A Conserved Apicomplexan Microneme Protein Contributes to Toxoplasma gondii Invasion and Virulence

Abstract

The obligate intracellular parasite Toxoplasma gondii critically relies on host cell invasion during infection. Proteins secreted from the apical micronemes are central components for host cell recognition, invasion, egress, and virulence. Although previous work established that the sporozoite protein with an altered thrombospondin repeat (SPATR) is a micronemal protein conserved in other apicomplexan parasites, including Plasmodium, Neospora, and Eimeria, no genetic evidence of its contribution to invasion has been reported. SPATR contains a predicted epidermal growth factor domain and two thrombospondin type 1 repeats, implying a role in host cell recognition. In this study, we assess the contribution of T. gondii SPATR (TgSPATR) to T. gondii invasion by genetically ablating it and restoring its expression by genetic complementation. ⌬spatr parasites were ϳ50% reduced in invasion compared to parental strains, a defect that was reversed in the complemented strain. In mouse virulence assays, ⌬spatr parasites were significantly attenuated, with ϳ20% of mice surviving infection. Given the conservation of this protein among the Apicomplexa, we assessed whether the Plasmodium falciparum SPATR ortholog (PfSPATR) could complement the absence of the TgSPATR. Although PfSPATR showed correct micronemal localization, it did not reverse the invasion deficiency of ⌬spatr parasites, because of an apparent failure in secretion. Overall, the results suggest that TgSPATR contributes to invasion and virulence, findings that have implications for the many genera and life stages of apicomplexans that express SPATR.

The Rhoptry Pseudokinase ROP54 Modulates Toxoplasma gondii Virulence and Host GBP2 Loading

“…The PCR product contained the endogenous promoter, ROP54 gene, 3×HA tag, and the HXGPRT 3′-untranslated region from the tagging construct. The amplicon was ligated into a complementation vector with the 3′ and 5′ flanks of the deleted Ku80 locus and selectable marker HXGPRT (provided by Vern Carruthers, University of Michigan) ( 53 ). The plasmid was linearized with BssHII, transfected into the Δrop54 II clone, and selected with MX medium.…”

Intersection of endocytic and exocytic systems in Toxoplasma gondii

“…The presence of a plant‐like lysosome and a plant‐specific proton pump (TgVP1) within the T. gondii endolysosomal system suggests that endocytic trafficking in T. gondii may resemble plants as proposed by Pieperhoff et al However, exocytic trafficking of proteins destined for the parasite’s regulated secretory organelles, the micronemes and rhoptries, proceeds through the TGN and ELCs, and requires clathrin, dynamin and Rab5 for transit . In contrast to the ingestion pathway, which leads to the destruction of its cargo, most microneme and rhoptry proteins have propeptides that are cleaved off during transit to the microneme and rhoptry organelles, but must otherwise remain intact to orchestrate parasite invasion, egress and defense against host immune attack . Without these exocytic proteins and organelles, the parasite cannot establish a successful infection.…”

“…[9][10][11][12][13][14][15][16] In contrast to the ingestion pathway, which leads to the destruction of its cargo, most microneme and rhoptry proteins have propeptides that are cleaved off during transit to the microneme and rhoptry organelles, but must otherwise remain intact to orchestrate parasite invasion, egress and defense against host immune attack. [17][18][19][20][21][22][23][24][25][26][27] Without these exocytic proteins and organelles, the parasite cannot establish a successful infection.…”

Secreted protein with altered thrombospondin repeat (SPATR) is essential for asexual blood stages but not required for hepatocyte invasion by the malaria parasite Plasmodium berghei

“…High‐activity binding peptides (HABPs) from P. falciparum SPATR bound to RBCs rather than hepatocytes and inhibited in vitro invasion of merozoites (Curtidor et al, ). The role of SPATR was also determined in invasion of host cells by T. gondii using genetic ablation (Huynh, Boulanger, & Carruthers, ). These findings suggest the importance of SPATR in invasion but no genetic evidence has been reported in Plasmodium .…”