation","journal":"Physiological reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain- and loss-of-function with Co-IP and functional migration readout, single lab","pmids":["33991460"],"is_preprint":false},{"year":2022,"finding":"De novo PPP2CA missense variant p.Cys196Arg causes severe catalytic impairment, mildly reduced A-subunit binding, and moderately decreased binding to B/B55, B\"/PR72, and most B56 subunits (except B56γ1), consistent with partial loss of function. C-terminal methylation and STRN3 binding were unaffected.","method":"In vitro phosphatase activity assay, co-immunoprecipitation for A and B subunit binding, methylation assay","journal":"Frontiers in cell and developmental biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple orthogonal biochemical characterizations of variant (activity, multiple binding partners, methylation) in one rigorous study","pmids":["36531959"],"is_preprint":false},{"year":2023,"finding":"PP2Ac deficiency in glioma cells enhanced dsDNA production and activated cGAS-STING-type I IFN signaling, increased MHC-I expression, and elevated tumor mutational burden. PP2Ac genetic ablation sensitized glioma tumors to immune-checkpoint blockade and radiotherapy, establishing PP2Ac as an inhibitor of cGAS-STING signaling in glioma cells.","method":"Genetic ablation (knockout), cGAS-STING signaling assays, MHC-I expression, dsDNA measurement, co-culture with DCs and T cells, in vivo tumor model with checkpoint blockade/radiotherapy, single-cell analysis","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic KO with multiple orthogonal functional readouts (molecular signaling, co-culture, in vivo), replicated in companion paper (PMID 36757811)","pmids":["37219874"],"is_preprint":false},{"year":2023,"finding":"PP2A with its specific B regulatory subunit STRN4 (forming PP2A/STRN4 holoenzyme) negatively regulates STING-type I IFN signaling in macrophages by dephosphorylating Hippo kinase MST1/2 and stabilizing YAP/TAZ to antagonize STING activation. STING agonists induced dissociation of PP2A from MST1/2 in normal but not in tumor-conditioned macrophages.","method":"Macrophage-specific PP2A conditional knockout, Co-IP (PP2Ac-STRN4-MST1/2 complex), dephosphorylation assay, STING signaling assay, in vivo tumor model","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional KO, biochemical Co-IP showing complex, dephosphorylation assay, in vivo validation; multiple orthogonal methods","pmids":["36757811"],"is_preprint":false},{"year":2023,"finding":"PTEN directly binds to and dephosphorylates the C terminus of PP2Ac to increase its enzymatic activity; elevated PP2Ac activity in turn dephosphorylates deoxycytidine kinase (DCK) at Ser74 to diminish gemcitabine efficacy. PTEN deficiency therefore results in decreased PP2Ac activity and higher DCK phosphorylation, facilitating gemcitabine action.","method":"Direct binding assay, phosphatase activity assay, DCK phosphorylation assay, cell-based drug sensitivity assays, CDX and PDX xenograft models","journal":"Science translational medicine","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct binding and enzymatic dephosphorylation assays combined with in vivo PDX validation and substrate identification","pmids":["37437018"],"is_preprint":false},{"year":2023,"finding":"Luteolin directly binds to KDM4C, blocking KDM4C-induced histone demethylation of the PPP2CA promoter, inhibiting PPP2CA transcription. Reduced PPP2CA expression then impairs PPP2CA-mediated YAP dephosphorylation, thereby attenuating YAP activity and ovarian cancer stem cell stemness.","method":"Direct binding assay (luteolin-KDM4C), ChIP/promoter methylation, luciferase reporter, YAP phosphorylation assay, functional stemness assays","journal":"Biomedicine & pharmacotherapy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding assay plus epigenetic and functional readouts, single lab, multiple methods","pmids":["36804120"],"is_preprint":false},{"year":2024,"finding":"dTAG-mediated selective degradation of PPP2CA in HEK293 cells followed by global phospho-proteomics identified 2,204 proteins with significantly increased phosphorylation, constituting a broad substrate landscape. A pSP/pTP motif was identified as the predominant PPP2CA target sequence. PPP2CA substrates are enriched for functions in spliceosome, cell cycle, RNA transport, and ubiquitin-mediated proteolysis.","method":"dTAG proteolysis-targeting chimera degradation of knock-in dTAG-PPP2CA, unbiased global phospho-proteomics, immunoblotting validation","journal":"iScience","confidence":"High","confidence_rationale":"Tier 1 / Strong — selective protein degradation combined with unbiased global phospho-proteomics and immunoblot validation; rigorous substrate identification","pmids":["38450154"],"is_preprint":false},{"year":2024,"finding":"PKC (classical isoforms PKCα and PKCβ, but not novel PKCδ or PKCε) phosphorylates PP2Ac at Ser24. This phosphorylation is necessary and sufficient to trigger the PP2A switch (involving IGBP1/alpha4), inducing dephosphorylation and inactivation of PI3K and AKT, and leading to JNK-dependent apoptosis upon GqPCR activation.","method":"Phospho-mass spectrometry to identify Ser24 site, specific phospho-antibody generation, S24A and S24E substitution mutants, co-immunoprecipitation, proximity ligation assay, TUNEL apoptosis assay","journal":"Cell communication and signaling : CCS","confidence":"High","confidence_rationale":"Tier 1 / Strong — phospho-MS site identification, phosphomimetic/unphosphorylatable mutants, specific antibody, multiple functional readouts","pmids":["38419089"],"is_preprint":false},{"year":2024,"finding":"PP2Ac regulates ULK1 phosphorylation (dephosphorylation at Ser637) during osteoclastogenesis to control autophagy. mTORC1 inhibition facilitated PP2Ac expression, and PP2Ac-mediated autophagy was dependent on ULK1 phosphorylation activity. Knockdown or inhibition of PP2Ac weakened autophagy during osteoclastogenesis.","method":"siRNA knockdown, phosphorylation assay (ULK1 Ser637), mTORC1 inhibition, in vivo OA rat model","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KD with phosphorylation site analysis and in vivo validation, single lab","pmids":["39041921"],"is_preprint":false},{"year":2025,"finding":"MKRN2 E3 ligase interacts with PPP2CA and promotes K48-linked ubiquitination at PPP2CA's K41 residue, leading to proteasomal degradation of PPP2CA. Consequently, MKRN2-mediated PPP2CA repression increased β-catenin phosphorylation and decreased its levels, inactivating Wnt signaling in clear cell renal cell carcinoma.","method":"Co-immunoprecipitation, ubiquitination assay (K48-linkage, K41 site identification), Western blot, in vivo xenograft","journal":"International journal of biological sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination linkage and site characterization, functional epistasis, single lab","pmids":["40959281"],"is_preprint":false},{"year":2025,"finding":"Sirt2 regulates PP2Ac activation through controlling PP2Ac acetylation at Lys136. Colchicine enhances Sirt2 expression, which deacetylates and activates PP2Ac, leading to increased phosphorylation of NLRP3 at Ser5 and reduced NLRP3 inflammasome activation, thereby inhibiting vascular calcification.","method":"Phosphorylation/acetylation assays, Sirt2 knockout mice, PP2Ac activity assay, vascular calcification models in vitro and in vivo","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KO mouse model with biochemical site identification (Lys136 acetylation, Ser5 phosphorylation), single lab","pmids":["40523615"],"is_preprint":false},{"year":2025,"finding":"Carboxy-methylation of PP2Ac is highly sensitive to intracellular SAM levels. Overexpression of PME-1 (which demethylates PP2A) or expression of a Leu309-deleted unmethylated PP2A mimetic was sufficient to reduce cancer cell proliferation even in methionine-independent cells, establishing a mechanistic link between methionine/SAM availability, PP2Ac methylation status, and cell proliferation.","method":"PME-1 overexpression, Leu309-deleted PP2Ac expression, SAM measurement, cell proliferation assays","journal":"Biomolecules","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain-of-function/methylation-deficient mutant with defined proliferation readout, single lab, two orthogonal approaches","pmids":["41008516"],"is_preprint":false},{"year":2025,"finding":"RCN2 facilitates PPP2CA ubiquitination and proteasomal degradation in a manner dependent on the HECT domain of UBR5 E3 ligase; RCN2 physically interacts with both PPP2CA and UBR5. This PPP2CA degradation activates PI3K-AKT signaling to promote ESCC metastasis and cisplatin resistance.","method":"Co-immunoprecipitation, GST pull-down, Western blot for ubiquitination, RNA-seq, LC-MS/MS, in vivo xenograft","journal":"Drug resistance updates","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, pull-down, ubiquitination assay confirming UBR5 HECT-domain dependence; single lab, multiple methods","pmids":["41411970"],"is_preprint":false},{"year":2025,"finding":"LRRK2 phosphorylates PP2Ac at residue T304 in vitro; this LRRK2-mediated T304 phosphorylation alters C-terminal methylation of PP2Ac, impairing PP2A holoenzyme formation and catalytic activity. Reciprocally, PP2A dephosphorylates sites in the RocCOR-GTPase domain of LRRK2, de-stabilizing LRRK2 dimers and reducing its kinase activity. WT PPP2CA expression protected from LRRK2-G2019S-induced neuronal cell death, while T304 mutants failed to do so.","method":"In vitro dephosphorylation assay (PP2A on LRRK2), kinase assay (LRRK2 on PP2Ac T304), PP2Ac methylation assay, holoenzyme formation assay, neuronal cell death rescue experiment with WT vs. T304 mutant PP2CA","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro biochemical reconstitution of reciprocal phosphorylation, methylation assay, and functional rescue, but preprint not yet peer-reviewed","pmids":["bio_10.1101_2025.10.07.680857"],"is_preprint":true},{"year":2026,"finding":"PP2Ac (Mts in Drosophila) is sufficient to trigger cell migration by activating the JNK signaling