PfMDR1 (P-glycoprotein homologue 1, Pgh1; PF3D7_0523000) is a full-length type-1 ABC transporter of the ABCB/multidrug-resistance-exporter subfamily, built from two transmembrane domains and two cytosolic nucleotide-binding domains. It resides in the membrane of the parasite's digestive (food) vacuole, the acidic, lysosome-like organelle in which endocytosed host haemoglobin is degraded, and is expressed throughout the asexual intraerythrocytic cycle as well as in gametocytes. The protein binds and hydrolyses ATP at its nucleotide-binding domains and couples that hydrolysis to the movement of amphipathic solutes across the vacuolar membrane. Because its nucleotide-binding domains face the parasite cytosol, the transport cycle runs inward with respect to the organelle, pumping substrates from the cytosol into the digestive vacuole lumen rather than out of the cell. Cryo-EM structures show the canonical ABC exporter cycle, from an inward-facing apo state with a large cytosol-open cavity to an ATP-bound, NBD-dimerized state opening toward the vacuolar lumen, together with an unusual N-terminal helical regulatory element that hinders NBD dimerization and damps ATPase activity. PfMDR1 accepts a broad range of amphipathic xenobiotics, including most quinoline and arylamino-alcohol antimalarials and classic human P-glycoprotein substrates; by concentrating such compounds in the vacuole it changes their distribution between vacuole and cytosol, and naturally occurring sequence variants that alter transport capacity shift parasite susceptibility to several antimalarial drugs. Its endogenous physiological substrate is not known.
Definition: The lipid bilayer surrounding the food vacuole, the acidic lysosome-like compartment of apicomplexan parasites in which endocytosed host-cell material such as haemoglobin is digested.
Justification: GO contains GO:0020020 food vacuole but no corresponding membrane class (OLS returns no match for "food vacuole membrane", and GO:0020020 has no children). Integral membrane proteins of the Plasmodium digestive vacuole membrane, of which PfMDR1 and PfCRT are the best characterized and are major antimalarial-resistance determinants, can therefore only be annotated to the generic GO:0005774 vacuolar membrane, losing the apicomplexan-specific compartment that makes their biology interpretable. A dedicated child of GO:0005774 would let the location and the organelle be stated in one term, as is already possible for lysosomal membrane.
Parent term: vacuolar membrane
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005524 ATP binding | IEA GO_REF:0000002 | ACCEPT | Summary: PfMDR1 carries two canonical ABC nucleotide-binding domains; nucleotide binding is directly demonstrated by 8-azido-ATP photoaffinity labelling of the protein in purified digestive vacuoles, and ATP-bound cryo-EM structures resolve the nucleotides at the NBD dimer interface. Reason: The InterPro-based electronic assertion is confirmed by direct experimental evidence on this very protein. Photoaffinity labelling with 8-N3-ATP is competed by ATP, ADP, GTP and GDP but not by the monophosphates, showing genuine nucleotide binding rather than nonspecific labelling, and the cryo-EM work resolves ATP bound at the NBDs with the conserved A-loop and Q-loop contacts. ATP binding is an integral part of the core transport activity, so this is kept as core. Supporting Evidence: PMID:8097560 Inhibition of photoaffinity labeling of PGH1 could be achieved with ATP, ADP, GTP and GDP but not with AMP or GMP. PMID:37527341 Upon ATP binding and dimerization of the nucleotide-binding domains (NBDs), PfMDR1 displays an outward-facing conformation with a cavity toward the DV lumen. |
| GO:0005737 cytoplasm | IEA GO_REF:0000117 | MODIFY | Summary: An ARBA machine-learning assignment of the generic compartment "cytoplasm". PfMDR1 is a multi-pass integral membrane protein of the digestive vacuole membrane, which is a cytoplasmic organelle, so the term is not false but conveys nothing about where the protein acts. Reason: The claim is true only in the trivial sense that the food vacuole lies within the cytoplasm; the evidence supports a far more precise location. Immunofluorescence and immunoelectron microscopy place Pgh1 on the digestive vacuole membrane, and UniProt records the subcellular location as vacuole membrane, multi-pass membrane protein. Replacing with the supported descendant GO:0005774 vacuolar membrane, which the gene already carries, rather than removing a true-but-uninformative row. Proposed replacements: vacuolar membrane Supporting Evidence: PMID:1674943 Immunofluorescence and immunoelectron microscopy has demonstrated that Pgh1 is localized on the membrane of the digestive vacuole of mature parasites. |
