Affinage

ATP8A1

Phospholipid-transporting ATPase IA · UniProt Q9Y2Q0

Length
1164 aa
Mass
131.4 kDa
Annotated
2026-06-09
16 papers in source corpus 12 papers cited in narrative 12 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 7/8 claims corpus-supported (88%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

ATP8A1 is a P4-type ATPase phospholipid flippase that uses vanadate-sensitive, phosphorylation-dependent ATP hydrolysis to translocate aminophospholipids from the exoplasmic/luminal leaflet to the cytoplasmic leaflet of cellular membranes, with phosphatidylserine (PS) serving as the principal activating substrate and phosphatidylethanolamine (PE) as a secondary one (PMID:16618126, PMID:20224745, PMID:22007859). It functions as a heteromeric complex with the CDC50A chaperone, which associates with ATP8A1 and recruits it to the plasma membrane (PMID:23269685). Catalysis requires an intact P-type phosphorylation site, as mutation of this residue abolishes translocase activity and instead causes PS externalization (PMID:22007859). Through this lipid-flipping activity ATP8A1 controls diverse membrane-dependent processes: it maintains plasma membrane aminophospholipid asymmetry in hippocampal neurons (PMID:22007859), drives PE-dependent membrane ruffling and cell migration (PMID:23269685), and on a synaptic-vesicle subset translocates PS to the cytoplasmic face to recruit synapsin and sustain high-frequency neurotransmitter release (PMID:37723322). Its subcellular targeting is governed by the AP-3 adaptor, which recognizes a C-terminal dileucine signal to sort ATP8A1 from endosomes to lamellar bodies and synaptic vesicles (PMID:33990468, PMID:37723322). ATP8A1 also acts at the trans-Golgi network, where its cytosolic C-terminal tail binds and stimulates the Sec7 domain of the ARF-GEFs BIG1 and BIG2 to promote ARF activation and AP1/GGA2/clathrin coat recruitment (PMID:38879142), and at Rab7-positive late endosomes, where it regulates PS topology on multivesicular body limiting membranes to control ESCRT recruitment, intraluminal vesicle cargo sorting, and EGFR signaling (PMID:40083718). Loss of ATP8A1 lipid asymmetry activity feeds into downstream signaling, modulating PTEN/PI3K-AKT-mTORC1 and YAP1 outputs that govern hematopoietic stem cell proliferation and AT2 cell behavior (PMID:33990468, PMID:36930333). During platelet apoptosis, the cysteine protease calpain cleaves and inactivates ATP8A1, permitting sustained PS exposure (PMID:30674456).

Mechanistic history

Synthesis pass · year-by-year structured walk · 12 steps
  1. 2006 High

    Established that ATP8A1 is a P-type ATPase whose activity is specifically driven by phosphatidylserine, defining its substrate preference at the biochemical level.

    Evidence In vitro ATPase assay with purified Atp8a1 from insect cells against a panel of phospholipid structural variants

    PMID:16618126

    Open questions at the time
    • ATPase stimulation alone does not demonstrate vectorial lipid transport
    • no in vivo membrane context
    • CDC50A requirement not yet known
  2. 2008 Medium

    Demonstrated that ATP8A1 is a bona fide ATP-dependent flippase that vectorially translocates PS across a membrane, moving beyond ATPase stimulation to actual transport.

    Evidence PS translocation assay in purified yeast secretory vesicles expressing Atp8a1; ATPase assay; RBC membrane fractionation

    PMID:20224745

    Open questions at the time
    • heterologous yeast system may not reflect native partner requirements
    • single lab
    • physiological membrane localization unresolved
  3. 2011 High

    Linked ATP8A1 catalytic mechanism to a physiological function by showing its phosphorylation site is required for plasma membrane aminophospholipid translocase activity and PS asymmetry in neurons.

    Evidence Atp8a1 knockout mice with annexin V PS-externalization readout; phosphorylation-site mutagenesis and APLT kinetics in N18 neuronal cells

    PMID:22007859

    Open questions at the time
    • PS externalization in knockout did not increase apoptosis, leaving downstream consequences unclear
    • chaperone subunit not addressed
    • did not distinguish direct vs indirect contribution to APLT
  4. 2012 High

    Identified CDC50A as the obligate partner that recruits ATP8A1 to the plasma membrane and resolved a distinct PE-translocation role driving membrane ruffling and migration.

