Microbiology
An apically located hybrid guanylate cyclase–ATPase is critical for the initiation of Ca2+ signaling and motility in Toxoplasma gondii

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Protozoan parasites of the phylum Apicomplexa actively move through tissue to initiate and perpetuate infection. The regulation of parasite motility relies on cyclic nucleotide-dependent kinases, but how these kinases are activated remains unknown. Here, using an array of biochemical and cell biology approaches, we show that the apicomplexan parasite Toxoplasma gondii expresses a large guanylate cyclase (TgGC) protein, which contains several upstream ATPase transporter-like domains. We show that TgGC has a dynamic localization, being concentrated at the apical tip in extracellular parasites, which then relocates to a more cytosolic distribution during intracellular replication. Conditional TgGC knockdown revealed that this protein is essential for acute-stage tachyzoite growth, as TgGC-deficient parasites were defective in motility, host cell attachment, invasion, and subsequent host cell egress. We show that TgGC is critical for a rapid rise in cytosolic [Ca2+] and for secretion of microneme organelles upon stimulation with a cGMP agonist, but these deficiencies can be bypassed by direct activation of signaling by a Ca2+ ionophore. Furthermore, we found that TgGC is required for transducing changes in extracellular pH and [K+] to activate cytosolic [Ca2+] flux. Together, the results of our work implicate TgGC as a putative signal transducer that activates Ca2+ signaling and motility in Toxoplasma.

calcium
Toxoplasma gondii
cell motility
cyclic GMP (cGMP)
cyclic nucleotide

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This work was supported by Australian Research Council (ARC) Future Fellowship FT120100164 (to C. J. T.); a China Council scholarship (to L. Y.); Victorian State Government Operational Infrastructure Support and the Australian Government National Health and Medical Research Council (NHMRC) Independent Medical Research Institutes Infrastructure Support Scheme (IRIISS); Agence Nationale de la Recherche, France Grant ANR-12-PDOC-0028-Project Apicolipid, the Atip-Avenir and Finovi programs (CNRS-INSERM-FinoviAtip-Avenir Apicolipid projects), and Laboratoire d’ Excellence Parafrap, France Grant ANR-11-LABX-0024 (to C. Y. B., Y. Y.-B., and N. J. K.); the Laboratoire International Associé (LIA) CNRS Program (Apicolipid project) (to C. Y. B. and C. J. T.); and an NHMRC principle research fellowship (to M. J. M.). The authors declare that they have no conflicts of interest with the contents of this article.

This article contains Fig. S1 and Movies S1–S4.