| 2004 |
TgGAP45 was identified as a component of the myosin XIV motor complex in Toxoplasma gondii, together with TgMyoA, TgMLC1, and TgGAP50. The complex is localized to the inner membrane complex (IMC), where it is anchored by TgGAP50, an integral membrane glycoprotein. Assembly occurs in two spatially controlled stages. |
Co-immunoprecipitation, subcellular fractionation, immunofluorescence microscopy |
The Journal of cell biology |
High |
15123738
|
| 2007 |
The myosin XIV motor complex (MyoA/MLC1/GAP45/GAP50) is firmly immobilized in the plane of the IMC. This immobilization requires cholesterol and is associated with detergent-resistant membrane (DRM) domains, but does not appear to depend on direct interactions with cytoskeletal elements. |
Fluorescence recovery after photobleaching (FRAP), detergent-resistant membrane fractionation, cholesterol depletion |
Molecular biology of the cell |
Medium |
17538016
|
| 2008 |
GAP45 is phosphorylated on multiple residues including Ser163 and Ser167. Phospho-mimetic mutation of both residues to Glu prevents association of the MyoA-MLC1-GAP45 soluble complex with the membrane anchor GAP50, demonstrating that phosphorylation of these sites controls the final step in myosin XIV motor complex assembly. Phospho-null mutations (Ala) do not block this association. |
Mass spectrometry phosphosite mapping, site-directed mutagenesis (Ser→Glu / Ser→Ala), co-immunoprecipitation |
Eukaryotic cell |
High |
19047362
|
| 2010 |
GAP45 is anchored to both the plasma membrane (via N-terminal acylation) and the IMC (via its C-terminal extremity). The C-terminal region of GAP45 recruits MLC1-MyoA to the IMC, while the N-terminal acylation and coiled-coil domain preserve pellicle integrity during invasion. GAP45 is essential for gliding, invasion, and egress. Plasmodium falciparum GAP45 can functionally replace TgGAP45, demonstrated by transgenera complementation. |
Conditional/inducible knockout, domain truncation/mutagenesis, transgenera complementation, immunofluorescence localization, gliding/invasion/egress assays |
Cell host & microbe |
High |
20951968
|
| 2010 |
The in vitro motility activity of the TgMyoA motor complex (containing TgMLC1, TgGAP45, and TgGAP50) is significantly reduced when TgMLC1 carries a compound-induced post-translational modification, providing evidence that the mechanical activity of the Class XIV myosin complex can be modulated by modifications to its associated light chain. |
In vitro motility assay comparing complexes from control vs. compound-treated parasites, electrophoretic mobility shift to detect TgMLC1 modification |
PLoS pathogens |
Medium |
20084115
|
| 2011 |
Calcium-dependent phosphorylation of six residues across GAP45, MLC1, and MyoA is correlated with invasion motor activity, as determined by SILAC-based quantitative LC-MS/MS analysis of Ca2+-stimulated tachyzoites. |
SILAC quantitative LC-MS/MS, 32P-labeling, 2-DE, Ca2+-mobilizing drug stimulation |
PLoS pathogens |
Medium |
21980283
|
| 2014 |
GAP45 is the acylated glideosome component that recruits the MyoA motor complex to the IMC pellicle and contributes to pellicle cohesion during motor traction. Conditional deletion of GAP45 reduces but does not fully abolish gliding or invasion, indicating that other mechanisms (e.g., MyoC-glideosome) can compensate. Loss of MyoA leads to relocalization of MyoC along the pellicle and apical cap, accounting for residual invasion. |
Inducible knockout (tetracycline-regulatable), gliding/invasion/egress assays, immunofluorescence |
PLoS pathogens |
High |
24632839 25393004
|
| 2014 |
Phospho-mimetic and phospho-null mutations of the twelve-residue phosphorylation cluster in TgGAP45, and of phosphorylation sites in TgMLC1, do not impair glideosome assembly or parasite motility when tested in a complementation strategy using an inducible GAP45 knockout background. The previously reported loss-of-interaction phenotype of a phospho-mimetic GAP45 mutant (Ser163/167Glu) with GAP50 was an artifact caused by competition with a co-existing wild-type copy. Thus, GAP45 phosphorylation is not critical for glideosome assembly or function. |
Allelic replacement, phospho-mimetic/phospho-null mutagenesis, inducible knockout complementation, co-immunoprecipitation, motility assays |
Cellular microbiology |
Medium |
24779470
|
| 2016 |
TgCaMKrk (a calmodulin-independent CaMK homolog localizing to the apical end of tachyzoites) phosphorylates TgGAP45 in vitro, identifying it as a candidate kinase for GAP45 phosphorylation. |
In vitro kinase assay with purified proteins |
Parasites & vectors |
Low |
27444499
|
| 2021 |
TgMLC1 palmitoylation on Cys8 and Cys11 is required for its binding to TgGAP45. Blocking palmitoylation completely abrogates TgMLC1–TgGAP45 interaction. Surprisingly, loss of this interaction has little effect on parasite motility, challenging the necessity of MLC1-GAP45 binding for force transduction. |
Site-directed mutagenesis of palmitoylation sites, co-immunoprecipitation, gliding motility assays |
mSphere |
Medium |
34011689
|