IMC43

IMC43 is an essential daughter inner membrane complex (IMC) protein in Toxoplasma gondii that is recruited at the earliest stages of daughter bud initiation and scaffolds nascent daughter IMC assembly during endodyogeny (PMID:37782662). Conditional depletion produces aberrant morphology, dysregulated endodyogeny, and severe replication defects, establishing IMC43 as required for parasite fitness (PMID:37782662). IMC43 acts through its essential C-terminal domain, which directly binds IMC32 and IMC44 and is required to regulate the stage-specific localization of both partners (PMID:37782662). Together IMC43 and IMC32 form an essential daughter bud assembly complex that founds the daughter IMC scaffold (PMID:37782662, PMID:39024411), and IMC43 is additionally required for recruitment of BCC0 to nascent buds (PMID:39024411). BCC0 does not bind IMC43 directly but associates with the coiled-coil region of IMC32; an IMC32–BCC0 subcomplex is first recruited to buds via palmitoylation of IMC32 and is subsequently locked into the scaffold when IMC32 engages IMC43 during bud elongation (PMID:39024411).

1. 2023 High

   Identifying when and where a new IMC component appears during replication established IMC43 as an early marker of daughter bud formation, defining the temporal window it acts in.

   Evidence fluorescence microscopy localization across endodyogeny in Toxoplasma gondii

   PMID:37782662

   Open questions at the time

   • localization alone does not establish function
   • recruitment signal directing IMC43 to nascent buds not defined
2. 2023 High

   Conditional depletion tested whether IMC43 is dispensable and showed it is essential, linking loss of the protein directly to failed daughter cell assembly and replication.

   Evidence auxin-inducible degron depletion with morphological and replication readouts

   PMID:37782662

   Open questions at the time

   • does not resolve which molecular interactions underlie the phenotype
   • timing of the block within endodyogeny not fully dissected
3. 2023 Medium

   Domain mapping determined which part of IMC43 carries its function and localization determinants, pinpointing the C-terminal domain as the essential functional module.

   Evidence deletion analysis with localization and functional complementation

   PMID:37782662

   Open questions at the time

   • no structural model of the C-terminal domain
   • residue-level binding determinants not defined
4. 2023 High

   Interaction mapping asked what IMC43 binds and showed its C-terminal domain directly engages IMC32 and IMC44, providing the molecular basis for its scaffolding role.

   Evidence TurboID proximity labeling and pairwise yeast two-hybrid with IMC43 as bait

   PMID:37782662

   Open questions at the time

   • binding affinities and stoichiometry unknown
   • no reconstitution of the complex in vitro
5. 2023 High

   Depleting IMC43 and tracking its partners showed it governs the stage-specific localization of IMC32 and IMC44, elevating IMC43 from a binding partner to an organizing scaffold for the daughter IMC.

   Evidence AID depletion of IMC43 followed by fluorescence microscopy of IMC32 and IMC44

   PMID:37782662

   Open questions at the time

   • mechanism by which IMC43 positions partners not resolved
   • does not define the assembly order relative to other IMC components
6. 2024 High

   Extending the depletion approach to BCC0 established IMC43 (with IMC32) as required for BCC0 recruitment, expanding the assembly complex beyond the IMC43–IMC32 core.

   Evidence conditional depletion of IMC32 and IMC43 with fluorescence microscopy of BCC0

   PMID:39024411

   Open questions at the time

   • does not distinguish direct versus indirect recruitment routes on its own
7. 2024 Medium

   Dissecting the wiring of the tripartite complex resolved the recruitment order, showing BCC0 binds IMC32 (not IMC43) and that a palmitoylation-primed IMC32–BCC0 subcomplex is subsequently locked into the scaffold via IMC32–IMC43 binding during bud elongation.

   Evidence pairwise yeast two-hybrid, proximity labeling, and deletion/complementation analyses

   PMID:39024411

   Open questions at the time

   • palmitoylation enzyme(s) for IMC32 not identified
   • temporal transition from initial recruitment to lock-in not directly imaged
   • single lab, no in vitro reconstitution

• The structural basis of IMC43 scaffolding and how its assembly is regulated in space and time during bud elongation remain open.
• no atomic structure of IMC43 or its complexes
• functional role of IMC44 beyond binding not established
• regulatory signals timing the IMC32–IMC43 lock-in unknown