{"gene":"MPP1","run_date":"2026-06-10T02:59:51","timeline":{"discoveries":[{"year":2007,"finding":"MPP1/p55 interacts with MPP5/Pals1 at the outer limiting membrane (OLM) of the retina via heterodimerization of their MAGUK modules in a directional fashion, linking the Usher protein network to the Crumbs polarity complex.","method":"Protein interaction assays (binding/interaction studies), co-localization in retina, domain mapping","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — single lab, co-localization and interaction data with domain-level mechanistic detail, but methods described at abstract level without full reconstitution","pmids":["17584769"],"is_preprint":false},{"year":2007,"finding":"MPP1 interacts with whirlin (a multi-PDZ scaffold protein of the Usher protein network) via both a classical PDZ domain-to-PDZ binding motif (PBM) mechanism and a mechanism involving internal epitopes; they co-localize at the OLM, outer synaptic layer, basal bodies, and ciliary axoneme in the retina.","method":"Protein interaction assays, co-localization by immunofluorescence in retinal tissue","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — single lab, two interaction mechanisms identified and co-localization demonstrated, but abstract-level description without full reconstitution","pmids":["17584769"],"is_preprint":false},{"year":2009,"finding":"p55/MPP1 knockout neutrophils form multiple transient pseudopods upon chemotactic stimulation and do not migrate efficiently in vitro; upon agonist stimulation, p55 is recruited to the leading edge. In p55(-/-) neutrophils, Akt phosphorylation is significantly decreased and PIP3 is diffusely localized rather than accumulated at the leading edge, despite normal PI3Kgamma activity and normal total PIP3 levels. Thus MPP1 regulates neutrophil polarity by controlling spatial restriction of PIP3 and downstream Akt activation independently of PI3Kgamma.","method":"p55 knockout mouse model, chemotaxis assays in vitro, immunofluorescence for PIP3 localization, Akt phosphorylation assays, PI3Kgamma activity immunoprecipitation assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean KO mouse with defined cellular phenotype, multiple orthogonal methods (migration assay, kinase activity, PIP3 localization, Akt phosphorylation), mechanistic pathway placement","pmids":["19897731"],"is_preprint":false},{"year":2009,"finding":"The FERM domain of NF2/merlin directly binds erythrocyte p55/MPP1 with a KD of 3.7 nM as measured by surface plasmon resonance; both proteins co-localize in non-myelin-forming Schwann cells.","method":"Surface plasmon resonance, co-localization by immunofluorescence, monoclonal antibody development","journal":"Experimental biology and medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding measured by SPR with defined affinity, co-localization in vivo, single lab","pmids":["19144871"],"is_preprint":false},{"year":2012,"finding":"DHHC17 is the palmitoylating enzyme (acyltransferase) responsible for MPP1 palmitoylation in red blood cells; loss of DHHC17-mediated palmitoylation of MPP1 results in reduced detergent-resistant membrane (DRM) material and decreased membrane order, linking MPP1 palmitoylation to lateral membrane organization in erythrocytes.","method":"Clinical patient RBC analysis (lacking DHHC17), chemical inhibition with 2-bromopalmitic acid, FLIM analysis of membrane order, DRM biochemical fractionation","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (clinical cases, chemical inhibition, FLIM, DRM fractionation), identification of specific writer enzyme, functional consequence on membrane organization","pmids":["22496366"],"is_preprint":false},{"year":2013,"finding":"MPP1 palmitoylation or MPP1 gene silencing (knockdown) in HEL erythroid precursor cells leads to a dramatic decrease in DRM fraction and reduction in membrane order; MPP1 knockdown also significantly impairs MAP-kinase signaling via raft-dependent RTK receptors.","method":"MPP1 gene silencing (knockdown), palmitoylation inhibition, DRM fractionation, FLIM, MAP-kinase signaling assays","journal":"Biochimica et biophysica acta","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (KD, chemical inhibition, FLIM, DRM, signaling assays) in a single study replicating and extending the erythrocyte findings","pmids":["23507198"],"is_preprint":false},{"year":2015,"finding":"MPP1 modulates membrane fluidity and phase separation capability of giant plasma membrane-derived vesicles (GPMVs) from live cells, demonstrating that membrane physicochemical domain properties can be tuned by MPP1 protein without major changes in lipid composition.","method":"Giant plasma membrane-derived vesicles (GPMVs), fluorescence microscopy of phase separation, miscibility phase transition temperature analysis","journal":"Biophysical journal","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct biophysical assay in GPMVs with functional consequence on membrane phase behavior, single lab, single method type","pmids":["25954878"],"is_preprint":false},{"year":2017,"finding":"MPP1 directly binds flotillin 1 and flotillin 2 in erythrocyte membrane; these MPP1-flotillin interactions are distinct from the known protein 4.1-dependent interactions of MPP1. Loss of MPP1-flotillin interactions results in significant changes in RBC membrane fluidity as shown by FLIM, indicating physiological importance for raft domain organization.","method":"Multiple protein interaction methods (Co-IP, pulldown), FLIM analysis of membrane fluidity, native RBC membrane complex analysis","journal":"Biochimica et biophysica