Quantitative proteomic measurement of the Ca2+-dependent excretory-secretory antigen secretome of Toxoplasma gondii

ABSTRACT: Apicomplexan parasites discharge specialized organelles called rhoptries upon host cell contact to mediate invasion. The events that drive rhoptry discharge are poorly understood, yet essential to sustain the apicomplexan parasitic life cycle. Rhoptry discharge appears to depend on proteins secreted from another set of organelles called micronemes, which in Toxoplasma gondii includes MIC8 and the microneme-associated CRMP complex. Here, we examine the function of the microneme protein CLAMP, uncovering its essential role in rhoptry discharge. CLAMP forms a distinct complex with two other microneme proteins, the invasion-associated SPATR, and a previously uncharacterized protein we name CLAMP-linked invasion protein (CLIP). Using a quantitative mass spectrometry based approach to determine the calcium-dependent excretory-secretory antigen secretome (MS-ESA), we identify secretion of both known and previously unverified microneme proteins. Our MS-ESA assay demonstrates that depletion of CLAMP or CLIP limits secretion of other CLAMP complex members, while not affecting secretion of other microneme proteins. CLAMP-deficiency does not impact parasite adhesion, however, knockdown of any member of the CLAMP complex affects rhoptry discharge. Phylogenetic analysis suggests orthologs of the essential complex components, CLAMP and CLIP, are ubiquitous across apicomplexans. Nevertheless, SPATR, which appears to act as an accessory factor in Toxoplasma, is essential for invasion during Plasmodium falciparum blood stages. Our results reveal a new protein complex that mediates rhoptry discharge following host-cell contact.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Toxoplasma Gondii Gt1

SUBMITTER: Dylan Valleau  

LAB HEAD: Sebastian Lourido

PROVIDER: PXD044182 | Pride | 2023-10-30

REPOSITORIES: Pride