    Evidence Co-IP, siRNA knockdown, fluorescent-lipid translocation assays, and ruffle/migration assays in CHO cells with PE-binding peptide and PE-synthesis mutants

    PMID:23269685

    Open questions at the time
    • PE versus PS substrate split across studies/systems not fully reconciled
    • stoichiometry of the ATP8A1-CDC50A complex not defined here
    • in vivo migration relevance untested
  5. 2019 Medium

    Showed how ATP8A1 activity is terminated during apoptosis, identifying calpain cleavage as the switch permitting sustained platelet PS exposure.

    Evidence Western blotting of platelet fractions with calpain/caspase inhibitors and calcium chelation across apoptotic versus thrombin/collagen activation

    PMID:30674456

    Open questions at the time
    • cleavage site not mapped
    • single lab
    • direct demonstration that cleavage abolishes flippase activity not shown
  6. 2021 High

    Defined the trafficking logic of ATP8A1, showing AP-3 recognizes a C-terminal dileucine signal to deliver it to lamellar bodies and linking mislocalization to PS topology and YAP-driven migration.

    Evidence Dileucine mutagenesis, Co-IP, fractionation, PS-exposure and YAP reporter assays in AP-3-deficient (HPS2) AT2 cell models

    PMID:33990468

    Open questions at the time
    • mechanism coupling cytosolic PS to YAP activation not resolved
    • single tissue context
    • did not test other adaptor contributions
  7. 2021 Medium

    Tested whether the ATP8A1-IFT27 interaction confers a reproductive function, establishing ATP8A1 is dispensable for spermatogenesis despite being IFT27's strongest binding partner.

    Evidence Pulldown interaction screening plus Atp8a1 knockout mouse fertility, sperm, and histology analysis

    PMID:33821543

    Open questions at the time
    • functional meaning of the IFT27 interaction unexplained
    • possible redundancy with paralogs not excluded
    • no ciliary phenotype assessment beyond reproduction
  8. 2023 High

    Connected AP-3 targeting to a defined neuronal function, showing ATP8A1 flips PS on a synaptic-vesicle subset to recruit synapsin and enable high-frequency release.

    Evidence AP-3 SV proteomics, Atp8a1 knockout mice, hippocampal slice electrophysiology, and synapsin recruitment/live SV imaging

    PMID:37723322

    Open questions at the time
    • how synapsin reads cytoplasmic-leaflet PS mechanistically not detailed
    • fraction of SVs carrying ATP8A1 not quantified structurally
    • interplay with other SV flippases unknown
  9. 2023 Medium

    Extended ATP8A1's role to hematopoietic stem cells, linking loss of PS asymmetry to PTEN/PI3K-AKT-mTORC1 and YAP1 signaling that boosts HSC proliferation and repopulation.

    Evidence Atp8a1 knockout mouse with flow cytometry, competitive transplantation, RNA-seq, and pathway western blots

    PMID:36930333

    Open questions at the time
    • direct mechanistic link between membrane PS and PTEN regulation not established
    • single lab
    • causality versus correlation among pathway changes unresolved
  10. 2023 Low

    Provided a computational model of the transport cycle, placing phospholipid binding at the E2P to E2Pi-PL transition and highlighting transmembrane electrostatics.

    Evidence Coarse-grained molecular dynamics and binding free energy calculations on the cryo-EM ATP8A1-CDC50 structure

    PMID:36831082

    Open questions at the time
    • computational only, no experimental validation of predicted residues or states
    • predicted key residues not mutationally tested
    • model assumes the cryo-EM conformations sampled
  11. 2024 Medium

    Revealed a moonlighting role at the TGN in which the ATP8A1 cytosolic tail stimulates BIG1/BIG2 GEF activity to couple lipid flipping to ARF activation and vesicle coat assembly.