pathway. Genetic epistasis analyses in Drosophila showed Mts acts upstream of Slpr (MLK kinase) in the JNK cascade. Affinity purification-mass spectrometry identified Rho1 as a mediator; Mts activates JNK by increasing Rho1 protein levels, and Rho1 acts downstream of Mts/upstream of Slpr-JNK. PP2Ac also promoted cell migration in human pancreatic cancer cells associated with RhoA levels and JNK activation.","method":"Drosophila genetic epistasis (double mutant analysis), affinity purification-mass spectrometry (AP-MS), PP2AC overexpression in human PAAD cells, Western blot for RhoA/JNK","journal":"FASEB journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis in Drosophila plus AP-MS interactor identification plus mammalian cell validation, multiple orthogonal methods","pmids":["42144974"],"is_preprint":false},{"year":2026,"finding":"Spry2 sequesters PP2Ac upon LPS stimulation (Spry2 becomes serine-phosphorylated and associates with PP2Ac), preventing PP2Ac from dephosphorylating p65, thus heightening NF-κB nuclear translocation and cytokine production. In Spry2-deficient macrophages, PP2Ac freely interacts with p65, enhancing its dephosphorylation and reducing its nuclear translocation; PP2Ac inhibitor treatment rescued this defect.","method":"Co-immunoprecipitation, Spry2 knockout macrophages, PP2Ac inhibitor rescue, p65 nuclear translocation assay, cytokine measurement","journal":"Journal of immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, KO macrophages with pharmacological rescue, nuclear translocation assay, single lab","pmids":["41847846"],"is_preprint":false}],"current_model":"PPP2CA encodes the principal catalytic subunit of PP2A (PP2Ac), a serine/threonine phosphatase whose activity and substrate specificity are governed by reversible C-terminal Leu309 carboxylmethylation (written by LCMT-1, erased by PME-1) and direct phosphorylation at Thr304 (by LRRK2), Ser24 (by classical PKCs), and Thr (by CDK1), with these modifications controlling holoenzyme assembly with scaffolding (A) and regulatory (B/B'/B''/B''') subunits; PP2Ac dephosphorylates a broad pSP/pTP-motif substrate landscape (>2,000 proteins including MEK/ERK, AKT, CaMKIIα, YAP, MST1/2, RIPK1, DCK, and IKK) to regulate cell cycle entry/exit, apoptosis, immune signaling (MEK-ERK-DNMT1 axis, cGAS-STING, NF-κB), metabolic reprogramming, autophagy, and cell migration (via Rho1-JNK), and is subject to ubiquitin-mediated degradation by MID1, EDD, UBR5, and MKRN2 E3 ligases."},"narrative":{"mechanistic_narrative":"PPP2CA encodes the catalytic subunit of protein phosphatase 2A (PP2Ac), a serine/threonine phosphatase that dephosphorylates a broad pSP/pTP-motif substrate landscape of >2,000 proteins enriched for cell cycle, RNA processing, and ubiquitin-proteolysis functions to govern proliferation, apoptosis, immune signaling, and cell migration [PMID:38450154, PMID:30595372]. PP2Ac assembles with scaffolding (A) and regulatory (B/B'/B''/STRN) subunits into functional holoenzymes; the conserved C-terminal Leu309 region and an intact catalytic core are required for productive B-subunit binding and activity, and active-site or truncation mutants act dominant-negatively by titrating regulatory subunits and substrates [PMID:10446173, PMID:11129046, PMID:36531959]. Holoenzyme assembly and activity are tuned by reversible C-terminal carboxylmethylation at Leu309—written by LCMT-1, erased by PME-1, and sensitive to intracellular SAM/methionine availability—which controls B-subunit recruitment, proliferation, and downstream programs [PMID:24475092, PMID:41008516, PMID:32492475]; additional post-translational control comes from direct phosphorylation by CDK1, which disrupts PP2A-B55 to promote mitotic entry [PMID:32900880], by classical PKCα/β at Ser24, which triggers an IGBP1/alpha4-dependent switch that inactivates PI3K/AKT and drives JNK-dependent apoptosis [PMID:38419089], and from acetylation at Lys136 controlled by Sirt2 [PMID:40523615]. Through defined substrates PP2Ac executes its biology: it dephosphorylates MEK/ERK to suppress the DNMT1 epigenetic axis [PMID:23775084], MST1/2 and YAP to control Hippo signaling, migration and inflammasome output [PMID:34521817, PMID:36757811], RIPK1 in glucose-deprivation cell death [PMID:29317521], IKKβ and p65 in NF-κB signaling [PMID:28402257, PMID:41847846], DCK to modulate gemcitabine sensitivity [PMID:37437018], ULK1 in autophagy [PMID:39041921], and acts via Rho1/RhoA-JNK to drive cell migration [PMID:42144974]; it also restrains cGAS-STING type-I interferon responses in tumor cells [PMID:37219874]. The alpha4 (IGBP1) chaperone binds PP2Ac through its catalytic domain and protects it from ubiquitin-mediated degradation, while the E3 ligases MID1, EDD, UBR5, and MKRN2 target PP2Ac for proteasomal turnover [PMID:25207814, PMID:24145130, PMID:40959281, PMID:41411970, PMID:12963337]. De novo PPP2CA missense and loss-of-function variants cause intellectual disability through haploinsufficient or dominant-negative impairment of phosphatase activity, A/B-subunit binding, and C-terminal methylation [PMID:30595372, PMID:36531959].","teleology":[{"year":1999,"claim":"Established that human PP2Ac is a functionally conserved catalytic subunit whose activity and proper complex assembly are essential, defining the dominant-negative behavior of catalytically dead forms.","evidence":"Yeast complementation, mutagenesis and in vitro phosphatase assays with active-site and C-terminal mutants","pmids":["10446173"],"confidence":"High","gaps":["Did not map all regulatory subunit contacts in human holoenzymes","Leu309 modification chemistry not yet addressed"]},{"year":2000,"claim":"Linked the conserved C-terminal leucine to selective B-subunit binding and mitotic checkpoint function, showing modification of this residue differentially gates holoenzyme composition.","evidence":"Yeast genetics, two-hybrid binding, and microtubule-drug phenotyping of Pph22p Leu mutants","pmids":["11129046"],"confidence":"High","gaps":["B-subunit selectivity demonstrated in yeast orthologs, not human subunits directly","Methylation status not measured"]},{"year":2004,"claim":"Defined the region of the scaffolding A subunit required for physical contact with PP2Ac, anchoring the architecture of the core dimer.","evidence":"Co-IP of PPP2R1B deletion/missense mutants from colorectal cancer tissue","pmids":["14767517"],"confidence":"Medium","gaps":["Single method, no structural validation","Functional consequence of disrupted binding not assayed"]},{"year":2006,"claim":"Identified the alpha4/PP2Ac complex as a cytoplasmic regulator of CaMKIIα, connecting the phosphatase to learning and memory through compartment-specific substrate dephosphorylation.","evidence":"Neuron-specific alpha4 conditional KO, Co-IP, fractionation and behavioral assays","pmids":["16516168"],"confidence":"High","gaps":["Effects attributed via alpha4 KO rather than direct PP2Ac perturbation","Direct CaMKIIα dephosphorylation site not mapped"]},{"year":2005,"claim":"Mapped distinct binding surfaces on PP2Ac for alpha4 and substrate S6K1, showing alpha4 bridges PP2Ac to substrates under immune stimulation.","evidence":"Pull-down with PP2Ac deletion constructs and LPS-stimulated B cell Co-IP","pmids":["15796902"],"confidence":"Medium","gaps":["S6K1 dephosphorylation not directly demonstrated","Single lab domain mapping"]},{"year":2013,"claim":"Placed PP2Ac upstream of the MEK/ERK/DNMT1 axis, defining a route by which the phosphatase controls DNA methylation in T cells.","evidence":"siRNA and chemical inhibition with phospho-ERK, DNMT activity and methylation readouts","pmids":["23775084"],"confidence":"High","gaps":["Direct MEK/ERK dephosphorylation by PP2Ac vs indirect not resolved","Relevant holoenzyme/B-subunit unspecified"]},{"year":2013,"claim":"Identified MID1, EDD as E3 ligases that ubiquitinate PP2Ac and established alpha4 as a protective chaperone modulating PP2Ac turnover.","evidence":"In vitro ubiquitination, alpha4 deletion-construct Co-IP, siRNA and MG132 treatment","pmids":["25207814","24145130"],"confidence":"High","gaps":["Ubiquitination site(s) on PP2Ac not mapped in these studies","Physiological triggers of degradation partly inferred"]},{"year":2014,"claim":"Demonstrated receptor- and PI3K-controlled Leu309 carboxylmethylation governs PP2Ac localization and showed PP2Ac suppresses β-catenin/NF-κB via Akt-GSK3β/IκBα, establishing methylation and signaling control of the phosphatase.","evidence":"Carboxylmethylation assays with Gi-receptor agonists; gain/loss-of-function with reporter and orthotopic models in prostate cancer","pmids":["24475092","24642616"],"confidence":"High","gaps":["Whether methylation directly dictates specific B-subunit recruitment in these contexts untested","Direct substrate of PP2Ac in β-catenin/NF-κB axis not identified"]},{"year":2016,"claim":"Provided structural and biochemical basis for C-terminal-tail recognition of PP2Ac by the regulator TIPRL, which preferentially binds unmethylated PP2Ac and occludes the active site.","evidence":"TIPRL crystal structure at 2.15 Å, mutagenesis, pull-down, HDX-MS and docking","pmids":["27489114"],"confidence":"High","gaps":["Cellular consequence of TIPRL-mediated active-site blockade not quantified here","No structure of full PP2Ac-TIPRL complex"]},{"year":2018,"claim":"Established PPP2CA as an intellectual-disability gene and dissected how de novo variants impair activity, subunit binding and methylation by haploinsufficient or dominant-negative mechanisms.","evidence":"Functional expression, phosphatase activity, A/B-subunit Co-IP and methylation assays across patient variants","pmids":["30595372"],"confidence":"High","gaps":["In vivo neurodevelopmental mechanism not modeled","Substrate-level consequences in neurons not defined"]},{"year":2018,"claim":"Linked glucose-sensing through Cav1.3/CAMK1/PPME1 to PP2Ac demethylation and RIPK1-dependent cell death, defining a metabolic control of PP2Ac inactivation.","evidence":"Channel pharmacology, CAMK1/PPME1 knockdown and demethylation/cell-death assays","pmids":["29317521"],"confidence":"High","gaps":["Direct