acta. Biomembranes","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (Co-IP/pulldown plus FLIM functional readout), identification of two binding partners, mechanistic distinction from known 4.1-dependent interactions","pmids":["28865798"],"is_preprint":false},{"year":2018,"finding":"MPP1 knockdown in HEL cells impairs insulin receptor signaling specifically at the level of H-Ras, causing impaired GDP-to-GTP exchange and reduced interaction between H-Ras and its effector Raf; H-Ras DRM localization was not sensitive to insulin treatment upon MPP1 knockdown, suggesting MPP1-dependent membrane domain organization is required for H-Ras activation.","method":"MPP1 knockdown, H-Ras GTP loading assay, Raf interaction assay, DRM fractionation, insulin stimulation experiments","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean KD with defined signaling phenotype, multiple assays identifying pathway position, single lab","pmids":["29719614"],"is_preprint":false},{"year":2021,"finding":"High-affinity MPP1-flotillin complexes are formed via a specific 'flotillin binding motif' within the D5 domain of MPP1; overexpression of peptides containing this motif inhibited endogenous MPP1-flotillin interaction in erythroid precursor cells, causing lateral disorganization of raft domains (reduced plasma membrane order) and markedly decreased activation of raft-dependent insulin receptor signaling.","method":"Molecular dynamics simulations, surface plasmon resonance, dominant-negative peptide overexpression, membrane order measurement (FLIM), insulin receptor signaling assays","journal":"Scientific reports","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — binding affinity measured by SPR, domain identified by MD simulations, functional validation by dominant-negative peptide approach with multiple orthogonal readouts (FLIM, signaling)","pmids":["34285255"],"is_preprint":false},{"year":2022,"finding":"MPP1 acts as a key raft-capturing molecule that regulates temporal immobilization of flotillin-based nanoclusters and controls local concentration and confinement of sphingomyelin and Thy-1 in raft nanodomains in erythroid cells, as revealed by super-resolution structured illumination imaging, FRAP, and spot-variation fluorescence correlation spectroscopy (svFCS).","method":"Structured illumination microscopy (super-resolution), FRAP, spot-variation FCS (svFCS)","journal":"Cells","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal spectroscopic and imaging methods, direct measurement of molecular mobility and confinement, builds on prior replicated findings","pmids":["35159121"],"is_preprint":false},{"year":2023,"finding":"MPP1 co-localizes with the angiotensin II AT1 receptor (AGTR1) on sarcolemmal membranes in vivo; transgenic overexpression of MPP1 (2-fold) in mice causes heart failure with reduced ejection fraction, cardiac enlargement and dilation, and increased AGTR1 protein levels. MPP1 also directly increases AGTR1 protein in HEK cells, demonstrating MPP1 can upregulate AGTR1 protein levels.","method":"Transgenic mouse model (Tg-MPP1), echocardiography, histology, co-localization in vivo, AGTR1eYFP fluorescence measurement in HEK cells","journal":"Biochemical pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — transgenic mouse with defined cardiac phenotype plus cell-based validation of AGTR1 upregulation, single lab, co-localization without direct binding assay","pmids":["37683843"],"is_preprint":false},{"year":2024,"finding":"MPP1 binds USP12 (identified by co-immunoprecipitation and mass spectrometry) and promotes CCL5 expression via the MPP1/USP12/CCL5 cascade in urothelial carcinoma cells, thereby enhancing CD8+ T-cell chemotaxis and inhibiting immune escape.","method":"Co-immunoprecipitation, mass spectrometry, RT-qPCR, western blotting, MPP1 overexpression vector, CD8+ T-cell coculture assay, in vivo tumorigenesis in HuNOG mice","journal":"International immunopharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus MS identifies binding partner, multiple functional assays in vitro and in vivo, single lab","pmids":["39700963"],"is_preprint":false},{"year":2025,"finding":"Reconstituted MPP1 with recombinant flotillins in giant unilamellar vesicles (GUVs) promotes membrane remodeling and triggers coexistence of liquid-ordered (Lo) and liquid-disordered (Ld) domains; palmitoylation of MPP1 exerts additional influence on membrane organization. Flotillin-MPP1 assemblies are sufficient and necessary to modulate lateral organization of lipid bilayers.","method":"Reconstitution in GUVs, FLIM, fluorescence microscopy of phase separation with recombinant proteins","journal":"Biochimica et biophysica acta. Biomembranes","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstitution with purified recombinant proteins in minimal membrane system, FLIM functional readout, establishes sufficiency of MPP1-flotillin complex for domain formation","pmids":["41061789"],"is_preprint":false}],"current_model":"MPP1/p55 is a palmitoylated MAGUK scaffolding protein that organizes plasma membrane raft nanodomains by directly interacting with flotillins (via a defined D5 domain motif), regulating flotillin nanocluster immobilization and confinement of raft-associated lipids (sphingomyelin, Thy-1); its palmitoylation (written by DHHC17 in erythrocytes) is required for lateral membrane order and DRM formation, and it controls downstream signaling cascades including H-Ras/Raf activation, Akt/PIP3 spatial restriction for neutrophil polarity, RTK/MAP-kinase signaling, and insulin receptor-dependent pathways, while also interacting with MPP5/Pals1 (Crumbs complex), whirlin (Usher network), and NF2/merlin to link polarity and cytoskeletal networks in retina, erythrocytes, and non-erythroid cells."