| GO:0005774 vacuolar membrane | IEA GO_REF:0000044 | ACCEPT | Summary: Localization to the vacuolar membrane, mapped from the UniProt subcellular location annotation, which is itself backed by experimental evidence on this protein (immunolocalization to the digestive vacuole membrane and photoaffinity labelling of purified vacuoles). Reason: This is the correct and, given the current ontology, the most precise membrane term available for PfMDR1. GO has no "food vacuole membrane" class (checked via OLS; GO:0020020 has no children), so GO:0005774 together with GO:0020020 food vacuole is the best representation of an integral protein of the apicomplexan digestive vacuole membrane. This is a core location. Supporting Evidence: PMID:1674943 localized on the membrane of the digestive vacuole of mature parasites. PMID:35507548 PfMDR1 and PfCRT reside in the membrane of the parasite’s digestive vacuole (DV), the lysosomal-type compartment (pH 5.0 to 5.5) in which many antimalarials accumulate, act, and/or are activated. |
| GO:0008559 ABC-type xenobiotic transporter activity | IEA GO_REF:0000003 | ACCEPT | Summary: The EC 7.6.2.2 mapping asserts ATP-hydrolysis-coupled xenobiotic transport. Direct measurements in a Xenopus oocyte expression system show that PfMDR1 moves a structurally diverse set of amphipathic drugs across a membrane in an ATP-dependent manner, and purified protein hydrolyses ATP. Reason: This is the core molecular function, and it is experimentally grounded for this accession rather than merely inferred from the EC mapping. Wild-type PfMDR1 transports lumefantrine, mefloquine, chloroquine, quinine, quinidine, amodiaquine, piperaquine, dihydroartemisinin and quinacrine, as well as the human P-glycoprotein substrates rhodamine B and vinblastine, while failing to transport amantadine, and transport is stimulated by injected ATP. UniProt records the corresponding experimental catalytic activity, EC 7.6.2.2, for this entry. Supporting Evidence: PMID:35507548 We show that wild-type PfMDR1 transports diverse pharmacons, including lumefantrine, mefloquine, dihydroartemisinin, piperaquine, amodiaquine, methylene blue, and chloroquine (but not the antiviral drug amantadine). file:PLAF7/MDR1/MDR1-uniprot.txt Reaction=ATP + H2O + xenobioticSide 1 = ADP + phosphate + xenobioticSide 2. |
| GO:0009410 response to xenobiotic stimulus | IDA PMID:10706290 Pgh1 modulates sensitivity and resistance to multiple antima... | MODIFY | Summary: Allelic-exchange experiments showed that mutations in Pgh1 confer resistance to mefloquine, quinine and halofantrine and modulate chloroquine and artemisinin susceptibility. This establishes that the protein determines how the parasite handles these drugs, but what PfMDR1 itself executes is transmembrane drug transport, not a cellular response program. Reason: The essence of the annotation is sound and the underlying experiment is strong, but "response to xenobiotic stimulus" describes a change in cell state triggered by a xenobiotic, whereas the step PfMDR1 performs is ATP-driven movement of the drug across the digestive vacuole membrane. Later direct transport measurements identify that step explicitly. Replacing with GO:0006855 xenobiotic transmembrane transport (verified via OLS as a descendant of both GO:0042908 and GO:0055085), which states what this gene product does and still captures the drug handling the IDA demonstrates. No part of the claim is being discarded. Proposed replacements: xenobiotic transmembrane transport Supporting Evidence: PMID:10706290 mutations in Pgh1 can confer resistance to mefloquine, quinine and halofantrine. PMID:35507548 Lumefantrine, mefloquine, and chloroquine are weak bases that enter the DV via 2 main routes: (1) simple diffusion of the neutral species across the membrane and subsequent protonation within the acidic DV lumen; and (2) ATP-driven import via PfMDR1. |