RIPK1 dephosphorylation by PP2Ac not formally shown","B-subunit specifying RIPK1 targeting unknown"]},{"year":2020,"claim":"Showed CDK1 directly phosphorylates a PP2Ac threonine to disrupt PP2A-B55 and promote mitotic entry, adding a direct layer to CDK1-PP2A crosstalk.","evidence":"Quantitative chemical proteomics with holoenzyme enrichment and kinase profiling","pmids":["32900880"],"confidence":"High","gaps":["Precise threonine residue and its structural effect not fully resolved","In vivo cell-cycle requirement not tested"]},{"year":2020,"claim":"Demonstrated that widely used phospho-Tyr307 antibodies are non-specific and instead report Leu309 methylation and Thr304 phosphorylation, forcing reinterpretation of prior PP2Ac regulatory studies.","evidence":"Targeted mass spectrometry and antibody testing with phosphomimetic mutants, replicated by two groups","pmids":["32130916","32130915"],"confidence":"High","gaps":["Physiological role of genuine pTyr307 remains undefined","Does not establish the true functional reader of these modifications"]},{"year":2021,"claim":"Connected PP2Ac to Hippo/YAP signaling and metabolism by showing PDCD10-stimulated PP2Ac dephosphorylates YAP to drive tumor migration, and methylation-dependent PP2Ac regulates macrophage mitophagy.","evidence":"Co-IP, phosphatase activity, siRNA epistasis and LB100 with xenografts; methylation manipulation with mitophagy readouts","pmids":["34521817","33912173"],"confidence":"High","gaps":["Direct YAP dephosphorylation site not mapped","B-subunit directing YAP/MST targeting unspecified"]},{"year":2023,"claim":"Defined PP2Ac as a brake on innate antitumor immunity, restraining cGAS-STING type-I IFN signaling in glioma and, via the STRN4 holoenzyme, dephosphorylating MST1/2 to antagonize STING in macrophages.","evidence":"Genetic ablation/conditional KO, cGAS-STING and dephosphorylation assays, co-culture and in vivo tumor models","pmids":["37219874","36757811"],"confidence":"High","gaps":["Direct STING-pathway substrate(s) in glioma not pinpointed","Generality across tumor types not established"]},{"year":2023,"claim":"Identified PTEN as a direct PP2Ac activator that dephosphorylates its C-terminus, and DCK Ser74 as a substrate controlling gemcitabine efficacy, linking PP2Ac to chemotherapy response.","evidence":"Direct binding, phosphatase and DCK phosphorylation assays with CDX/PDX models","pmids":["37437018"],"confidence":"High","gaps":["Mechanism by which C-terminal dephosphorylation raises activity not structurally defined","Phosphatase activity of PTEN on PP2Ac vs scaffolding role"]},{"year":2024,"claim":"Provided an unbiased, system-wide definition of the PP2Ac substrate landscape and its pSP/pTP target motif, anchoring its broad regulatory reach.","evidence":"dTAG-mediated PPP2CA degradation with global phospho-proteomics","pmids":["38450154"],"confidence":"High","gaps":["Holoenzyme/B-subunit assignment per substrate not resolved","Direct vs indirect substrates not fully distinguished"]},{"year":2024,"claim":"Identified PKCα/β phosphorylation of PP2Ac at Ser24 as the trigger of an IGBP1/alpha4-dependent PP2A switch that inactivates PI3K/AKT and drives JNK-dependent apoptosis.","evidence":"Phospho-MS, S24A/S24E mutants, specific antibody, Co-IP, PLA and TUNEL","pmids":["38419089"],"confidence":"High","gaps":["Structural basis of the alpha4-mediated switch not resolved","Generality beyond GqPCR stimulation untested"]},{"year":2025,"claim":"Expanded the PP2Ac degradation network and modification control, identifying MKRN2 (K48/K41) and RCN2-UBR5 as degradation routes and Sirt2-controlled Lys136 acetylation as an activity switch.","evidence":"Co-IP, ubiquitination linkage/site mapping, acetylation and activity assays with KO mice and xenografts","pmids":["40959281","41411970","40523615"],"confidence":"Medium","gaps":["Each E3-PP2Ac axis from single labs without cross-validation","Relative contribution of competing E3 ligases in vivo unclear"]},{"year":2025,"claim":"Established that PP2Ac carboxymethylation is tightly coupled to SAM/methionine availability and that demethylated PP2Ac suppresses cancer proliferation, linking one-carbon metabolism to PP2A activity.","evidence":"PME-1 overexpression, Leu309-deletion mimetic, SAM measurement and proliferation assays","pmids":["41008516"],"confidence":"Medium","gaps":["Downstream substrate program of demethylated PP2Ac not defined","Single-lab gain-of-function evidence"]},{"year":2026,"claim":"Defined a PP2Ac-Rho1/RhoA-JNK migration axis and a Spry2-mediated sequestration mechanism controlling p65 dephosphorylation in NF-κB signaling.","evidence":"Drosophila genetic epistasis with AP-MS plus mammalian validation; Spry2-KO macrophages with Co-IP and inhibitor rescue","pmids":["42144974","41847846"],"confidence":"High","gaps":["Direct Rho1/RhoA substrate relationship vs level control not fully resolved","Spry2 mechanism single-lab"]},{"year":2026,"claim":"Proposed reciprocal regulation between LRRK2 and PP2Ac in a neuronal context, with LRRK2 phosphorylating PP2Ac at Thr304 and PP2Ac dephosphorylating LRRK2.","evidence":"In vitro kinase/phosphatase and methylation assays with neuronal cell-death rescue (preprint)","pmids":["bio_10.1101_2025.10.07.680857"],"confidence":"Medium","gaps":["Preprint, not peer-reviewed","Thr304-methylation crosstalk not validated in vivo"]},{"year":null,"claim":"How specific holoenzyme B-subunit compositions are matched to the >2,000 substrate landscape, and how the layered modification code (methylation, Ser24/Thr304/Thr phosphorylation, Lys136 acetylation) is integrated to dictate substrate choice, remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No comprehensive map linking individual B-subunits to defined substrates","Combinatorial effect of co-occurring PP2Ac modifications not deconvolved","Structures of substrate-engaged human holoenzymes lacking"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,30]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[7,28,30,27]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[19,31]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[5,6,13]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[6,10]}],"pathway":[{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[19,1]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[7,22,31,38]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[26,27,39]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[17,31]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[32,23]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[8,9,33,36]}],"complexes":["PP2A holoenzyme","PP2A-B55","PP2A-B56","PP2A/STRN4"],"partners":["PPP2R1B","IGBP1","TIPRL","PDCD10","PTEN","MID1","UBR5","MKRN2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P67775","full_name":"Serine/threonine-protein phosphatase 2A catalytic subunit alpha isoform","aliases":["Replication protein C","RP-C"],"length_aa":309,"mass_kda":35.6,"function":"Catalytic subunit of protein phosphatase 2A (PP2A), a serine/threonine phosphatase involved in the regulation of a wide variety of enzymes, signal transduction pathways, and cellular events (PubMed:10801873, PubMed:12473674, PubMed:17245430, PubMed:22613722, PubMed:33243860, PubMed:34004147, PubMed:9920888). PP2A is the major phosphatase for microtubule-associated proteins (MAPs) (PubMed:22613722). PP2A can modulate the activity of phosphorylase B kinase casein kinase 2, mitogen-stimulated S6 kinase, and MAP-2 kinase (PubMed:22613722). Cooperates with SGO2 to protect centromeric cohesin from separase-mediated cleavage in oocytes specifically during meiosis I (By similarity). Can dephosphorylate various proteins, such as SV40 large T antigen, AXIN1, p53/TP53, PIM3, WEE1 (PubMed:10801873, PubMed:12473674, PubMed:17245430, PubMed:9920888). Activates RAF1 by dephosphorylating it at 'Ser-259' (PubMed:10801873). Mediates dephosphorylation of WEE1, preventing its ubiquitin-mediated proteolysis, increasing WEE1 protein levels, and promoting the G2/M checkpoint (PubMed:33108758). Mediates dephosphorylation of MYC; promoting its ubiquitin-mediated proteolysis: interaction with AMBRA1 enhances interaction between PPP2CA and MYC (PubMed:25438055). Mediates dephosphorylation of FOXO3; promoting its stabilization: interaction with AMBRA1 enhances interaction between PPP2CA and FOXO3 (PubMed:30513302). Catalyzes dephosphorylation of the pyrin domain of NLRP3, promoting assembly of the NLRP3 inflammasome (By similarity). Together with RACK1 adapter, mediates dephosphorylation of AKT1 at 'Ser-473', preventing AKT1 activation and AKT-mTOR signaling pathway (By similarity). Dephosphorylation of AKT1 is essential for regulatory T-cells (Treg) homeostasis and stability (By similarity). Catalyzes dephosphorylation of PIM3, promoting PIM3 ubiquitination and proteasomal degradation (PubMed:12473674). Part of the striatin-interacting phosphatase and kinase (STRIPAK) complexes (PubMed:33633399). STRIPAK complexes have critical roles in protein (de)phosphorylation and are regulators of multiple signaling pathways including Hippo, MAPK, nuclear receptor and cytoskeleton remodeling (PubMed:33633399). Different types of STRIPAK complexes are involved in a variety of biological processes such as cell growth, differentiation, apoptosis, metabolism and immune regulation (PubMed:33633399). Key mediator of a quality checkpoint during transcription elongation as part of the Integrator-PP2A (INTAC) complex (PubMed:33243860, PubMed:34004147, PubMed:37080207). The INTAC complex drives premature transcription termination of transcripts that are unfavorably configured for transcriptional elongation: within the INTAC complex, PPP2CA catalyzes dephosphorylation of the C-terminal domain (CTD) of Pol II subunit POLR2A/RPB1 and SUPT5H/SPT5, thereby preventing transcriptional elongation (PubMed:33243860, PubMed:34004147, PubMed:37080207)","subcellular_location":"Cytoplasm; Nucleus; Chromosome; Chromosome, centromere; Cytoplasm, cytoskeleton, spindle pole","url":"https://www.uniprot.org/uniprotkb/P67775/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/PPP2CA","classification":"Common