},"narrative":{"mechanistic_narrative":"MPP1/p55 is a palmitoylated MAGUK scaffolding protein that organizes the lateral order of plasma membrane raft nanodomains and couples this organization to receptor signaling and cell polarity [PMID:23507198, PMID:35159121]. Its central biochemical activity is a direct, high-affinity interaction with flotillin-1 and flotillin-2 through a defined flotillin-binding motif in the D5 domain; this interaction is distinct from MPP1's protein 4.1-dependent contacts and is required to maintain erythrocyte membrane fluidity and order [PMID:28865798, PMID:34285255]. Reconstitution of MPP1 with recombinant flotillins in synthetic vesicles is sufficient to drive coexistence of liquid-ordered and liquid-disordered domains, establishing the MPP1-flotillin assembly as a minimal unit capable of remodeling membrane lateral organization [PMID:41061789]. Mechanistically, MPP1 captures and temporally immobilizes flotillin-based nanoclusters and confines raft lipids such as sphingomyelin and Thy-1 [PMID:35159121], and its palmitoylation — written by DHHC17 in red blood cells — is required for detergent-resistant membrane formation and membrane order [PMID:22496366, PMID:23507198]. This raft-organizing function underlies MPP1's control of multiple raft-dependent signaling outputs: spatial restriction of PIP3 and downstream Akt activation that drives neutrophil chemotactic polarity [PMID:19897731], H-Ras GDP/GTP exchange and H-Ras-Raf coupling in insulin receptor signaling [PMID:29719614, PMID:34285255], and RTK/MAP-kinase signaling in erythroid cells [PMID:23507198]. MPP1 additionally functions as a scaffold linking polarity and cytoskeletal networks through directional heterodimerization with MPP5/Pals1 and interaction with whirlin at the retinal outer limiting membrane [PMID:17584769] and through nanomolar-affinity binding to the FERM domain of NF2/merlin [PMID:19144871]. Roles in cardiac AGTR1 regulation [PMID:37683843] and in a USP12/CCL5 axis modulating anti-tumor immunity [PMID:39700963] have also been described.","teleology":[{"year":2007,"claim":"Established MPP1 as a scaffolding hub linking distinct polarity and Usher networks, answering how disparate retinal complexes are physically coupled.","evidence":"Interaction and domain-mapping assays with co-localization in retinal tissue, showing directional MAGUK heterodimerization with MPP5/Pals1 and dual-mode binding to whirlin","pmids":["17584769"],"confidence":"Medium","gaps":["No reconstitution of the multiprotein complex","Functional consequence of these interactions for polarity not tested by perturbation"]},{"year":2009,"claim":"Defined MPP1 as a spatial regulator of PIP3/Akt signaling required for cell polarity, distinguishing localization control from kinase activity itself.","evidence":"p55 knockout mouse neutrophils with chemotaxis assays, PIP3 immunolocalization, Akt phosphorylation, and PI3Kgamma activity assays","pmids":["19897731"],"confidence":"High","gaps":["Molecular mechanism by which MPP1 restricts PIP3 not resolved","Link to raft organization not yet established in this study"]},{"year":2009,"claim":"Quantified a direct, high-affinity MPP1-merlin interaction, placing MPP1 in cytoskeletal/tumor-suppressor scaffolding contexts beyond erythrocytes.","evidence":"Surface plasmon resonance (KD 3.7 nM) between NF2/merlin FERM domain and erythrocyte p55, with co-localization in Schwann cells","pmids":["19144871"],"confidence":"Medium","gaps":["Functional consequence of the merlin interaction not tested","Single lab, no cellular perturbation"]},{"year":2012,"claim":"Identified DHHC17 as the writer of MPP1 palmitoylation and tied this modification causally to membrane order, defining the post-translational requirement for MPP1 raft function.","evidence":"Clinical DHHC17-deficient RBCs, 2-bromopalmitate inhibition, FLIM membrane order, and DRM fractionation","pmids":["22496366"],"confidence":"High","gaps":["Palmitoylation site(s) on MPP1 not mapped","Whether DHHC17 acts on MPP1 in non-erythroid cells unknown"]},{"year":2013,"claim":"Connected MPP1 levels and palmitoylation to raft-dependent RTK/MAP-kinase signaling, extending the membrane-order role to a signaling output.","evidence":"MPP1 knockdown and palmitoylation inhibition in HEL erythroid cells with DRM fractionation, FLIM, and MAP-kinase assays","pmids":["23507198"],"confidence":"High","gaps":["Which RTKs are affected not enumerated","Direct molecular link between MPP1 and the receptors not shown"]},{"year":2015,"claim":"Demonstrated MPP1 tunes membrane phase behavior directly, showing domain properties depend on the protein rather than lipid composition changes.","evidence":"GPMVs from live cells analyzed for phase separation and miscibility transition temperature","pmids":["25954878"],"confidence":"Medium","gaps":["Binding partner driving phase modulation not identified in this study","Single biophysical method type"]},{"year":2017,"claim":"Identified flotillins 1 and 2 as direct MPP1 partners distinct from 4.1-dependent contacts, providing the molecular basis for MPP1's raft-organizing activity.","evidence":"Co-IP and pulldown in erythrocyte membranes plus FLIM membrane fluidity readout","pmids":["28865798"],"confidence":"High","gaps":["Binding interface not yet mapped at this stage","Stoichiometry of the complex unknown"]},{"year":2018,"claim":"Placed MPP1 upstream of H-Ras activation in insulin signaling, linking