Exit from host cells is an essential step in the life cycle of intracellular pathogens including Toxoplasma gondii, the agent of human toxoplasmosis. Related malaria parasites also egress from host cells after replication, and they additionally traverse cells by escaping from a transient vacuole in a manner similar to egress. Recent work has identified critical roles for pore-forming proteins and phospholipases in cell egress and traversal. But how these proteins facilitate egress and the extent to which they collaborate to disrupt membranes is unknown. Also, although recent reports suggest that acidification of the parasite-containing vacuole augments activity of the pore-forming protein(s), the proton pumps mediating acidification have not been identified. Precisely how acidification facilitates pore-formation and egress is also unclear. The absence of such knowledge precludes rationale design of interference strategies to alter the course of infection and disease. We will address these key gaps by identifying the proton pump(s) responsible for vacuole acidification (Aim1), defining the structural basis for pH- augmented membrane binding by the pore-forming protein Perforin-Like Protein 1 (Aim 2), and measuring pH-dependent cooperativity between Perforin-Like Protein 1 and an LCAT phospholipase for membrane disruption during egress. The proposed studies will advance the field by delivering insight that is broadly relevant to egress by apicomplexan parasites and other cytolytic pathogens.

Public Health Relevance

These studies will identify how acidifying pumps and membrane disruptive proteins promote cell exit by the human intracellular parasite Toxoplasma gondii. Our work will discovery how membrane disrupting proteins released by T. gondii are regulated and define how function together in a manner that is relevant to a broad range of microbial infections and immune-related diseases.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Pathogenic Eukaryotes Study Section (PTHE)
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Mcgugan, Glen C
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University of Michigan Ann Arbor
Schools of Medicine
Ann Arbor
United States
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Di Cristina, Manlio; Carruthers, Vern B (2018) New and emerging uses of CRISPR/Cas9 to genetically manipulate apicomplexan parasites. Parasitology 145:1119-1126
Schultz, Aric J; Carruthers, Vern B (2018) Toxoplasma gondii LCAT Primarily Contributes to Tachyzoite Egress. mSphere 3:
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Huynh, My-Hang; Carruthers, Vern B (2016) A Toxoplasma gondii Ortholog of Plasmodium GAMA Contributes to Parasite Attachment and Cell Invasion. mSphere 1:
Pszenny, Viviana; Ehrenman, Karen; Romano, Julia D et al. (2016) A Lipolytic Lecithin:Cholesterol Acyltransferase Secreted by Toxoplasma Facilitates Parasite Replication and Egress. J Biol Chem 291:3725-46
Huynh, My-Hang; Liu, Bing; Henry, Maud et al. (2015) Structural basis of Toxoplasma gondii MIC2-associated protein interaction with MIC2. J Biol Chem 290:1432-41
Carruthers, Vern B (2015) Parasites and their heterophagic appetite for disease. PLoS Pathog 11:e1004803
Roiko, Marijo S; Svezhova, Nadezhda; Carruthers, Vern B (2014) Acidification Activates Toxoplasma gondii Motility and Egress by Enhancing Protein Secretion and Cytolytic Activity. PLoS Pathog 10:e1004488

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