Essential","n_dependent_lines":1047,"n_total_lines":1208,"dependency_fraction":0.8667218543046358},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000113575","cell_line_id":"CID000214","localizations":[{"compartment":"centrosome","grade":3},{"compartment":"cytoplasmic","grade":2},{"compartment":"nucleoplasm","grade":2},{"compartment":"membrane","grade":1}],"interactors":[{"gene":"CCT3","stoichiometry":10.0},{"gene":"CCT5","stoichiometry":10.0},{"gene":"CCT6A","stoichiometry":10.0},{"gene":"CCT4","stoichiometry":10.0},{"gene":"CCT8","stoichiometry":10.0},{"gene":"TCP1","stoichiometry":10.0},{"gene":"PPP2R1B","stoichiometry":10.0},{"gene":"PPP2R1A","stoichiometry":10.0},{"gene":"PDCD10","stoichiometry":10.0},{"gene":"CCT7","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/target/CID000214","total_profiled":1310},"omim":[{"mim_id":"618785","title":"CELL DIVISION CYCLE-ASSOCIATED PROTEIN 2; CDCA2","url":"https://www.omim.org/entry/618785"},{"mim_id":"618354","title":"HOUGE-JANSSENS SYNDROME 3; HJS3","url":"https://www.omim.org/entry/618354"},{"mim_id":"617249","title":"FAMILY WITH SEQUENCE SIMILARITY 122, MEMBER A; FAM122A","url":"https://www.omim.org/entry/617249"},{"mim_id":"616759","title":"NITRIC OXIDE SYNTHASE-INTERACTING PROTEIN; NOSIP","url":"https://www.omim.org/entry/616759"},{"mim_id":"616355","title":"HOUGE-JANSSENS SYNDROME 1; HJS1","url":"https://www.omim.org/entry/616355"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/PPP2CA"},"hgnc":{"alias_symbol":["PP2Calpha","PP2AC"],"prev_symbol":[]},"alphafold":{"accession":"P67775","domains":[{"cath_id":"3.60.21.10","chopping":"43-89_113-289","consensus_level":"high","plddt":98.0595,"start":43,"end":289}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P67775","model_url":"https://alphafold.ebi.ac.uk/files/AF-P67775-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P67775-F1-predicted_aligned_error_v6.png","plddt_mean":95.06},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PPP2CA","jax_strain_url":"https://www.jax.org/strain/search?query=PPP2CA"},"sequence":{"accession":"P67775","fasta_url":"https://rest.uniprot.org/uniprotkb/P67775.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P67775/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P67775"}},"corpus_meta":[{"pmid":"23775084","id":"PMC_23775084","title":"The catalytic 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The invariant C-terminal leucine residue (Leu-309) is dispensable for basic function but truncated or active-site mutant forms of PP2Ac exert dominant-negative effects by titrating regulatory subunits/substrates into non-productive complexes; the L199P active-site mutant was catalytically impaired despite binding Tpd3p.\",\n      \"method\": \"Yeast complementation assay, mutagenesis, in vitro phosphatase activity assay, computer modeling of active site\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution in yeast, mutagenesis, in vitro activity assay, multiple orthogonal methods in a single rigorous study\",\n      \"pmids\": [\"10446173\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"The C-terminal Leu-377 residue of yeast Pph22p (equivalent to human PP2Ac Leu-309) is required for binding of the PR55/B subunit (Cdc55p) but not the PR61/B' subunit (Rts1p). Mutation of this leucine enhanced sensitivity to microtubule destabilization, phenocopying cdc55Δ cells with impaired spindle checkpoint function, linking PP2Ac C-terminal leucine to mitotic checkpoint regulation via B-subunit binding.\",\n      \"method\": \"Yeast genetics, in vitro phosphatase activity assay, yeast two-hybrid/binding assay, phenotypic analysis with microtubule-destabilizing drugs\",\n      \"journal\": \"Molecular & general genetics : MGG\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — mutagenesis combined with functional assay and genetic epistasis, replicated mechanistic insight from companion paper\",\n      \"pmids\": [\"11129046\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Purified PP2Ac (but not PP4c or PP6c) bound to GST-alpha4 in pull-down assays and co-immunoprecipitated with endogenous and ectopic myc-tagged alpha4. The alpha4/PP2Ac interaction was disrupted by okadaic acid and microcystin-LR (phosphatase inhibitors) but was unaffected by rapamycin, implying the alpha4 binding site on PP2Ac includes the phosphatase catalytic domain.\",\n      \"method\": \"GST pull-down, co-immunoprecipitation, microcystin-Sepharose affinity purification, pharmacological inhibition\",\n      \"journal\": \"Protein expression and purification\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP and pull-down with purified proteins, single lab, two orthogonal methods\",\n      \"pmids\": [\"12963337\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Missense mutations and deletions within the PP2A-Aβ subunit (PPP2R1B) at amino acids 412–601 (the PP2Ac-binding region) inhibited co-immunoprecipitation of PP2A-Aβ and PP2Ac, demonstrating that this region of the A subunit is required for physical interaction with the catalytic subunit.\",\n      \"method\": \"Co-immunoprecipitation from colorectal cancer tissues, sequence analysis\",\n      \"journal\": \"Oncology reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP from patient tissues, replicated across multiple cancer samples but single lab, single method\",\n      \"pmids\": [\"14767517\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Alpha4 interacts with S6K1 through PP2Ac; pull-down assays defined that S6K1 binds the region from aa 88–309 of PP2Ac, while alpha4 binds two separated regions (aa 19–22 and aa 150–164) of PP2Ac. Stimulation of B cells with LPS induced the interaction of alpha4/PP2Ac with S6K1, suggesting alpha4 regulates S6K1 activity through PP2Ac.\",\n      \"method\": \"Co-immunoprecipitation, pull-down assay with deletion constructs, LPS stimulation of primary B cells\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain mapping by pull-down and Co-IP, single lab, two orthogonal methods\",\n      \"pmids\": [\"15796902\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"CaMKIIα associates with PP2Ac and alpha4 in brain neurons. Neuron-specific alpha4 knockout mice showed increased CaMKIIα activity in hippocampus and impaired learning/memory. Alpha4 and PP2Ac were localized in the cytoplasm (not in the PSD), indicating cytoplasmic dephosphorylation of CaMKIIα by the alpha4/PP2Ac complex.\",\n      \"method\": \"Co-immunoprecipitation, neuron-specific conditional knockout mouse, behavioral assays (water maze, shuttle-box), subcellular fractionation/localization\",\n      \"journal\": \"Brain research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional KO with defined behavioral phenotype, Co-IP, subcellular localization, multiple orthogonal approaches\",\n      \"pmids\": [\"16516168\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"siRNA-mediated depletion of PP2Ac in pancreatic INS-1 832/13 β-cells markedly attenuated PP2A activity and glucose-stimulated insulin secretion (GSIS). PP2Ac was detected in all subcellular fractions (cytosol > microsomes > secretory granules = nucleus > mitochondria), and its catalytic activity (regulated by carboxylmethylation) is required for GSIS signaling.\",\n      \"method\": \"siRNA knockdown, phosphatase activity assay, GSIS measurement, subcellular fractionation\",\n      \"journal\": \"Endocrine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA KD with defined functional readout and subcellular localization, single lab\",\n      \"pmids\": [\"17906371\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"PP2Ac suppression (chemical or siRNA) in T-cells resulted in sustained phosphorylation of MEK and ERK after stimulation, increased DNMT enzyme activity, and DNA hypermethylation. Conversely, elevated PP2Ac dephosphorylates MEK/ERK to reduce DNMT1 expression and promote DNA hypomethylation, placing PP2Ac upstream of the MEK/ERK/DNMT1 axis in T-cell epigenetic regulation.\",\n      \"method\": \"siRNA knockdown, pharmacological inhibition, MEK/ERK phosphorylation assays, DNMT enzyme activity assay, gene expression analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — siRNA KD plus chemical inhibition with multiple functional readouts in both normal and SLE T-cells, multiple orthogonal methods\",\n      \"pmids\": [\"23775084\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"MID1 E3 ubiquitin ligase catalyzes in vitro ubiquitination of PP2Ac in the absence of alpha4. In the presence of alpha4, the level of PP2Ac ubiquitination is reduced. Bbox1 domain mutations found in X-linked Opitz syndrome abolish alpha4 polyubiquitination but not PP2Ac ubiquitination, indicating distinct MID1 substrate-binding mechanisms for PP2Ac versus alpha4.\",\n      \"method\": \"In vitro ubiquitination assay, domain deletion/mutation constructs\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of ubiquitination with mutagenesis, single lab but rigorous biochemical assay\",\n      \"pmids\": [\"25207814\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"EDD E3 ubiquitin ligase binds the C-terminus of alpha4 and promotes polyubiquitination and proteasomal degradation of PP2Ac; siEDD knockdown reduced polyubiquitinated PP2Ac species, and progesterone induction of EDD correlated with decreased PP2Ac levels.\",\n      \"method\": \"Co-immunoprecipitation with alpha4 deletion constructs, siRNA knockdown, proteasomal inhibitor (MG132) treatment, Western blot\",\n      \"journal\": \"Molecular and cellular endocrinology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP domain mapping and siRNA KD with proteasomal inhibitor confirmation, single lab, two orthogonal methods\",\n      \"pmids\": [\"24145130\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"PPP2CA restoration in prostate cancer cells decreased nuclear accumulation and transcriptional activity of β-catenin and NF-κB. Akt mediated PPP2CA loss-induced nuclear accumulation of β-catenin/NF-κB through inactivation of Gsk3-β and IκB-α, respectively. Restitution of β-catenin/NF-κB activity abrogated PPP2CA-induced reversal of EMT.