membrane domain organization to GDP/GTP exchange and effector coupling.","evidence":"MPP1 knockdown in HEL cells with H-Ras GTP-loading and Raf interaction assays plus DRM fractionation under insulin stimulation","pmids":["29719614"],"confidence":"Medium","gaps":["Mechanism connecting raft order to H-Ras nucleotide exchange not resolved","Single cell system"]},{"year":2021,"claim":"Localized the flotillin-binding activity to a specific D5 motif and proved its necessity for raft order and insulin signaling via a dominant-negative peptide.","evidence":"Molecular dynamics, SPR affinity measurement, dominant-negative motif-peptide overexpression, FLIM, and insulin receptor signaling assays in erythroid precursors","pmids":["34285255"],"confidence":"High","gaps":["Structure of the full MPP1-flotillin complex not solved","Whether the same motif governs non-erythroid functions untested"]},{"year":2022,"claim":"Resolved the physical mechanism of raft organization as MPP1-driven immobilization of flotillin nanoclusters and confinement of specific raft lipids.","evidence":"Super-resolution structured illumination imaging, FRAP, and spot-variation FCS in erythroid cells measuring mobility and confinement of sphingomyelin and Thy-1","pmids":["35159121"],"confidence":"High","gaps":["Kinetics of nanocluster capture not detailed","Generalizability beyond erythroid cells not tested"]},{"year":2023,"claim":"Extended MPP1 function to cardiac AGTR1 regulation, showing MPP1 dosage controls receptor protein levels and cardiac physiology.","evidence":"Transgenic MPP1-overexpressing mice with echocardiography and histology, in vivo co-localization, and AGTR1 protein measurement in HEK cells","pmids":["37683843"],"confidence":"Medium","gaps":["No direct MPP1-AGTR1 binding assay","Mechanism of AGTR1 upregulation unknown"]},{"year":2024,"claim":"Linked MPP1 to immune modulation through a USP12/CCL5 axis affecting T-cell chemotaxis in urothelial carcinoma.","evidence":"Co-IP and mass spectrometry identifying USP12 binding, RT-qPCR/western blot, CD8+ T-cell coculture, and HuNOG tumorigenesis assays","pmids":["39700963"],"confidence":"Medium","gaps":["How MPP1-USP12 binding drives CCL5 expression not mechanistically resolved","Relationship to MPP1's raft-organizing role unclear"]},{"year":2025,"claim":"Proved sufficiency: reconstituted MPP1-flotillin assemblies alone generate ordered/disordered domain coexistence in minimal bilayers.","evidence":"Reconstitution of recombinant MPP1 and flotillins in GUVs with FLIM and phase-separation microscopy, with palmitoylation modulating organization","pmids":["41061789"],"confidence":"High","gaps":["Quantitative stoichiometry and structural arrangement in the bilayer not defined","How signaling receptors partition into reconstituted domains not addressed"]},{"year":null,"claim":"How MPP1's raft-organizing scaffold activity is mechanistically reconciled with its discrete roles in AGTR1 regulation and the USP12/CCL5 immune axis remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking membrane-order function to AGTR1/USP12 phenotypes","Structure of MPP1-flotillin complex unsolved","Tissue-specificity of partner usage not mapped"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,1,3,7,9]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[3]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[4,7,10,11]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[2,8,9]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[2,12]}],"complexes":[],"partners":["FLOT1","FLOT2","MPP5","WHRN","NF2","USP12","AGTR1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q00013","full_name":"55 kDa erythrocyte membrane protein","aliases":["Membrane protein, palmitoylated 1"],"length_aa":466,"mass_kda":52.3,"function":"Essential regulator of neutrophil polarity. Regulates neutrophil polarization by regulating AKT1 phosphorylation through a mechanism that is independent of PIK3CG activity (By similarity)","subcellular_location":"Cell membrane; Cell projection, stereocilium","url":"https://www.uniprot.org/uniprotkb/Q00013/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/MPP1","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"SRP9","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/MPP1","total_profiled":1310},"omim":[{"mim_id":"607928","title":"WHIRLIN; WHRN","url":"https://www.omim.org/entry/607928"},{"mim_id":"607350","title":"KINESIN FAMILY MEMBER 13B; KIF13B","url":"https://www.omim.org/entry/607350"},{"mim_id":"606959","title":"PROTEIN ASSOCIATED WITH LIN7 2, MAGUK p55 FAMILY MEMBER; PALS2","url":"https://www.omim.org/entry/606959"},{"mim_id":"606958","title":"PROTEIN ASSOCIATED WITH LIN7 1, MAGUK p55 FAMILY MEMBER; PALS1","url":"https://www.omim.org/entry/606958"},{"mim_id":"605498","title":"KINESIN FAMILY MEMBER 20B; KIF20B","url":"https://www.omim.org/entry/605498"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Plasma membrane","reliability":"Supported"},{"location":"Centriolar satellite","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"bone marrow","ntpm":110.2}],"url":"https://www.proteinatlas.org/search/MPP1"},"hgnc":{"alias_symbol":["PEMP"],"prev_symbol":["DXS552E"]},"alphafold":{"accession":"Q00013","domains":[{"cath_id":"2.30.42.10","chopping":"69-148","consensus_level":"high","plddt":84.7943,"start":69,"end":148},{"cath_id":"2.30.30.40","chopping":"160-227","consensus_level":"high","plddt":91.3188,"start":160,"end":227},{"cath_id":"3.40.50.300","chopping":"278-309_376-466","consensus_level":"medium","plddt":94.4454,"start":278,"end":466},{"cath_id":"3.30.63.10","chopping":"312-373","consensus_level":"medium","plddt":95.8552,"start":312,"end":373}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q00013","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q00013-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q00013-F1-predicted_aligned_error_v6.png","plddt_mean":78.12},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=MPP1","jax_strain_url":"https://www.jax.org/strain/search?query=MPP1"},"sequence":{"accession":"Q00013","fasta_url":"https://rest.uniprot.org/uniprotkb/Q00013.