PIKfyve

In Toxoplasma gondii, PIKfyve is a lipid kinase that phosphorylates PI(3)P to generate PI(3,5)P2, an activity required for apicoplast homeostasis, since conditional disruption of PIKfyve enlarges the apicoplast (PMID:25329540). PIKfyve functions together with its regulatory protein ArPIKfyve, whose conditional knockdown phenocopies the enlarged-apicoplast defect, establishing that the PIKfyve–ArPIKfyve complex maintains this organelle (PMID:25329540). The PI(3)P pool at the apicoplast appears to act principally as a precursor for PI(3,5)P2 synthesis rather than through PX/FYVE-domain effectors, as depletion of six candidate PI(3)P-binding proteins did not impair growth or apicoplast biology (PMID:25329540). Beyond this apicoplast role and the inferred kinase reaction, no further mechanistic detail has been characterized in the available corpus.

1. 2014 Medium

   Established that PIKfyve provides the PI(3,5)P2-generating activity needed for apicoplast homeostasis, defining a specific lipid-signaling requirement for maintaining this organelle.

   Evidence Conditional gene disruption of PIKfyve with apicoplast morphology readout in T. gondii, with kinase activity inferred from substrate identity (PI(3)P → PI(3,5)P2)

   PMID:25329540

   Open questions at the time

   • Kinase activity inferred from substrate identity rather than directly reconstituted in vitro
   • Mechanism by which PI(3,5)P2 controls apicoplast size not resolved
   • Subcellular localization of PIKfyve relative to the apicoplast not defined in the corpus
2. 2014 Medium

   Showed that PIKfyve acts within a complex with its regulatory partner ArPIKfyve, since knockdown of ArPIKfyve reproduces the apicoplast enlargement phenotype.

   Evidence Conditional knockdown of ArPIKfyve with apicoplast morphology readout in T. gondii

   PMID:25329540

   Open questions at the time

   • Physical association of PIKfyve and ArPIKfyve not directly demonstrated by biochemical interaction data in the corpus
   • How ArPIKfyve regulates PIKfyve kinase output is unknown
3. 2014 Medium

   Distinguished the biological function of PI(3)P at the apicoplast, indicating it serves mainly as a precursor for PI(3,5)P2 rather than acting through PX/FYVE-domain effectors.

   Evidence Conditional depletion panel of six PX/FYVE-domain candidate PI(3)P effectors with growth and apicoplast phenotype readout (negative result) in T. gondii

   PMID:25329540

   Open questions at the time

   • Negative essentiality result does not exclude redundancy among the candidate effectors
   • Direct measurement of apicoplast PI(3)P versus PI(3,5)P2 pools not reported

• How PI(3,5)P2 mechanistically maintains apicoplast size and inheritance, and the localization and direct enzymatic characterization of PIKfyve, remain unresolved.
• No downstream effector of PI(3,5)P2 at the apicoplast identified
• No in vitro reconstitution of the PIKfyve kinase reaction in the corpus
• No structural or interaction data on the PIKfyve–ArPIKfyve complex