\",\n      \"method\": \"Stable overexpression/silencing, luciferase promoter-reporter assay, immunoblot, immunofluorescence, orthotopic mouse model\",\n      \"journal\": \"British journal of cancer\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — loss- and gain-of-function with epistasis rescue experiments and in vivo validation, multiple orthogonal methods\",\n      \"pmids\": [\"24642616\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Gi-coupled receptor stimulation (A1R, M2, AT2) induces PP2Ac carboxylmethylation at Leu309 in adult rat ventricular myocytes via a Gβγ-PI3K signaling pathway. A1R stimulation increased PP2Ac association with its methyltransferase LCMT-1 and promoted PP2Ac translocation to the particulate fraction; these effects were blocked by PI3K inhibition or Gαt1 expression.\",\n      \"method\": \"Carboxylmethylation assay, pharmacological receptor stimulation, adenoviral Gαt1 expression, subcellular fractionation, co-immunoprecipitation\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple receptor agonists, genetic (adenoviral) and pharmacological epistasis, subcellular fractionation with functional consequence, multiple orthogonal methods\",\n      \"pmids\": [\"24475092\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"TIPRL crystal structure solved at 2.15 Å reveals a conserved cleft that binds the C-terminal tail of PP2Ac. Mutagenesis, pull-down, and hydrogen/deuterium exchange mass spectrometry confirmed TIPRL preferentially binds the unmodified (unmethylated) PP2Ac C-terminal peptide (DYFL) over the tyrosine-phosphorylated version. A docking model suggests TIPRL blocks the phosphatase active site.\",\n      \"method\": \"Crystal structure determination, mutagenesis, GST pull-down, hydrogen/deuterium exchange mass spectrometry, docking modeling\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure at 2.15 Å combined with mutagenesis and HDX-MS, multiple orthogonal methods in one study\",\n      \"pmids\": [\"27489114\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"PP2Ac (alpha4/PP2Ac complex) dephosphorylates CaMKIIα in the cytoplasm, regulating its activity and influencing learning/memory. PP2Ac and alpha4 are localized in the cytoplasm and not in the post-synaptic density, indicating compartment-specific regulation of CaMKIIα.\",\n      \"method\": \"Conditional neuronal knockout, co-immunoprecipitation, subcellular fractionation, overexpression in neuronal cell lines\",\n      \"journal\": \"Brain research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — replicated and extended (2006 paper same group), conditional KO plus localization plus in vitro functional rescue\",\n      \"pmids\": [\"16516168\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"PP2Ac promotes Raf-MEK-ERK signaling in endothelial cells; inhibition of PP2Ac by okadaic acid or siRNA depletion led to significant inactivation of Raf-MEK-ERK and reduced glutaminolysis (measured by cellular glutamate levels and KGA expression). TGF-β1-induced glutaminolysis required PP2Ac-dependent Raf-MEK-ERK activation.\",\n      \"method\": \"Pharmacological inhibition (okadaic acid), siRNA knockdown, MEK inhibitor (U0126), Western blot, glutamate measurement\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA KD and pharmacological inhibition with defined metabolic readout, single lab, two complementary approaches\",\n      \"pmids\": [\"27612201\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"DDX3 forms a complex with PP2A-C and IKK-β (identified by co-immunoprecipitation and mass spectrometry). DDX3 modulates PP2A-C activity by controlling phosphorylation of PP2A-C, enabling PP2A-C to dephosphorylate IKK-β and regulate NF-κB signaling; this regulatory effect was independent of DDX3's ATPase or helicase activity.\",\n      \"method\": \"Co-immunoprecipitation, mass spectrometry, phosphorylation assays, siRNA knockdown\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP/MS identification of complex plus functional phosphorylation assay, single lab\",\n      \"pmids\": [\"28402257\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"PP2Ac promotes DNA hypomethylation by dephosphorylating MEK/ERK, reducing DNMT1 expression. De novo PPP2CA mutations in patients with intellectual disability showed mutation-specific biochemical distortions including poor expression, altered A-subunit and B-subunit binding, impaired phosphatase activity, and impaired C-terminal methylation. Four variants caused complete PP2A dysfunction consistent with haploinsufficiency; four others had dominant-negative mechanisms correlating with severe ID. All pathogenic variants impair PP2A-B56δ functionality.\",\n      \"method\": \"Functional expression assays, phosphatase activity measurement, co-immunoprecipitation (A-subunit and B-subunit binding), methylation assays\",\n      \"journal\": \"American journal of human genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal biochemical assays on 16 patient variants, phosphatase activity, binding assays, methylation, large cohort\",\n      \"pmids\": [\"30595372\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Glucose deprivation triggers rapid plasma membrane depolarization and Ca2+ influx through Cav1.3, activating CAMK1, which together with PPME1 demethylates and inactivates PP2Ac. PP2Ac demethylation leads to RIPK1 phosphorylation and cell death. Glucose (but not its metabolizable function, since 2-DG also prevented it) prevents PP2Ac demethylation and cell death, identifying glucose as a signaling molecule protecting cells from depolarization-induced PP2Ac inactivation.\",\n      \"method\": \"Calcium channel pharmacology, CAMK1/PPME1 knockdown, PP2Ac demethylation assay, cell death assays (RIPK1 phosphorylation)\",\n      \"journal\": \"Science signaling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple genetic (KD) and pharmacological approaches with orthogonal cell death readouts, mechanistic pathway defined\",\n      \"pmids\": [\"29317521\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"LB-100, an experimental antitumor drug, is a catalytic inhibitor of both PP2Ac (PPP2CA) and PPP5C in vitro using purified enzymes. The crystal structure of PPP5C co-crystallized with LB-100 at 1.65 Å resolution revealed that the 7-oxabicyclo[2.2.1]heptane-2,3-dicarbonyl moiety coordinates with metal ions and conserved active-site residues shared by both PP2Ac and PPP5C.\",\n      \"method\": \"In vitro inhibition assay with purified enzymes, crystal structure determination (1.65 Å), cell-based genetic disruption of PPP5C\",\n      \"journal\": \"Molecular cancer therapeutics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution with purified enzymes, crystal structure, and cell-based genetic validation\",\n      \"pmids\": [\"30679389\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"CDK1 directly phosphorylates a threonine residue on the PP2A catalytic subunit (PP2Ac), disrupting its holoenzyme formation with the regulatory subunit B55. This consequent decrease in PP2A-B55 substrate dephosphorylation promotes mitotic entry, constituting an additional layer of CDK1-PP2A regulation beyond the previously known indirect mechanism.\",\n      \"method\": \"Mass spectrometry-based chemical proteomics (PPP holoenzyme enrichment and quantification), kinase profiling, phosphorylation site identification\",\n      \"journal\": \"Science signaling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — quantitative chemical proteomics with kinase profiling identifying direct phosphorylation site and functional consequence on holoenzyme assembly\",\n      \"pmids\": [\"32900880\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"PP2Ac phospho-Tyr307 antibodies (clones E155 and F-8) are not specific for phosphorylated Tyr307 but are instead sensitive to other PP2Ac modifications including Leu309 methylation and Thr304 phosphorylation. pTyr307 was identified by targeted mass spectrometry only under overexpression conditions, and none of the tested antibodies showed exclusive pTyr307 specificity, requiring reinterpretation of over 180 prior studies.\",\n      \"method\": \"Targeted mass spectrometry, antibody specificity testing with phosphomimetic mutants, Western blot\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — replicated by two independent groups (PMIDs 32130916 and 32130915), mass spectrometry and multiple antibody validations\",\n      \"pmids\": [\"32130916\", \"32130915\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Reduction in PP2Ac methylation (caused by ethanol-induced decrease in hepatic methylation capacity) led to increased degradation of the regulatory Bα subunit, promoting phosphorylation and nuclear exclusion of FOXO1, reducing FOXO1 transcriptional activity, and ultimately increasing TXNIP expression and hepatic lipid accumulation.\",\n      \"method\": \"Betaine supplementation (methyl donor), Western blot, phosphorylation analysis, subcellular fractionation, in vivo mouse model\",\n      \"journal\": \"Toxicology letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological manipulation of methylation with mechanistic pathway readouts in vitro and in vivo, single lab\",\n      \"pmids\": [\"32492475\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"PDCD10 directly binds to PP2Ac and increases its enzymatic activity, leading to YAP dephosphorylation, YAP nuclear translocation and transcriptional activation. Knockdown of PP2Ac abolished PDCD10-mediated HCC cell migration, invasion and EMT; PP2Ac inhibitor LB100 restricted tumor growth and metastasis in HCC with high PDCD10 expression.\",\n      \"method\": \"Co-immunoprecipitation, phosphatase activity assay, siRNA knockdown, LB100 pharmacological inhibition, in vivo xenograft\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP, enzymatic activity assay, siRNA epistasis, and in vivo validation; multiple orthogonal methods\",\n      \"pmids\": [\"34521817\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Taurine supplementation reduces SAM availability, which is sensed by LCMT-1 and PME-1 to reduce PP2Ac methylation. PP2Ac methylation was found necessary for M1 macrophage polarization including positive regulation of VDAC1 and PINK1, and its activation promotes PINK1-mediated mitophagy for metabolic adaptation to glycolysis during M1 polarization.