GAP40

GAP40 is a polytopic integral membrane protein of the Toxoplasma gondii inner membrane complex (IMC) that functions as a structural anchor for the glideosome, the actomyosin motor complex driving parasite gliding motility (PMID:20951968). It anchors the MyoA-MLC1 motor together with the integral membrane protein GAP50 (PMID:20951968), and this GAP40-GAP50 interaction depends on N-glycosylation of GAP50 (PMID:21610105). GAP40 is shared between distinct glideosomes, associating with both the MyoA-containing motor complex and the class XIV MyoC-containing glideosome at the basal polar ring (PMID:25393004). Beyond its scaffolding role in motility, GAP40 is essential for IMC biogenesis during intracellular replication: its loss causes rapid collapse of developing parasites and redistribution of other glideosome components (PMID:26845335).

1. 2010 High

   Establishing what physically holds the gliding motor to the IMC, GAP40 was defined as a polytopic membrane anchor for the MyoA-MLC1 motor complex, identifying a structural foundation for glideosome assembly.

   Evidence Biochemical fractionation and reciprocal co-immunoprecipitation of glideosome components

   PMID:20951968

   Open questions at the time

   • Membrane topology not resolved at residue level
   • Direct binding interface with GAP50 not mapped
2. 2011 Medium

   To determine how GAP40 is recruited into the motor scaffold, the GAP40-GAP50 interaction was shown to require N-glycosylation of GAP50, linking a post-translational modification to glideosome partner assembly.

   Evidence Comparative affinity purification with glycosylated vs. unglycosylated GAP50 bait and N-glycosylation-site mutagenesis

   PMID:21610105

   Open questions at the time

   • Whether GAP40 itself is glycosylated not addressed
   • Single lab, no reciprocal structural validation
3. 2014 Medium

   Addressing whether GAP40 serves only the MyoA motor, it was found to also associate with the MyoC-containing glideosome at the basal polar ring, showing GAP40 is a shared anchor across distinct motor complexes.

   Evidence Co-immunoprecipitation and localization studies of MyoC-glideosome components

   PMID:25393004

   Open questions at the time

   • Whether GAP40 simultaneously binds both motors or in spatially separate pools unclear
   • Single lab
4. 2014 Medium

   To test which components' modifications are required for assembly, mutagenesis showed GAP40 anchors the MyoA motor via GAP50 independently of GAP45/MLC1 phosphorylation, delimiting the regulatory inputs to assembly.

   Evidence Phospho-mimetic/phospho-null mutagenesis with complementation in inducible knockout background

   PMID:24779470

   Open questions at the time

   • Direct role of GAP40 phosphorylation not tested
   • Single lab
5. 2016 High

   Moving from binding partner to cellular requirement, conditional loss of GAP40 revealed it is essential for IMC biogenesis, establishing a structural role in daughter-cell formation beyond motor anchoring.

   Evidence Conditional deletion/disruption of GAP40 with readout of IMC formation, parasite replication, and glideosome marker redistribution

   PMID:26845335

   Open questions at the time

   • Molecular mechanism linking GAP40 to IMC membrane biogenesis unresolved
   • Whether IMC collapse is direct or secondary to loss of glideosome integrity not distinguished

• How GAP40's polytopic membrane architecture mechanistically couples IMC biogenesis to motor anchoring, and whether the two roles are separable, remains open.
• No structural model of GAP40 or its membrane topology
• No mapped interaction interfaces with GAP50 or myosin partners
• Mechanism of IMC membrane collapse upon GAP40 loss uncharacterized