| GO:0016020 membrane | IBA GO_REF:0000033 | MODIFY | Summary: Phylogenetic inference that the ancestral ABCB/MDR node acts in a membrane. The node placement is sound and PfMDR1 is unambiguously inside that clade, being a full-length type-1 ABC exporter with twelve predicted transmembrane helices; the only issue is specificity. Reason: The IBA is correct about the gene product acting in a membrane, and the ancestral-node judgment is not in dispute. It is, however, less specific than direct evidence allows for this protein, which is localized experimentally to the digestive vacuole membrane. Refining to GO:0005774 vacuolar membrane, an is_a descendant of membrane that the gene already carries from UniProt subcellular location. The same replacement is applied to the IEA row for this term. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN002517083 SUPPORTS TRANSFER Ancestral ABCB/MDR node; membrane localization is correct for every descendant including PfMDR1, so the node placement is not at issue, only the granularity of the term for this target. Proposed replacements: vacuolar membrane Supporting Evidence: PMID:1674943 localized on the membrane of the digestive vacuole of mature parasites. |
| GO:0016020 membrane | IEA GO_REF:0000002 | MODIFY | Summary: InterPro ABC transporter transmembrane-domain signatures imply membrane localization. True, and matched by twelve predicted transmembrane segments in the UniProt record, but uninformative given the experimentally determined compartment. Reason: Correct but general. The protein is an integral component of a specific organellar membrane, demonstrated by immunoelectron microscopy and by photoaffinity labelling of purified digestive vacuoles. Refined to GO:0005774 vacuolar membrane, consistent with the action taken on the IBA row carrying the same term. Proposed replacements: vacuolar membrane Supporting Evidence: PMID:1674943 localized on the membrane of the digestive vacuole of mature parasites. |
| GO:0016887 ATP hydrolysis activity | IEA GO_REF:0000002 | ACCEPT | Summary: ATPase activity is inferred from the ABC nucleotide-binding-domain signature and confirmed directly. Purified PfMDR1 hydrolyses ATP, the activity is abolished by mutating the two catalytic glutamates, and it is modulated by drugs and by resistance-associated substitutions. Reason: Directly measured on purified protein rather than merely inferred. The wild-type protein shows substantial ATPase activity that is lost in the E588Q/E1337Q catalytic mutant, and UniProt records a KM of 0.33 mM for ATP with a Vmax of 198.3 nmol/min/mg. ATP hydrolysis is the energizing half-reaction of the core transport activity, so this is retained as core. Supporting Evidence: PMID:37527341 To confirm that the PfMDR1 protein purified in a detergent solution is functionally active, we measured the ATP hydrolysis activity at different ATP concentrations. PMID:37527341 Compared to the EQ mutant protein in which the key catalytic glutamates were mutated to glutamines, the wild-type PfMDR1 protein showed significant ATPase activity. |
| GO:0020020 food vacuole | IDA PMID:1674943 A P-glycoprotein homologue of Plasmodium falciparum is local... | ACCEPT | Summary: Immunofluorescence and immunoelectron microscopy localize Pgh1 to the membrane of the digestive vacuole of mature asexual parasites; the food vacuole is the apicomplexan-specific acidic compartment in which host haemoglobin is digested and in which many antimalarials accumulate. Reason: A correct, organism-appropriate organelle term supported by direct experimental localization and independently confirmed by biochemical labelling of purified digestive vacuoles and by two subsequent structural and transport studies. This is the core site of action, and GO offers no more specific food-vacuole membrane class, so the term is kept as is and paired with GO:0005774. Supporting Evidence: PMID:1674943 Immunofluorescence and immunoelectron microscopy has demonstrated that Pgh1 is localized on the membrane of the digestive vacuole of mature parasites. PMID:8097560 involved in a nucleotide-regulated transport function across the membrane of the digestive vacuole. |