    Evidence Reciprocal Co-IP, confocal co-localization, tail-deletion mutagenesis, ARF activation assays, and AP1/GGA2/clathrin recruitment quantification

    PMID:38879142

    Open questions at the time
    • whether GEF stimulation requires catalytic flippase activity untested
    • single lab
    • in vivo TGN trafficking consequence not shown
  12. 2025 Medium

    Established a late-endosomal function, showing ATP8A1 controls MVB-limiting-membrane PS topology to regulate ESCRT recruitment, ILV cargo sorting, and EGFR signaling.

    Evidence siRNA knockdown, GFP-LactC2 PS biosensor, fractionation, cargo trafficking and ESCRT recruitment assays, EGFR western blot

    PMID:40083718

    Open questions at the time
    • mechanism by which luminal-leaflet PS recruits ESCRT not defined
    • single lab
    • relationship to AP-3-dependent endosomal sorting not integrated

Open questions

Synthesis pass · forward-looking unresolved questions
  • How a single flippase coordinates its distinct PS- versus PE-flipping activities across plasma membrane, TGN, endosomes, and synaptic vesicles, and how cytosolic-leaflet aminophospholipid changes are transduced into PTEN/YAP/synapsin/ESCRT outputs, remains unresolved.
  • unified structural basis for substrate switching not established
  • direct molecular sensors of cytosolic PS/PE in each pathway unidentified
  • tissue-specific phenotypic hierarchy of ATP8A1 loss not defined

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0140104 molecular carrier activity 4 GO:0140657 ATP-dependent activity 3 GO:0008289 lipid binding 2 GO:0016787 hydrolase activity 2 GO:0098772 molecular function regulator activity 1
Localization
GO:0005768 endosome 2 GO:0005886 plasma membrane 2 GO:0005764 lysosome 1 GO:0005794 Golgi apparatus 1 GO:0031410 cytoplasmic vesicle 1
Pathway
R-HSA-5653656 Vesicle-mediated transport 3 R-HSA-112316 Neuronal System 2 R-HSA-162582 Signal Transduction 2 R-HSA-9609507 Protein localization 2
Partners
BIG1BIG2CDC50AIFT27
Complex memberships
ATP8A1-CDC50A flippase complex