\",\n      \"method\": \"Metabolic assays, Western blot for PP2Ac methylation, PINK1/VDAC1 expression, mitophagy flux assays, pharmacological inhibition of PP2Ac methylation (ABL127)\",\n      \"journal\": \"Frontiers in immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological manipulation with mechanistic pathway readouts, single lab, multiple downstream markers assessed\",\n      \"pmids\": [\"33912173\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"TRPC1-mediated Ca2+ signaling increases levels of alpha4 and PP2Ac proteins and promotes alpha4/PP2Ac association in intestinal epithelial cells, enhancing cell migration after wounding. siRNA silencing of either alpha4 or PP2Ac destabilized alpha4/PP2Ac complexes and repressed migration; cellular polyamines regulated this pathway by controlling alpha4/PP2Ac expression and association.\",\n      \"method\": \"Stable TRPC1 overexpression, siRNA knockdown, co-immunoprecipitation, wound migration assay, polyamine manipulation\",\n      \"journal\": \"Physiological reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain- and loss-of-function with Co-IP and functional migration readout, single lab\",\n      \"pmids\": [\"33991460\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"De novo PPP2CA missense variant p.Cys196Arg causes severe catalytic impairment, mildly reduced A-subunit binding, and moderately decreased binding to B/B55, B\\\"/PR72, and most B56 subunits (except B56γ1), consistent with partial loss of function. C-terminal methylation and STRN3 binding were unaffected.\",\n      \"method\": \"In vitro phosphatase activity assay, co-immunoprecipitation for A and B subunit binding, methylation assay\",\n      \"journal\": \"Frontiers in cell and developmental biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple orthogonal biochemical characterizations of variant (activity, multiple binding partners, methylation) in one rigorous study\",\n      \"pmids\": [\"36531959\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PP2Ac deficiency in glioma cells enhanced dsDNA production and activated cGAS-STING-type I IFN signaling, increased MHC-I expression, and elevated tumor mutational burden. PP2Ac genetic ablation sensitized glioma tumors to immune-checkpoint blockade and radiotherapy, establishing PP2Ac as an inhibitor of cGAS-STING signaling in glioma cells.\",\n      \"method\": \"Genetic ablation (knockout), cGAS-STING signaling assays, MHC-I expression, dsDNA measurement, co-culture with DCs and T cells, in vivo tumor model with checkpoint blockade/radiotherapy, single-cell analysis\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic KO with multiple orthogonal functional readouts (molecular signaling, co-culture, in vivo), replicated in companion paper (PMID 36757811)\",\n      \"pmids\": [\"37219874\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PP2A with its specific B regulatory subunit STRN4 (forming PP2A/STRN4 holoenzyme) negatively regulates STING-type I IFN signaling in macrophages by dephosphorylating Hippo kinase MST1/2 and stabilizing YAP/TAZ to antagonize STING activation. STING agonists induced dissociation of PP2A from MST1/2 in normal but not in tumor-conditioned macrophages.\",\n      \"method\": \"Macrophage-specific PP2A conditional knockout, Co-IP (PP2Ac-STRN4-MST1/2 complex), dephosphorylation assay, STING signaling assay, in vivo tumor model\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional KO, biochemical Co-IP showing complex, dephosphorylation assay, in vivo validation; multiple orthogonal methods\",\n      \"pmids\": [\"36757811\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PTEN directly binds to and dephosphorylates the C terminus of PP2Ac to increase its enzymatic activity; elevated PP2Ac activity in turn dephosphorylates deoxycytidine kinase (DCK) at Ser74 to diminish gemcitabine efficacy. PTEN deficiency therefore results in decreased PP2Ac activity and higher DCK phosphorylation, facilitating gemcitabine action.\",\n      \"method\": \"Direct binding assay, phosphatase activity assay, DCK phosphorylation assay, cell-based drug sensitivity assays, CDX and PDX xenograft models\",\n      \"journal\": \"Science translational medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct binding and enzymatic dephosphorylation assays combined with in vivo PDX validation and substrate identification\",\n      \"pmids\": [\"37437018\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Luteolin directly binds to KDM4C, blocking KDM4C-induced histone demethylation of the PPP2CA promoter, inhibiting PPP2CA transcription. Reduced PPP2CA expression then impairs PPP2CA-mediated YAP dephosphorylation, thereby attenuating YAP activity and ovarian cancer stem cell stemness.\",\n      \"method\": \"Direct binding assay (luteolin-KDM4C), ChIP/promoter methylation, luciferase reporter, YAP phosphorylation assay, functional stemness assays\",\n      \"journal\": \"Biomedicine & pharmacotherapy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding assay plus epigenetic and functional readouts, single lab, multiple methods\",\n      \"pmids\": [\"36804120\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"dTAG-mediated selective degradation of PPP2CA in HEK293 cells followed by global phospho-proteomics identified 2,204 proteins with significantly increased phosphorylation, constituting a broad substrate landscape. A pSP/pTP motif was identified as the predominant PPP2CA target sequence. PPP2CA substrates are enriched for functions in spliceosome, cell cycle, RNA transport, and ubiquitin-mediated proteolysis.\",\n      \"method\": \"dTAG proteolysis-targeting chimera degradation of knock-in dTAG-PPP2CA, unbiased global phospho-proteomics, immunoblotting validation\",\n      \"journal\": \"iScience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — selective protein degradation combined with unbiased global phospho-proteomics and immunoblot validation; rigorous substrate identification\",\n      \"pmids\": [\"38450154\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PKC (classical isoforms PKCα and PKCβ, but not novel PKCδ or PKCε) phosphorylates PP2Ac at Ser24. This phosphorylation is necessary and sufficient to trigger the PP2A switch (involving IGBP1/alpha4), inducing dephosphorylation and inactivation of PI3K and AKT, and leading to JNK-dependent apoptosis upon GqPCR activation.\",\n      \"method\": \"Phospho-mass spectrometry to identify Ser24 site, specific phospho-antibody generation, S24A and S24E substitution mutants, co-immunoprecipitation, proximity ligation assay, TUNEL apoptosis assay\",\n      \"journal\": \"Cell communication and signaling : CCS\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — phospho-MS site identification, phosphomimetic/unphosphorylatable mutants, specific antibody, multiple functional readouts\",\n      \"pmids\": [\"38419089\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PP2Ac regulates ULK1 phosphorylation (dephosphorylation at Ser637) during osteoclastogenesis to control autophagy. mTORC1 inhibition facilitated PP2Ac expression, and PP2Ac-mediated autophagy was dependent on ULK1 phosphorylation activity. Knockdown or inhibition of PP2Ac weakened autophagy during osteoclastogenesis.\",\n      \"method\": \"siRNA knockdown, phosphorylation assay (ULK1 Ser637), mTORC1 inhibition, in vivo OA rat model\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KD with phosphorylation site analysis and in vivo validation, single lab\",\n      \"pmids\": [\"39041921\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MKRN2 E3 ligase interacts with PPP2CA and promotes K48-linked ubiquitination at PPP2CA's K41 residue, leading to proteasomal degradation of PPP2CA. Consequently, MKRN2-mediated PPP2CA repression increased β-catenin phosphorylation and decreased its levels, inactivating Wnt signaling in clear cell renal cell carcinoma.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay (K48-linkage, K41 site identification), Western blot, in vivo xenograft\",\n      \"journal\": \"International journal of biological sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination linkage and site characterization, functional epistasis, single lab\",\n      \"pmids\": [\"40959281\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Sirt2 regulates PP2Ac activation through controlling PP2Ac acetylation at Lys136. Colchicine enhances Sirt2 expression, which deacetylates and activates PP2Ac, leading to increased phosphorylation of NLRP3 at Ser5 and reduced NLRP3 inflammasome activation, thereby inhibiting vascular calcification.\",\n      \"method\": \"Phosphorylation/acetylation assays, Sirt2 knockout mice, PP2Ac activity assay, vascular calcification models in vitro and in vivo\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO mouse model with biochemical site identification (Lys136 acetylation, Ser5 phosphorylation), single lab\",\n      \"pmids\": [\"40523615\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Carboxy-methylation of PP2Ac is highly sensitive to intracellular SAM levels. Overexpression of PME-1 (which demethylates PP2A) or expression of a Leu309-deleted unmethylated PP2A mimetic was sufficient to reduce cancer cell proliferation even in methionine-independent cells, establishing a mechanistic link between methionine/SAM availability, PP2Ac methylation status, and cell proliferation.\",\n      \"method\": \"PME-1 overexpression, Leu309-deleted PP2Ac expression, SAM measurement, cell proliferation assays\",\n      \"journal\": \"Biomolecules\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain-of-function/methylation-deficient mutant with defined proliferation readout, single lab, two orthogonal approaches\",\n      \"pmids\": [\"41008516\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"RCN2 facilitates PPP2CA ubiquitination and proteasomal degradation in a manner dependent on the HECT domain of UBR5 E3 ligase; RCN2 physically interacts with both PPP2CA and UBR5. This PPP2CA degradation activates PI3K-AKT signaling to promote ESCC metastasis and cisplatin resistance.