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q00013/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q00013"}},"corpus_meta":[{"pmid":"17584769","id":"PMC_17584769","title":"MPP1 links the Usher protein network and the Crumbs protein complex in the retina.","date":"2007","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/17584769","citation_count":35,"is_preprint":false},{"pmid":"19897731","id":"PMC_19897731","title":"Erythrocyte scaffolding protein p55/MPP1 functions as an essential regulator of neutrophil polarity.","date":"2009","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/19897731","citation_count":33,"is_preprint":false},{"pmid":"22496366","id":"PMC_22496366","title":"Palmitoylation of MPP1 (membrane-palmitoylated protein 1)/p55 is crucial for lateral membrane organization in erythroid cells.","date":"2012","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/22496366","citation_count":27,"is_preprint":false},{"pmid":"23507198","id":"PMC_23507198","title":"The role of MPP1/p55 and its palmitoylation in resting state raft organization in HEL cells.","date":"2013","source":"Biochimica et biophysica acta","url":"https://pubmed.ncbi.nlm.nih.gov/23507198","citation_count":25,"is_preprint":false},{"pmid":"25954878","id":"PMC_25954878","title":"MPP1 as a Factor Regulating Phase Separation in Giant Plasma Membrane-Derived Vesicles.","date":"2015","source":"Biophysical journal","url":"https://pubmed.ncbi.nlm.nih.gov/25954878","citation_count":23,"is_preprint":false},{"pmid":"28865798","id":"PMC_28865798","title":"MPP1 directly interacts with flotillins in erythrocyte membrane - Possible mechanism of raft domain formation.","date":"2017","source":"Biochimica et biophysica acta. 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In p55(-/-) neutrophils, Akt phosphorylation is significantly decreased and PIP3 is diffusely localized rather than accumulated at the leading edge, despite normal PI3Kgamma activity and normal total PIP3 levels. Thus MPP1 regulates neutrophil polarity by controlling spatial restriction of PIP3 and downstream Akt activation independently of PI3Kgamma.\",\n      \"method\": \"p55 knockout mouse model, chemotaxis assays in vitro, immunofluorescence for PIP3 localization, Akt phosphorylation assays, PI3Kgamma activity immunoprecipitation assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean KO mouse with defined cellular phenotype, multiple orthogonal methods (migration assay, kinase activity, PIP3 localization, Akt phosphorylation), mechanistic pathway placement\",\n      \"pmids\": [\"19897731\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"The FERM domain of NF2/merlin directly binds erythrocyte p55/MPP1 with a KD of 3.7 nM as measured by surface plasmon resonance; both proteins co-localize in non-myelin-forming Schwann cells.\",\n      \"method\": \"Surface plasmon resonance, co-localization by immunofluorescence, monoclonal antibody development\",\n      \"journal\": \"Experimental biology and medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding measured by SPR with defined affinity, co-localization in vivo, single lab\",\n      \"pmids\": [\"19144871\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"DHHC17 is the palmitoylating enzyme (acyltransferase) responsible for MPP1 palmitoylation in red blood cells; loss of DHHC17-mediated palmitoylation of MPP1 results in reduced detergent-resistant membrane (DRM) material and decreased membrane order, linking MPP1 palmitoylation to lateral membrane organization in erythrocytes.\",\n      \"method\": \"Clinical patient RBC analysis (lacking DHHC17), chemical inhibition with 2-bromopalmitic acid, FLIM analysis of membrane order, DRM biochemical fractionation\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (clinical cases, chemical inhibition, FLIM, DRM fractionation), identification of specific writer enzyme, functional consequence on membrane organization\",\n      \"pmids\": [\"22496366\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"MPP1 palmitoylation or MPP1 gene silencing (knockdown) in HEL erythroid precursor cells leads to a dramatic decrease in DRM fraction and reduction in membrane order; MPP1 knockdown also significantly impairs MAP-kinase signaling via raft-dependent RTK receptors.\",\n      \"method\": \"MPP1 gene silencing (knockdown), palmitoylation inhibition, DRM fractionation, FLIM, MAP-kinase signaling assays\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (KD, chemical inhibition, FLIM, DRM, signaling assays) in a single study replicating and extending the erythrocyte findings\",\n      \"pmids\": [\"23507198\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"MPP1 modulates membrane fluidity and phase separation capability of giant plasma membrane-derived vesicles (GPMVs) from live cells, demonstrating that membrane physicochemical domain properties can be tuned by MPP1 protein without major changes in lipid composition.