| GO:0042626 ATPase-coupled transmembrane transporter activity | IBA GO_REF:0000033 | MODIFY | Summary: Phylogenetic inference of primary active transport for the ABCB/MDR ancestral node. Entirely correct for PfMDR1, which hydrolyses ATP and uses that hydrolysis to move drugs across the digestive vacuole membrane, but one level above the activity the direct evidence pins down. Reason: Not a challenge to the node placement, which is well supported and includes PfMDR1 squarely within the ABCB exporter clade. The specific activity has since been measured for this protein. It transports a broad range of amphipathic xenobiotics in an ATP-dependent manner, and UniProt records the EC 7.6.2.2 catalytic activity experimentally. Refining to the is_a descendant GO:0008559 ABC-type xenobiotic transporter activity, which the gene already carries. The same replacement is applied to the GO:0140359 row. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN000657189 SUPPORTS TRANSFER Ancestral ABCB exporter node; primary active transport is correctly inherited by PfMDR1, and the substrate class is simply better defined for this target than at the node. Proposed replacements: ABC-type xenobiotic transporter activity Supporting Evidence: PMID:35507548 We show that wild-type PfMDR1 transports diverse pharmacons, including lumefantrine, mefloquine, dihydroartemisinin, piperaquine, amodiaquine, methylene blue, and chloroquine (but not the antiviral drug amantadine). |
| GO:0042908 xenobiotic transport | IEA GO_REF:0000108 | MODIFY | Summary: Logically inferred from the ABC-type xenobiotic transporter activity term. The process claim is right, and the direct evidence additionally establishes that the transport is across a lipid bilayer, specifically the digestive vacuole membrane. Reason: True but less specific than warranted. GO:0006855 xenobiotic transmembrane transport is an is_a descendant of this term (verified via OLS) and is exactly what the oocyte transport assays demonstrate, namely ATP-driven movement of drugs from the cytosolic side into the vacuolar lumen across the vacuole membrane. Refining to that term, consistently with the other process rows in this review. Proposed replacements: xenobiotic transmembrane transport Supporting Evidence: PMID:35507548 Lumefantrine, mefloquine, and chloroquine are weak bases that enter the DV via 2 main routes: (1) simple diffusion of the neutral species across the membrane and subsequent protonation within the acidic DV lumen; and (2) ATP-driven import via PfMDR1. |
| GO:0055085 transmembrane transport | IBA GO_REF:0000033 | MODIFY | Summary: Phylogenetic inference that members of this ABCB clade participate in transmembrane transport. Correct for PfMDR1, whose transport across the digestive vacuole membrane has since been measured directly. Reason: The node placement is sound and PfMDR1 belongs inside the clade; the term is simply broader than the evidence now supports. Refining to GO:0006855 xenobiotic transmembrane transport, an is_a descendant of transmembrane transport, which matches the demonstrated substrates. Applied identically to the IEA row carrying the same term, and consistent with the GO:0042908 and GO:0009410 refinements. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN000657189 SUPPORTS TRANSFER Ancestral ABCB exporter node; transmembrane transport is correctly inherited, and the xenobiotic substrate class is established directly for PfMDR1 rather than being in dispute at the node. Proposed replacements: xenobiotic transmembrane transport Supporting Evidence: PMID:35507548 We show that wild-type PfMDR1 transports diverse pharmacons, including lumefantrine, mefloquine, dihydroartemisinin, piperaquine, amodiaquine, methylene blue, and chloroquine (but not the antiviral drug amantadine). |
| GO:0055085 transmembrane transport | IEA GO_REF:0000002 | MODIFY | Summary: InterPro-derived process assertion mirroring the IBA row. True of PfMDR1 but not as specific as the measured activity. Reason: Correct but general; refined to GO:0006855 xenobiotic transmembrane transport for the same reasons as the IBA row carrying this term, keeping the two rows consistent. Proposed replacements: xenobiotic transmembrane transport Supporting Evidence: PMID:35507548 Lumefantrine, mefloquine, and chloroquine are weak bases that enter the DV via 2 main routes: (1) simple diffusion of the neutral species across the membrane and subsequent protonation within the acidic DV lumen; and (2) ATP-driven import via PfMDR1. |
