Proc Natl Acad Sci U S A. 2012
Apr 20. [Epub ahead of print]
Intramembrane proteolysis of Toxoplasma apical membrane antigen 1 facilitates host-cell invasion but is dispensable for replication.
SourceDepartment of Microbiology and Molecular Genetics, University of Vermont,
Burlington, VT 05405.
Abstract
Apical membrane antigen 1 (AMA1) is a conserved transmembrane adhesin of
apicomplexan parasites that plays an important role in host-cell invasion.
Toxoplasma gondii AMA1 (TgAMA1) is secreted onto the parasite surface and
subsequently released by proteolytic cleavage within its transmembrane domain.
To elucidate the function of TgAMA1 intramembrane proteolysis, we used a
heterologous cleavage assay to characterize the determinants within the TgAMA1
transmembrane domain (ALIAGLAVGGVLLLALLGGGCYFA) that govern its processing.
Quantitative analysis revealed that the TgAMA1(L/G) mutation enhanced cleavage
by 13-fold compared with wild type. In contrast, the TgAMA1(AG/FF) mutation
reduced cleavage by 30-fold, whereas the TgAMA1(GG/FF) mutation had a minor
effect on proteolysis; mutating both motifs in a quadruple mutant blocked
cleavage completely. We then complemented a TgAMA1 conditional knockout parasite
line with plasmids expressing these TgAMA1 variants. Contrary to expectation,
variants that increased or decreased TgAMA1 processing by >10-fold had no
phenotypic consequences, revealing that the levels of rhomboid proteolysis in
parasites are not delicately balanced. Only parasites transgenically expressing
or carrying a true knock-in allele of the uncleavable TgAMA1(AG/FF+GG/FF) mutant
showed a growth defect, which resulted from inhibiting invasion without
perturbing intracellular replication. These data demonstrate that TgAMA1
cleavage plays a role in invasion, but refute a recently proposed model in which
parasite replication within the host cell is regulated by intramembrane
proteolysis of TgAMA1.
- PMID: 22523242
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