Evidence

Reading pass · 12 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2006 Purified murine Atp8a1 (ATPase II) is maximally activated by phosphatidylserine (PS) in a manner dependent on the sn-1,2-glycerol stereoisomer and multiple elements of the PS headgroup structure, is minimally activated by PE and phosphatidylglycerol, and is inactive in phosphatidylcholine, phosphatidic acid, or phosphatidylinositol micelles; its selectivity profile mirrors but is distinct from the plasma membrane PS flippase, and it is vanadate-sensitive, consistent with P-type ATPase mechanism. In vitro ATPase activity assay with purified Atp8a1 expressed in insect cells, tested against a panel of phospholipid structural variants Biochemistry High 16618126
2008 Atp8a1 is expressed in red blood cell precursors and is present in mature RBC membranes; its flippase activity was established in purified yeast secretory vesicles where it translocates PS across the vesicle membrane in an ATP-dependent manner, and its ATPase activity is stimulated by PS and PE. In vitro PS translocation assay in purified Saccharomyces cerevisiae secretory vesicles expressing Atp8a1; ATPase activity assay; membrane fractionation Journal of receptor, ligand and channel research Medium 20224745
2011 Atp8a1 is required for plasma membrane aminophospholipid translocase (APLT) activity in hippocampal neurons; Atp8a1 knockout mice show dramatic PS externalization in dentate gyrus, CA1, and CA3 cells without increased apoptosis; ectopic expression of wild-type Atp8a1 (but not a P-type phosphorylation-site mutant) increases the Vmax of PM-APLT activity in neuronal N18 cells, and expression of the phosphorylation-site mutant causes PS externalization. Atp8a1 knockout mouse model; annexin V-based PS externalization assay; ectopic expression and phosphorylation-site mutagenesis in N18 neuronal cells; APLT kinetic assays (Vmax, Km) Journal of neurochemistry High 22007859
2012 ATP8A1 forms a phospholipid flippase complex with CDC50A; CDC50A associates with ATP8A1 and recruits it to the plasma membrane; depletion of ATP8A1 specifically inhibits inward translocation of PE (but not PS) at the plasma membrane of CHO cells, impairs membrane ruffle formation, and severely reduces cell migration. Co-immunoprecipitation; siRNA knockdown; phospholipid translocation assay (fluorescent lipid analogs); cell spreading, ruffle formation, and migration assays; PE-binding peptide and PE-synthesis-defective mutant cell lines The Journal of biological chemistry High 23269685
2019 ATP8A1 is highly expressed in murine and human platelets but is not present in the plasma membrane; during apoptosis, ATP8A1 is cleaved by the cysteine protease calpain, and this cleavage is indirectly prevented by caspase inhibition through blockage of calcium influx and subsequent calpain activation. In contrast, ATP8A1 remains intact in platelets activated with thrombin and collagen that also expose PS. Western blotting of platelet fractions; calpain inhibitor and caspase inhibitor treatment; calcium chelation; immunofluorescence/subcellular fractionation Blood advances Medium 30674456
2021 In alveolar type 2 (AT2) cells, AP-3 sorts ATP8A1 from early endosomes to lamellar bodies (lysosome-related organelles) through recognition of a C-terminal dileucine-based signal on ATP8A1; disruption of the AP-3/ATP8A1 interaction causes ATP8A1 accumulation in early/recycling endosomes, increases phosphatidylserine exposure on the cytosolic leaflet, and activates Yes-associated protein (YAP), augmenting cell migration and AT2 cell numbers. Mutagenesis of dileucine signal; co-immunoprecipitation; subcellular fractionation and live imaging; PS exposure assay; YAP activity reporter; siRNA knockdown; AP-3-deficient (HPS2) cell models Proceedings of the National Academy of Sciences of the United States of America High 33990468
2023 AP-3 targets the phospholipid flippase ATP8A1 to a subset of synaptic vesicles (SVs) in mouse hippocampal neurons; ATP8A1 on these SVs translocates PS to the cytoplasmic face, which recruits synapsin to that SV subset; loss of ATP8A1 recapitulates the high-frequency stimulation-specific SV mobilization defect seen with AP-3 loss, establishing that ATP8A1-mediated PS translocation and consequent synapsin recruitment enables high-frequency neurotransmitter release. AP-3 SV proteomics (mass spectrometry); ATP8A1 knockout mice; electrophysiology of hippocampal slices (high-frequency stimulation); synapsin recruitment assay; live imaging of SV dynamics Nature neuroscience High 37723322
2023 Using coarse-grained molecular dynamics and binding free energy calculations on the ATP8A1-CDC50 complex, phospholipid binding to the transporter occurs early in the transport cycle when ATP8A1-CDC50 transitions from E2P to E2Pi-PL state, and electrostatic interactions of key transmembrane residues are critical drivers of the phospholipid transport free energy landscape. Coarse-grained molecular simulation; binding free energy calculations on cryo-EM-derived ATP8A1-CDC50 structure Biomedicines Low 36831082
2023 Atp8a1 knockout in mice causes loss of plasma membrane PS asymmetry in hematopoietic stem cells (HSCs), leading to decreased PTEN protein levels, activation of PI3K-AKT-mTORC1 signaling, increased JNK/AP-1 activity, and YAP1 phosphorylation changes, which collectively increase HSC proliferative activity and repopulation capacity. Atp8a1 knockout mouse; flow cytometry; competitive bone marrow transplantation; 5-FU stress assay; RNA sequencing; western blotting for PTEN, AKT, mTOR, JNK, YAP1; comet assay and immunofluorescence for DNA damage Cellular oncology Medium 36930333
2024 ATP8A1 co-localizes with BIG1 and BIG2 ARF-GEFs at the trans-Golgi Network (TGN); the cytosolic C-terminal tail of ATP8A1 binds the catalytic Sec7 domain of BIG1 and BIG2; expression of ATP8A1 (but not a C-terminal tail deletion mutant) increases generation of activated ARFs at the TGN and selectively increases recruitment of AP1, GGA2, and clathrin to TGN membranes, suggesting the ATP8A1 tail stimulates BIG1/BIG2 GEF catalytic activity to couple membrane deformation with vesicle coat assembly. Co-immunoprecipitation; co-localization by confocal microscopy; expression of wild-type vs. tail-deletion mutant ATP8A1; ARF activation assay; quantification of AP1/GGA2/clathrin recruitment by immunofluorescence Archives of biochemistry and biophysics Medium 38879142
2025 ATP8A1 is enriched in Rab7-positive late endosomal compartments and preferentially flips PS from the luminal to the cytosolic leaflet of endosomal membranes (but not the inner leaflet of the plasma membrane); ATP8A1 depletion accelerates cargo transfer into intraluminal vesicles (ILVs) of multivesicular bodies (MVBs), alters EGFR signaling, and promotes ESCRT component recruitment by increasing luminal-leaflet PS loading on MVB limiting membranes. siRNA knockdown; PS biosensor (GFP-LactC2); subcellular fractionation; live imaging; cargo trafficking assays; EGFR signaling (western blot); ESCRT recruitment by immunofluorescence iScience Medium 40083718
2021 ATP8A1 is identified as the strongest binding partner of IFT27 by pulldown/interaction screening; however, global Atp8a1 knockout mice are fully fertile with normal sperm count, motility, and testis/epididymis histology, demonstrating ATP8A1 is dispensable for spermatogenesis despite this interaction. Co-immunoprecipitation/binding partner identification; Atp8a1 knockout mouse; sperm count and motility analysis; histology Molecular reproduction and development Medium 33821543

