PfCRT (Q8IBZ9, PF3D7_0709000) — curation notes

Provenance note: automated deep research was unavailable in this environment
(Falcon auth failure, invalid OpenAI key), so no *-deep-research-*.md file was
generated. What follows is a manual literature review performed by reading the
UniProt flat file, the cached publications under publications/, and additional
primary papers fetched into the cache with ai-gene-review fetch-pmid.

1. Identity and family

• UniProt CRT_PLAF7 / Q8IBZ9, 424 aa, 10-TM integral membrane protein of the
  digestive-vacuole (food-vacuole) membrane of Plasmodium falciparum 3D7.
• Family assignment is bioinformatic: CRT belongs to a previously undefined family
  within the drug/metabolite transporter (DMT) superfamily
  PMID:15240840. That
  paper is explicitly a sequence analysis, not an experimental study
  [PMID:15240840 "The function of CRT is not known and the protein was originally described as a"
  transporter simply because it possesses 10 transmembrane domains].
• 3D7 is the chloroquine-sensitive wild-type allele; the resistance-associated
  isoforms (Dd2, 7G8, Ecu1110, carrying K76T etc.) are different alleles of the same
  gene. Several annotations on Q8IBZ9 derive from experiments on mutant isoforms,
  which matters for how strongly they apply to the reference (3D7) protein.

2. Localization

• The founding genetic/transfection paper localized the protein to the digestive
  vacuole by immuno-labelling
  PMID:11090624.
• UniProt records "Vacuole membrane ... Multi-pass membrane protein" and notes
  "Localizes to the parasite digestive vacuole, the site of chloroquine action."
• Trafficking to the DV membrane is signal-dependent and the protein is a
  phosphoprotein; T416 phosphorylation is implicated in sorting
  PMID:20015114.
  That study is the source of the ER (GO:0005783) IDA — an early biosynthetic
  transit compartment, not a functional site.
• GO has GO:0020020 food vacuole ("Vacuole within a parasite used for digestion of
  the host cell cytoplasm") but no "food vacuole membrane" term; the generic
  GO:0005774 vacuolar membrane is currently the most precise membrane term
  available (see proposed new term).

3. Native (physiological) function

The natural substrate question was unresolved for two decades; heterologous systems
gave contradictory answers (Cl- channel, H+ pump, cation channel, GSH transporter,
Fe2+/Fe3+ transporter) — enumerated and criticized in
PMID:32764664.

The best-supported current model: PfCRT exports host-haemoglobin-derived peptides
of 4–11 residues from the DV lumen to the parasite cytosol:

• PMID:32764664
• [PMID:32764664 "The transport of peptides and peptide mimics via PfCRT is saturable, blocked by known PfCRT inhibitors, and is dependent on protons as well as on a co-substrate that remains to b"e identified]
• Physiological consequence: PMID:32764664

An earlier proteoliposome study with purified protein reported broader polyspecific,
proton-gradient-driven export, including basic amino acids and polyamines
PMID:25733858.
UniProt's FUNCTION/CATALYTIC ACTIVITY blocks (arginine, lysine, histidine, histamine,
spermidine) come from this paper. Shafik et al. re-screened metabolites in oocytes and
in parasites and did not reproduce non-peptide transport, so the amino-acid/polyamine
substrates are treated here as in-vitro findings not yet corroborated in situ; peptide
export is the claim supported both in vitro and in situ.

Loss/perturbation phenotypes support the osmotic-homeostasis role: PfCRT mutations
C101F and L272F produce grossly swollen vacuoles
PMID:26420308, and the same
mutations cut chloroquine transport
PMID:26420308.

4. Drug (xenobiotic) transport

• Resistance mechanism: mutant PfCRT effluxes chloroquine out of the DV, away from its
  haem target. Wild-type does not transport CQ in oocytes
  [PMID:25378409 "mutant PfCRT transports the antimalarial drug"
  chloroquine away from its target, whereas the wild-type form of PfCRT does not].
• Mutant PfCRT is a genuine multidrug carrier
  [PMID:25378409 "mutant PfCRT also transports quinine, quinidine, and verapamil, indicating"
  that the protein behaves as a multidrug resistance carrier], with distinct but
  interacting substrate sites.
• Cryo-EM of the 7G8 isoform shows resistance mutations lining a central negatively
  charged cavity and confirms membrane-potential- and pH-dependent CQ transport
  PMID:31776516.
• Collateral sensitivity work shows CQ-resistant PfCRT isoforms, but not wild type,
  transport lumefantrine and mefloquine
  PMID:35507548.
• The purified-protein study did detect CQ and verapamil uptake by the CQ-sensitive
  protein as well PMID:25733858,
  so xenobiotic transporter activity is defensible for the gene product, but it is an
  acquired/secondary capability, strongly allele-dependent, and not the natural role.

5. Glutathione

• PMID:23256874 —
  note "mutant alleles"; the paper's own model is that GSH influx into the DV is a
  property of the CQR isoforms, contributing to resistance, not the normal function of
  the 3D7 protein.
• Shafik et al. explicitly list "a glutathione transporter" among the incompatible
  earlier claims and screened GSH without finding it to be a substrate or co-substrate
  PMID:32764664.
• The IBA for GO:0034635 glutathione transport is seeded by Arabidopsis CRT-like
  (CLT) plastidic glutathione transporters (AGI_LocusCode:AT4G24460, AT5G12170,
  AT5G19380). The plant CLT clade is a distinct, plastid-targeted branch of the
  CRT-like family; transferring the GSH-transport role onto the apicomplexan DV
  transporter is exactly the kind of family-level over-reach the in-situ data argue
  against. Both GSH rows are therefore marked as over-annotation rather than removed
  (the oocyte IDA is real, just allele-specific and not reproduced).

6. Protist-specific checks performed

• No plant/animal-only terms survive: the only plant-derived inference is the
  glutathione-transport IBA, handled above.
• Organelle terms are apicomplexan-appropriate: food vacuole (GO:0020020) is the
  correct digestive compartment; no apicoplast, rhoptry, microneme or
  parasitophorous-vacuole claims are made.
• No host-vs-symbiont process confusion: PfCRT acts entirely within the parasite's own
  DV membrane; the peptides transported are host-derived but the transport event is
  intra-parasite, so symbiont-host interaction terms are not warranted.
• Paralog check: PfMDR1 (an ABC transporter, separate family) also sits in the DV
  membrane and transports several of the same drugs PMID:35507548; drug-transport
  terms should not be propagated between the two on the basis of shared phenotypes.

7. Curation decisions (summary)

Note on the ER row: only the abstract of PMID:20015114 is cached and it does not
mention the endoplasmic reticulum, so no verbatim quote can be attached. The action is
unchanged — the IDA rests on full-text trafficking data the curator read — but the
supported_by entry was dropped rather than filled with the threonine-416 sorting
quote, which names no compartment.

Proposed new term: food vacuole membrane (the membrane bounding GO:0020020), needed
because the DV membrane is the functional site of PfCRT, PfMDR1, PfV-type ATPase and
several drug-resistance determinants, and GO currently forces the generic
GO:0005774 vacuolar membrane.