\",\n      \"method\": \"Co-immunoprecipitation, GST pull-down, Western blot for ubiquitination, RNA-seq, LC-MS/MS, in vivo xenograft\",\n      \"journal\": \"Drug resistance updates\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, pull-down, ubiquitination assay confirming UBR5 HECT-domain dependence; single lab, multiple methods\",\n      \"pmids\": [\"41411970\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"LRRK2 phosphorylates PP2Ac at residue T304 in vitro; this LRRK2-mediated T304 phosphorylation alters C-terminal methylation of PP2Ac, impairing PP2A holoenzyme formation and catalytic activity. Reciprocally, PP2A dephosphorylates sites in the RocCOR-GTPase domain of LRRK2, de-stabilizing LRRK2 dimers and reducing its kinase activity. WT PPP2CA expression protected from LRRK2-G2019S-induced neuronal cell death, while T304 mutants failed to do so.\",\n      \"method\": \"In vitro dephosphorylation assay (PP2A on LRRK2), kinase assay (LRRK2 on PP2Ac T304), PP2Ac methylation assay, holoenzyme formation assay, neuronal cell death rescue experiment with WT vs. T304 mutant PP2CA\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro biochemical reconstitution of reciprocal phosphorylation, methylation assay, and functional rescue, but preprint not yet peer-reviewed\",\n      \"pmids\": [\"bio_10.1101_2025.10.07.680857\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"PP2Ac (Mts in Drosophila) is sufficient to trigger cell migration by activating the JNK signaling pathway. Genetic epistasis analyses in Drosophila showed Mts acts upstream of Slpr (MLK kinase) in the JNK cascade. Affinity purification-mass spectrometry identified Rho1 as a mediator; Mts activates JNK by increasing Rho1 protein levels, and Rho1 acts downstream of Mts/upstream of Slpr-JNK. PP2Ac also promoted cell migration in human pancreatic cancer cells associated with RhoA levels and JNK activation.\",\n      \"method\": \"Drosophila genetic epistasis (double mutant analysis), affinity purification-mass spectrometry (AP-MS), PP2AC overexpression in human PAAD cells, Western blot for RhoA/JNK\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis in Drosophila plus AP-MS interactor identification plus mammalian cell validation, multiple orthogonal methods\",\n      \"pmids\": [\"42144974\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"Spry2 sequesters PP2Ac upon LPS stimulation (Spry2 becomes serine-phosphorylated and associates with PP2Ac), preventing PP2Ac from dephosphorylating p65, thus heightening NF-κB nuclear translocation and cytokine production. In Spry2-deficient macrophages, PP2Ac freely interacts with p65, enhancing its dephosphorylation and reducing its nuclear translocation; PP2Ac inhibitor treatment rescued this defect.\",\n      \"method\": \"Co-immunoprecipitation, Spry2 knockout macrophages, PP2Ac inhibitor rescue, p65 nuclear translocation assay, cytokine measurement\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, KO macrophages with pharmacological rescue, nuclear translocation assay, single lab\",\n      \"pmids\": [\"41847846\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"PPP2CA encodes the principal catalytic subunit of PP2A (PP2Ac), a serine/threonine phosphatase whose activity and substrate specificity are governed by reversible C-terminal Leu309 carboxylmethylation (written by LCMT-1, erased by PME-1) and direct phosphorylation at Thr304 (by LRRK2), Ser24 (by classical PKCs), and Thr (by CDK1), with these modifications controlling holoenzyme assembly with scaffolding (A) and regulatory (B/B'/B''/B''') subunits; PP2Ac dephosphorylates a broad pSP/pTP-motif substrate landscape (>2,000 proteins including MEK/ERK, AKT, CaMKIIα, YAP, MST1/2, RIPK1, DCK, and IKK) to regulate cell cycle entry/exit, apoptosis, immune signaling (MEK-ERK-DNMT1 axis, cGAS-STING, NF-κB), metabolic reprogramming, autophagy, and cell migration (via Rho1-JNK), and is subject to ubiquitin-mediated degradation by MID1, EDD, UBR5, and MKRN2 E3 ligases.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PPP2CA encodes the catalytic subunit of protein phosphatase 2A (PP2Ac), a serine/threonine phosphatase that dephosphorylates a broad pSP/pTP-motif substrate landscape of >2,000 proteins enriched for cell cycle, RNA processing, and ubiquitin-proteolysis functions to govern proliferation, apoptosis, immune signaling, and cell migration [#30, #16]. PP2Ac assembles with scaffolding (A) and regulatory (B/B'/B''/STRN) subunits into functional holoenzymes; the conserved C-terminal Leu309 region and an intact catalytic core are required for productive B-subunit binding and activity, and active-site or truncation mutants act dominant-negatively by titrating regulatory subunits and substrates [#0, #1, #25]. Holoenzyme assembly and activity are tuned by reversible C-terminal carboxylmethylation at Leu309—written by LCMT-1, erased by PME-1, and sensitive to intracellular SAM/methionine availability—which controls B-subunit recruitment, proliferation, and downstream programs [#11, #35, #21]; additional post-translational control comes from direct phosphorylation by CDK1, which disrupts PP2A-B55 to promote mitotic entry [#19], by classical PKCα/β at Ser24, which triggers an IGBP1/alpha4-dependent switch that inactivates PI3K/AKT and drives JNK-dependent apoptosis [#31], and from acetylation at Lys136 controlled by Sirt2 [#34]. Through defined substrates PP2Ac executes its biology: it dephosphorylates MEK/ERK to suppress the DNMT1 epigenetic axis [#7], MST1/2 and YAP to control Hippo signaling, migration and inflammasome output [#22, #27], RIPK1 in glucose-deprivation cell death [#17], IKKβ and p65 in NF-κB signaling [#15, #39], DCK to modulate gemcitabine sensitivity [#28], ULK1 in autophagy [#32], and acts via Rho1/RhoA-JNK to drive cell migration [#38]; it also restrains cGAS-STING type-I interferon responses in tumor cells [#26]. The alpha4 (IGBP1) chaperone binds PP2Ac through its catalytic domain and protects it from ubiquitin-mediated degradation, while the E3 ligases MID1, EDD, UBR5, and MKRN2 target PP2Ac for proteasomal turnover [#8, #9, #33, #36, #2]. De novo PPP2CA missense and loss-of-function variants cause intellectual disability through haploinsufficient or dominant-negative impairment of phosphatase activity, A/B-subunit binding, and C-terminal methylation [#16, #25].\",\n  \"teleology\": [\n    {\n      \"year\": 1999,\n      \"claim\": \"Established that human PP2Ac is a functionally conserved catalytic subunit whose activity and proper complex assembly are essential, defining the dominant-negative behavior of catalytically dead forms.\",\n      \"evidence\": \"Yeast complementation, mutagenesis and in vitro phosphatase assays with active-site and C-terminal mutants\",\n      \"pmids\": [\"10446173\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not map all regulatory subunit contacts in human holoenzymes\", \"Leu309 modification chemistry not yet addressed\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Linked the conserved C-terminal leucine to selective B-subunit binding and mitotic checkpoint function, showing modification of this residue differentially gates holoenzyme composition.\",\n      \"evidence\": \"Yeast genetics, two-hybrid binding, and microtubule-drug phenotyping of Pph22p Leu mutants\",\n      \"pmids\": [\"11129046\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"B-subunit selectivity demonstrated in yeast orthologs, not human subunits directly\", \"Methylation status not measured\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Defined the region of the scaffolding A subunit required for physical contact with PP2Ac, anchoring the architecture of the core dimer.\",\n      \"evidence\": \"Co-IP of PPP2R1B deletion/missense mutants from colorectal cancer tissue\",\n      \"pmids\": [\"14767517\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single method, no structural validation\", \"Functional consequence of disrupted binding not assayed\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Identified the alpha4/PP2Ac complex as a cytoplasmic regulator of CaMKIIα, connecting the phosphatase to learning and memory through compartment-specific substrate dephosphorylation.\",\n      \"evidence\": \"Neuron-specific alpha4 conditional KO, Co-IP, fractionation and behavioral assays\",\n      \"pmids\": [\"16516168\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Effects attributed via alpha4 KO rather than direct PP2Ac perturbation\", \"Direct CaMKIIα dephosphorylation site not mapped\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Mapped distinct binding surfaces on PP2Ac for alpha4 and substrate S6K1, showing alpha4 bridges PP2Ac to substrates under immune stimulation.\",\n      \"evidence\": \"Pull-down with PP2Ac deletion constructs and LPS-stimulated B cell Co-IP\",\n      \"pmids\": [\"15796902\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"S6K1 dephosphorylation not directly demonstrated\", \"Single lab domain mapping\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Placed PP2Ac upstream of the MEK/ERK/DNMT1 axis, defining a route by which the phosphatase controls DNA methylation in T cells.\",\n      \"evidence\": \"siRNA and chemical inhibition with phospho-ERK, DNMT activity and methylation readouts\",\n      \"pmids\": [\"23775084\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct MEK/ERK dephosphorylation by PP2Ac vs indirect not resolved\", \"Relevant holoenzyme/B-subunit unspecified\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Identified MID1, EDD as E3 ligases that ubiquitinate PP2Ac and established alpha4 as a protective chaperone modulating PP2Ac turnover.