\",\n      \"method\": \"Giant plasma membrane-derived vesicles (GPMVs), fluorescence microscopy of phase separation, miscibility phase transition temperature analysis\",\n      \"journal\": \"Biophysical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct biophysical assay in GPMVs with functional consequence on membrane phase behavior, single lab, single method type\",\n      \"pmids\": [\"25954878\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"MPP1 directly binds flotillin 1 and flotillin 2 in erythrocyte membrane; these MPP1-flotillin interactions are distinct from the known protein 4.1-dependent interactions of MPP1. Loss of MPP1-flotillin interactions results in significant changes in RBC membrane fluidity as shown by FLIM, indicating physiological importance for raft domain organization.\",\n      \"method\": \"Multiple protein interaction methods (Co-IP, pulldown), FLIM analysis of membrane fluidity, native RBC membrane complex analysis\",\n      \"journal\": \"Biochimica et biophysica acta. Biomembranes\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (Co-IP/pulldown plus FLIM functional readout), identification of two binding partners, mechanistic distinction from known 4.1-dependent interactions\",\n      \"pmids\": [\"28865798\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"MPP1 knockdown in HEL cells impairs insulin receptor signaling specifically at the level of H-Ras, causing impaired GDP-to-GTP exchange and reduced interaction between H-Ras and its effector Raf; H-Ras DRM localization was not sensitive to insulin treatment upon MPP1 knockdown, suggesting MPP1-dependent membrane domain organization is required for H-Ras activation.\",\n      \"method\": \"MPP1 knockdown, H-Ras GTP loading assay, Raf interaction assay, DRM fractionation, insulin stimulation experiments\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KD with defined signaling phenotype, multiple assays identifying pathway position, single lab\",\n      \"pmids\": [\"29719614\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"High-affinity MPP1-flotillin complexes are formed via a specific 'flotillin binding motif' within the D5 domain of MPP1; overexpression of peptides containing this motif inhibited endogenous MPP1-flotillin interaction in erythroid precursor cells, causing lateral disorganization of raft domains (reduced plasma membrane order) and markedly decreased activation of raft-dependent insulin receptor signaling.\",\n      \"method\": \"Molecular dynamics simulations, surface plasmon resonance, dominant-negative peptide overexpression, membrane order measurement (FLIM), insulin receptor signaling assays\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — binding affinity measured by SPR, domain identified by MD simulations, functional validation by dominant-negative peptide approach with multiple orthogonal readouts (FLIM, signaling)\",\n      \"pmids\": [\"34285255\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"MPP1 acts as a key raft-capturing molecule that regulates temporal immobilization of flotillin-based nanoclusters and controls local concentration and confinement of sphingomyelin and Thy-1 in raft nanodomains in erythroid cells, as revealed by super-resolution structured illumination imaging, FRAP, and spot-variation fluorescence correlation spectroscopy (svFCS).\",\n      \"method\": \"Structured illumination microscopy (super-resolution), FRAP, spot-variation FCS (svFCS)\",\n      \"journal\": \"Cells\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal spectroscopic and imaging methods, direct measurement of molecular mobility and confinement, builds on prior replicated findings\",\n      \"pmids\": [\"35159121\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MPP1 co-localizes with the angiotensin II AT1 receptor (AGTR1) on sarcolemmal membranes in vivo; transgenic overexpression of MPP1 (2-fold) in mice causes heart failure with reduced ejection fraction, cardiac enlargement and dilation, and increased AGTR1 protein levels. MPP1 also directly increases AGTR1 protein in HEK cells, demonstrating MPP1 can upregulate AGTR1 protein levels.\",\n      \"method\": \"Transgenic mouse model (Tg-MPP1), echocardiography, histology, co-localization in vivo, AGTR1eYFP fluorescence measurement in HEK cells\",\n      \"journal\": \"Biochemical pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — transgenic mouse with defined cardiac phenotype plus cell-based validation of AGTR1 upregulation, single lab, co-localization without direct binding assay\",\n      \"pmids\": [\"37683843\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"MPP1 binds USP12 (identified by co-immunoprecipitation and mass spectrometry) and promotes CCL5 expression via the MPP1/USP12/CCL5 cascade in urothelial carcinoma cells, thereby enhancing CD8+ T-cell chemotaxis and inhibiting immune escape.\",\n      \"method\": \"Co-immunoprecipitation, mass spectrometry, RT-qPCR, western blotting, MPP1 overexpression vector, CD8+ T-cell coculture assay, in vivo tumorigenesis in HuNOG mice\",\n      \"journal\": \"International immunopharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus MS identifies binding partner, multiple functional assays in vitro and in vivo, single lab\",\n      \"pmids\": [\"39700963\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Reconstituted MPP1 with recombinant flotillins in giant unilamellar vesicles (GUVs) promotes membrane remodeling and triggers coexistence of liquid-ordered (Lo) and liquid-disordered (Ld) domains; palmitoylation of MPP1 exerts additional influence on membrane organization. Flotillin-MPP1 assemblies are sufficient and necessary to modulate lateral organization of lipid bilayers.