Source papers

Stage 0 corpus · 16 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2012 Role for phospholipid flippase complex of ATP8A1 and CDC50A proteins in cell migration. The Journal of biological chemistry 76 23269685
2015 MiR-140-3p suppressed cell growth and invasion by downregulating the expression of ATP8A1 in non-small cell lung cancer. Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine 64 26415732
2006 Lipid specific activation of the murine P4-ATPase Atp8a1 (ATPase II). Biochemistry 64 16618126
2011 Atp8a1 deficiency is associated with phosphatidylserine externalization in hippocampus and delayed hippocampus-dependent learning. Journal of neurochemistry 60 22007859
2021 AP-3-dependent targeting of flippase ATP8A1 to lamellar bodies suppresses activation of YAP in alveolar epithelial type 2 cells. Proceedings of the National Academy of Sciences of the United States of America 32 33990468
2008 ATP8A1 activity and phosphatidylserine transbilayer movement. Journal of receptor, ligand and channel research 23 20224745
2016 Aberrant hippocampal Atp8a1 levels are associated with altered synaptic strength, electrical activity, and autistic-like behavior. Biochimica et biophysica acta 19 27287255
2019 Calpain cleaves phospholipid flippase ATP8A1 during apoptosis in platelets. Blood advances 13 30674456
2023 Adaptor protein AP-3 produces synaptic vesicles that release at high frequency by recruiting phospholipid flippase ATP8A1. Nature neuroscience 10 37723322
2017 The role of ATP8A1 in non-small cell lung cancer. International journal of clinical and experimental pathology 9 31966623
2023 Atp8a1 deletion increases the proliferative activity of hematopoietic stem cells by impairing PTEN function. Cellular oncology (Dordrecht, Netherlands) 6 36930333
2023 Exploring the Phospholipid Transport Mechanism of ATP8A1-CDC50. Biomedicines 4 36831082
2021 ATP8a1, an IFT27 binding partner, is dispensable for spermatogenesis and male fertility. Molecular reproduction and development 4 33821543
2024 The lipid flippase ATP8A1 regulates the recruitment of ARF effectors to the trans-Golgi Network. Archives of biochemistry and biophysics 3 38879142
2025 circRNA_Atp8a1 Promotes Glycolytic Reprogramming in Damage of Intestinal Mucosal Barrier by Upregulating IGF2 through miR-200b-3p. Tissue engineering and regenerative medicine 2 40679788
2025 ATP8A1-translocated endosomal phosphatidylserine fine-tunes the multivesicular body formation and the endo-lysosomal traffic. iScience 0 40083718

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