\",\n      \"evidence\": \"In vitro ubiquitination, alpha4 deletion-construct Co-IP, siRNA and MG132 treatment\",\n      \"pmids\": [\"25207814\", \"24145130\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Ubiquitination site(s) on PP2Ac not mapped in these studies\", \"Physiological triggers of degradation partly inferred\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Demonstrated receptor- and PI3K-controlled Leu309 carboxylmethylation governs PP2Ac localization and showed PP2Ac suppresses β-catenin/NF-κB via Akt-GSK3β/IκBα, establishing methylation and signaling control of the phosphatase.\",\n      \"evidence\": \"Carboxylmethylation assays with Gi-receptor agonists; gain/loss-of-function with reporter and orthotopic models in prostate cancer\",\n      \"pmids\": [\"24475092\", \"24642616\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether methylation directly dictates specific B-subunit recruitment in these contexts untested\", \"Direct substrate of PP2Ac in β-catenin/NF-κB axis not identified\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Provided structural and biochemical basis for C-terminal-tail recognition of PP2Ac by the regulator TIPRL, which preferentially binds unmethylated PP2Ac and occludes the active site.\",\n      \"evidence\": \"TIPRL crystal structure at 2.15 Å, mutagenesis, pull-down, HDX-MS and docking\",\n      \"pmids\": [\"27489114\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Cellular consequence of TIPRL-mediated active-site blockade not quantified here\", \"No structure of full PP2Ac-TIPRL complex\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Established PPP2CA as an intellectual-disability gene and dissected how de novo variants impair activity, subunit binding and methylation by haploinsufficient or dominant-negative mechanisms.\",\n      \"evidence\": \"Functional expression, phosphatase activity, A/B-subunit Co-IP and methylation assays across patient variants\",\n      \"pmids\": [\"30595372\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo neurodevelopmental mechanism not modeled\", \"Substrate-level consequences in neurons not defined\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Linked glucose-sensing through Cav1.3/CAMK1/PPME1 to PP2Ac demethylation and RIPK1-dependent cell death, defining a metabolic control of PP2Ac inactivation.\",\n      \"evidence\": \"Channel pharmacology, CAMK1/PPME1 knockdown and demethylation/cell-death assays\",\n      \"pmids\": [\"29317521\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct RIPK1 dephosphorylation by PP2Ac not formally shown\", \"B-subunit specifying RIPK1 targeting unknown\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Showed CDK1 directly phosphorylates a PP2Ac threonine to disrupt PP2A-B55 and promote mitotic entry, adding a direct layer to CDK1-PP2A crosstalk.\",\n      \"evidence\": \"Quantitative chemical proteomics with holoenzyme enrichment and kinase profiling\",\n      \"pmids\": [\"32900880\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Precise threonine residue and its structural effect not fully resolved\", \"In vivo cell-cycle requirement not tested\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Demonstrated that widely used phospho-Tyr307 antibodies are non-specific and instead report Leu309 methylation and Thr304 phosphorylation, forcing reinterpretation of prior PP2Ac regulatory studies.\",\n      \"evidence\": \"Targeted mass spectrometry and antibody testing with phosphomimetic mutants, replicated by two groups\",\n      \"pmids\": [\"32130916\", \"32130915\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological role of genuine pTyr307 remains undefined\", \"Does not establish the true functional reader of these modifications\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Connected PP2Ac to Hippo/YAP signaling and metabolism by showing PDCD10-stimulated PP2Ac dephosphorylates YAP to drive tumor migration, and methylation-dependent PP2Ac regulates macrophage mitophagy.\",\n      \"evidence\": \"Co-IP, phosphatase activity, siRNA epistasis and LB100 with xenografts; methylation manipulation with mitophagy readouts\",\n      \"pmids\": [\"34521817\", \"33912173\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct YAP dephosphorylation site not mapped\", \"B-subunit directing YAP/MST targeting unspecified\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined PP2Ac as a brake on innate antitumor immunity, restraining cGAS-STING type-I IFN signaling in glioma and, via the STRN4 holoenzyme, dephosphorylating MST1/2 to antagonize STING in macrophages.\",\n      \"evidence\": \"Genetic ablation/conditional KO, cGAS-STING and dephosphorylation assays, co-culture and in vivo tumor models\",\n      \"pmids\": [\"37219874\", \"36757811\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct STING-pathway substrate(s) in glioma not pinpointed\", \"Generality across tumor types not established\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identified PTEN as a direct PP2Ac activator that dephosphorylates its C-terminus, and DCK Ser74 as a substrate controlling gemcitabine efficacy, linking PP2Ac to chemotherapy response.\",\n      \"evidence\": \"Direct binding, phosphatase and DCK phosphorylation assays with CDX/PDX models\",\n      \"pmids\": [\"37437018\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism by which C-terminal dephosphorylation raises activity not structurally defined\", \"Phosphatase activity of PTEN on PP2Ac vs scaffolding role\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Provided an unbiased, system-wide definition of the PP2Ac substrate landscape and its pSP/pTP target motif, anchoring its broad regulatory reach.\",\n      \"evidence\": \"dTAG-mediated PPP2CA degradation with global phospho-proteomics\",\n      \"pmids\": [\"38450154\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Holoenzyme/B-subunit assignment per substrate not resolved\", \"Direct vs indirect substrates not fully distinguished\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified PKCα/β phosphorylation of PP2Ac at Ser24 as the trigger of an IGBP1/alpha4-dependent PP2A switch that inactivates PI3K/AKT and drives JNK-dependent apoptosis.\",\n      \"evidence\": \"Phospho-MS, S24A/S24E mutants, specific antibody, Co-IP, PLA and TUNEL\",\n      \"pmids\": [\"38419089\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of the alpha4-mediated switch not resolved\", \"Generality beyond GqPCR stimulation untested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Expanded the PP2Ac degradation network and modification control, identifying MKRN2 (K48/K41) and RCN2-UBR5 as degradation routes and Sirt2-controlled Lys136 acetylation as an activity switch.\",\n      \"evidence\": \"Co-IP, ubiquitination linkage/site mapping, acetylation and activity assays with KO mice and xenografts\",\n      \"pmids\": [\"40959281\", \"41411970\", \"40523615\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Each E3-PP2Ac axis from single labs without cross-validation\", \"Relative contribution of competing E3 ligases in vivo unclear\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Established that PP2Ac carboxymethylation is tightly coupled to SAM/methionine availability and that demethylated PP2Ac suppresses cancer proliferation, linking one-carbon metabolism to PP2A activity.\",\n      \"evidence\": \"PME-1 overexpression, Leu309-deletion mimetic, SAM measurement and proliferation assays\",\n      \"pmids\": [\"41008516\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Downstream substrate program of demethylated PP2Ac not defined\", \"Single-lab gain-of-function evidence\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Defined a PP2Ac-Rho1/RhoA-JNK migration axis and a Spry2-mediated sequestration mechanism controlling p65 dephosphorylation in NF-κB signaling.\",\n      \"evidence\": \"Drosophila genetic epistasis with AP-MS plus mammalian validation; Spry2-KO macrophages with Co-IP and inhibitor rescue\",\n      \"pmids\": [\"42144974\", \"41847846\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct Rho1/RhoA substrate relationship vs level control not fully resolved\", \"Spry2 mechanism single-lab\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Proposed reciprocal regulation between LRRK2 and PP2Ac in a neuronal context, with LRRK2 phosphorylating PP2Ac at Thr304 and PP2Ac dephosphorylating LRRK2.\",\n      \"evidence\": \"In vitro kinase/phosphatase and methylation assays with neuronal cell-death rescue (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.10.07.680857\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Preprint, not peer-reviewed\", \"Thr304-methylation crosstalk not validated in vivo\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How specific holoenzyme B-subunit compositions are matched to the >2,000 substrate landscape, and how the layered modification code (methylation, Ser24/Thr304/Thr phosphorylation, Lys136 acetylation) is integrated to dictate substrate choice, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No comprehensive map linking individual B-subunits to defined substrates\", \"Combinatorial effect of co-occurring PP2Ac modifications not deconvolved\", \"Structures of substrate-engaged human holoenzymes lacking\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 30]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [7, 28, 30, 27]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [19, 31]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [5, 6, 13]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [6, 10]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [19, 1]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [7, 22, 31, 38]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [26, 27, 39]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [17, 31]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [32, 23]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [8, 9, 33, 36]}\n    ],\n    \"complexes\": [\"PP2A holoenzyme\", \"PP2A-B55\", \"PP2A-B56\", \"PP2A/STRN4\"],\n    \"partners\": [\"PPP2R1B\", \"IGBP1\", \"TIPRL\", \"PDCD10\", \"PTEN\", \"MID1\", \"UBR5\", \"MKRN2\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}