\",\n      \"method\": \"Reconstitution in GUVs, FLIM, fluorescence microscopy of phase separation with recombinant proteins\",\n      \"journal\": \"Biochimica et biophysica acta. Biomembranes\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstitution with purified recombinant proteins in minimal membrane system, FLIM functional readout, establishes sufficiency of MPP1-flotillin complex for domain formation\",\n      \"pmids\": [\"41061789\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MPP1/p55 is a palmitoylated MAGUK scaffolding protein that organizes plasma membrane raft nanodomains by directly interacting with flotillins (via a defined D5 domain motif), regulating flotillin nanocluster immobilization and confinement of raft-associated lipids (sphingomyelin, Thy-1); its palmitoylation (written by DHHC17 in erythrocytes) is required for lateral membrane order and DRM formation, and it controls downstream signaling cascades including H-Ras/Raf activation, Akt/PIP3 spatial restriction for neutrophil polarity, RTK/MAP-kinase signaling, and insulin receptor-dependent pathways, while also interacting with MPP5/Pals1 (Crumbs complex), whirlin (Usher network), and NF2/merlin to link polarity and cytoskeletal networks in retina, erythrocytes, and non-erythroid cells.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MPP1/p55 is a palmitoylated MAGUK scaffolding protein that organizes the lateral order of plasma membrane raft nanodomains and couples this organization to receptor signaling and cell polarity [#5, #10]. Its central biochemical activity is a direct, high-affinity interaction with flotillin-1 and flotillin-2 through a defined flotillin-binding motif in the D5 domain; this interaction is distinct from MPP1's protein 4.1-dependent contacts and is required to maintain erythrocyte membrane fluidity and order [#7, #9]. Reconstitution of MPP1 with recombinant flotillins in synthetic vesicles is sufficient to drive coexistence of liquid-ordered and liquid-disordered domains, establishing the MPP1-flotillin assembly as a minimal unit capable of remodeling membrane lateral organization [#13]. Mechanistically, MPP1 captures and temporally immobilizes flotillin-based nanoclusters and confines raft lipids such as sphingomyelin and Thy-1 [#10], and its palmitoylation — written by DHHC17 in red blood cells — is required for detergent-resistant membrane formation and membrane order [#4, #5]. This raft-organizing function underlies MPP1's control of multiple raft-dependent signaling outputs: spatial restriction of PIP3 and downstream Akt activation that drives neutrophil chemotactic polarity [#2], H-Ras GDP/GTP exchange and H-Ras-Raf coupling in insulin receptor signaling [#8, #9], and RTK/MAP-kinase signaling in erythroid cells [#5]. MPP1 additionally functions as a scaffold linking polarity and cytoskeletal networks through directional heterodimerization with MPP5/Pals1 and interaction with whirlin at the retinal outer limiting membrane [#0, #1] and through nanomolar-affinity binding to the FERM domain of NF2/merlin [#3]. Roles in cardiac AGTR1 regulation [#11] and in a USP12/CCL5 axis modulating anti-tumor immunity [#12] have also been described.\",\n  \"teleology\": [\n    {\n      \"year\": 2007,\n      \"claim\": \"Established MPP1 as a scaffolding hub linking distinct polarity and Usher networks, answering how disparate retinal complexes are physically coupled.\",\n      \"evidence\": \"Interaction and domain-mapping assays with co-localization in retinal tissue, showing directional MAGUK heterodimerization with MPP5/Pals1 and dual-mode binding to whirlin\",\n      \"pmids\": [\"17584769\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No reconstitution of the multiprotein complex\", \"Functional consequence of these interactions for polarity not tested by perturbation\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Defined MPP1 as a spatial regulator of PIP3/Akt signaling required for cell polarity, distinguishing localization control from kinase activity itself.\",\n      \"evidence\": \"p55 knockout mouse neutrophils with chemotaxis assays, PIP3 immunolocalization, Akt phosphorylation, and PI3Kgamma activity assays\",\n      \"pmids\": [\"19897731\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism by which MPP1 restricts PIP3 not resolved\", \"Link to raft organization not yet established in this study\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Quantified a direct, high-affinity MPP1-merlin interaction, placing MPP1 in cytoskeletal/tumor-suppressor scaffolding contexts beyond erythrocytes.\",\n      \"evidence\": \"Surface plasmon resonance (KD 3.7 nM) between NF2/merlin FERM domain and erythrocyte p55, with co-localization in Schwann cells\",\n      \"pmids\": [\"19144871\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of the merlin interaction not tested\", \"Single lab, no cellular perturbation\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Identified DHHC17 as the writer of MPP1 palmitoylation and tied this modification causally to membrane order, defining the post-translational requirement for MPP1 raft function.\",\n      \"evidence\": \"Clinical DHHC17-deficient RBCs, 2-bromopalmitate inhibition, FLIM membrane order, and DRM fractionation\",\n      \"pmids\": [\"22496366\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Palmitoylation site(s) on MPP1 not mapped\", \"Whether DHHC17 acts on MPP1 in non-erythroid cells unknown\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Connected MPP1 levels and palmitoylation to raft-dependent RTK/MAP-kinase signaling, extending the membrane-order role to a signaling output.