| GO:0140359 ABC-type transporter activity | IEA GO_REF:0000002 | MODIFY | Summary: Generic ABC-type transporter activity from the InterPro ABC transmembrane domain signature. Correct at the family level; the substrate class is now known for this protein. Reason: The parent term is right but the evidence supports the specific child. PfMDR1 transports amphipathic xenobiotics including most quinoline and arylamino-alcohol antimalarials, and UniProt assigns the experimental catalytic activity EC 7.6.2.2. Refining to GO:0008559 ABC-type xenobiotic transporter activity, an is_a descendant that the gene already carries, consistently with the GO:0042626 row. Proposed replacements: ABC-type xenobiotic transporter activity Supporting Evidence: PMID:35507548 We show that wild-type PfMDR1 transports diverse pharmacons, including lumefantrine, mefloquine, dihydroartemisinin, piperaquine, amodiaquine, methylene blue, and chloroquine (but not the antiviral drug amantadine). |
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Download this section (compressed HTML)Q: What is the endogenous substrate of PfMDR1? Given that its digestive vacuole neighbour PfCRT exports haemoglobin-derived peptides, is the natural cargo a product of haemoglobin digestion, a lipid, or a parasite metabolite?
Suggested experts: Rowena E. Martin, Sarah H. Shafik
Q: What relieves the autoinhibition imposed by the N-terminal regulatory element in vivo? Unlike the CFTR R domain it lacks phosphorylation sites, so is regulation by ligand binding, pH, or proteolysis?
Suggested experts: Maojun Li, Kun Si
Q: Is the direction of transport in the parasite strictly cytosol-to-lumen for all substrates, and does the acidic vacuolar pH contribute to net accumulation independently of the pump?
Q: Does PfMDR1 copy-number amplification alter any process beyond drug distribution, for example haemoglobin catabolism or vacuolar homeostasis, that would justify an additional biological process annotation?
Experiment: Perform untargeted metabolomic and peptidomic comparison of isolated digestive vacuoles and cytosolic fractions from parasites with conditional knockdown of pfmdr1 versus controls, and test candidate hits directly in the established Xenopus oocyte PfMDR1 expression system for saturable, ATP-dependent uptake.
Hypothesis: PfMDR1 transports a product of host haemoglobin digestion, analogous to the peptide-export role of PfCRT.
Type: conditional knockdown plus organellar metabolomics and heterologous transport assay
Experiment: Generate parasite lines expressing the N-terminally truncated transporter at the endogenous locus and compare vacuolar accumulation of radiolabelled mefloquine and lumefantrine, drug susceptibility, and growth rate against isogenic wild-type parasites.
Hypothesis: The N-terminal regulatory helices autoinhibit PfMDR1 in the parasite, and their removal increases drug sequestration in the digestive vacuole.
Type: allelic replacement with drug accumulation assays
Experiment: Use a conditional degradation or DiCre excision system to deplete PfMDR1 in the absence of drug pressure and quantify the effect on intraerythrocytic development, haemozoin formation, and digestive vacuole morphology by electron microscopy.
Hypothesis: PfMDR1 is essential for asexual blood-stage growth independently of drug exposure.
Type: conditional protein depletion with phenotypic profiling
What is not known — curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: No clean genetic test of pfmdr1 essentiality or of its drug-independent cellular role has been reported; essentiality is asserted in reviews rather than demonstrated by conditional depletion in the literature read here.
BIOLOGY BP_DARK
What is known: Copy-number variation and point mutations at this locus are extensively characterized and demonstrably shift antimalarial susceptibility, and the protein's biochemistry and structure are solved. What is missing is the phenotype of acute loss of the protein in the absence of drug pressure.
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