\",\n      \"evidence\": \"MPP1 knockdown and palmitoylation inhibition in HEL erythroid cells with DRM fractionation, FLIM, and MAP-kinase assays\",\n      \"pmids\": [\"23507198\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Which RTKs are affected not enumerated\", \"Direct molecular link between MPP1 and the receptors not shown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Demonstrated MPP1 tunes membrane phase behavior directly, showing domain properties depend on the protein rather than lipid composition changes.\",\n      \"evidence\": \"GPMVs from live cells analyzed for phase separation and miscibility transition temperature\",\n      \"pmids\": [\"25954878\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Binding partner driving phase modulation not identified in this study\", \"Single biophysical method type\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Identified flotillins 1 and 2 as direct MPP1 partners distinct from 4.1-dependent contacts, providing the molecular basis for MPP1's raft-organizing activity.\",\n      \"evidence\": \"Co-IP and pulldown in erythrocyte membranes plus FLIM membrane fluidity readout\",\n      \"pmids\": [\"28865798\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Binding interface not yet mapped at this stage\", \"Stoichiometry of the complex unknown\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Placed MPP1 upstream of H-Ras activation in insulin signaling, linking membrane domain organization to GDP/GTP exchange and effector coupling.\",\n      \"evidence\": \"MPP1 knockdown in HEL cells with H-Ras GTP-loading and Raf interaction assays plus DRM fractionation under insulin stimulation\",\n      \"pmids\": [\"29719614\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism connecting raft order to H-Ras nucleotide exchange not resolved\", \"Single cell system\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Localized the flotillin-binding activity to a specific D5 motif and proved its necessity for raft order and insulin signaling via a dominant-negative peptide.\",\n      \"evidence\": \"Molecular dynamics, SPR affinity measurement, dominant-negative motif-peptide overexpression, FLIM, and insulin receptor signaling assays in erythroid precursors\",\n      \"pmids\": [\"34285255\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of the full MPP1-flotillin complex not solved\", \"Whether the same motif governs non-erythroid functions untested\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Resolved the physical mechanism of raft organization as MPP1-driven immobilization of flotillin nanoclusters and confinement of specific raft lipids.\",\n      \"evidence\": \"Super-resolution structured illumination imaging, FRAP, and spot-variation FCS in erythroid cells measuring mobility and confinement of sphingomyelin and Thy-1\",\n      \"pmids\": [\"35159121\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Kinetics of nanocluster capture not detailed\", \"Generalizability beyond erythroid cells not tested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Extended MPP1 function to cardiac AGTR1 regulation, showing MPP1 dosage controls receptor protein levels and cardiac physiology.\",\n      \"evidence\": \"Transgenic MPP1-overexpressing mice with echocardiography and histology, in vivo co-localization, and AGTR1 protein measurement in HEK cells\",\n      \"pmids\": [\"37683843\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct MPP1-AGTR1 binding assay\", \"Mechanism of AGTR1 upregulation unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Linked MPP1 to immune modulation through a USP12/CCL5 axis affecting T-cell chemotaxis in urothelial carcinoma.\",\n      \"evidence\": \"Co-IP and mass spectrometry identifying USP12 binding, RT-qPCR/western blot, CD8+ T-cell coculture, and HuNOG tumorigenesis assays\",\n      \"pmids\": [\"39700963\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"How MPP1-USP12 binding drives CCL5 expression not mechanistically resolved\", \"Relationship to MPP1's raft-organizing role unclear\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Proved sufficiency: reconstituted MPP1-flotillin assemblies alone generate ordered/disordered domain coexistence in minimal bilayers.\",\n      \"evidence\": \"Reconstitution of recombinant MPP1 and flotillins in GUVs with FLIM and phase-separation microscopy, with palmitoylation modulating organization\",\n      \"pmids\": [\"41061789\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative stoichiometry and structural arrangement in the bilayer not defined\", \"How signaling receptors partition into reconstituted domains not addressed\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How MPP1's raft-organizing scaffold activity is mechanistically reconciled with its discrete roles in AGTR1 regulation and the USP12/CCL5 immune axis remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking membrane-order function to AGTR1/USP12 phenotypes\", \"Structure of MPP1-flotillin complex unsolved\", \"Tissue-specificity of partner usage not mapped\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 1, 3, 7, 9]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [4, 7, 10, 11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [2, 8, 9]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [2, 12]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"FLOT1\", \"FLOT2\", \"MPP5\", \"WHRN\", \"NF2\", \"USP12\", \"AGTR1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":6,"faith_total":7,"faith_pct":85.71428571428571}}