Egress from host cells is a crucial step in the life cycle of intracellular pathogens including the parasitic protozoan Toxoplasma gondii. T. gondii can cause severe disease during pregnancy or immune dysfunction, and debilitating ocular pathology in otherwise healthy people. After intracellular replication, the parasite ruptures from host cells in a manner shared by other parasites including Plasmodium. Lytic egress frees the parasite to infect new host cells but the ensuing damage also fuels inflammation and fever, hallmarks of toxoplasmosis and malaria. We recently showed that T. gondii rapid egress and fatal acute disease both depend on the timely secretion of a pore-forming protein, TgPLP1. Our finding that TgPLP1 is released from apical micronemes exposed a novel role for these regulated secretory organelles. How TgPLP1 lyses host membranes during egress is not known. The absence of such knowledge precludes strategic efforts to extinguish its activity and alter the course of infection. Our long-term goal is to understand the roles of key microneme proteins in T. gondii infection. The objective for this funding period is to determine how TgPLP1 selectively and rapidly lyses host cell membranes during egress without causing parasite self-damage. We hypothesize that TgPLP1 pore formation is dictated by the lipid composition of its target membranes, environmental factors, and specific structural features of the protein. This assertion is based on preliminary data identifying host lipid receptors targeted for selective lysis, potential roles for pH and proteolysis in regulating pore formation, and conserved structural features of TgPLP1 predicted to drive membrane binding and oligomerization.
The specific aims are: (1) Reveal the mechanism for selective cytolysis of host but not parasite membranes;(2) Uncover how TgPLP1 is active during egress but not during invasion;and (3) Identify the molecular basis for rapid pore formation. By providing a deeper mechanistic understanding of an essential step in the T. gondii life cycle, we expect the proposed work will create novel opportunities to interrupt infection and potentially ameliorate disease.

Public Health Relevance

We seek to understand how a putative pore forming protein called TgPLP1 facilitates cell egress by the human parasite Toxoplasma gondii. TgPLP1 belongs to a family of pore forming proteins are used for both for microbial aggression and host defense. Our work will reveal mechanistic and regulatory features of pore formation that are relevant to a broad range of microbial infections and immune-related diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI046675-14
Application #
8451408
Study Section
Pathogenic Eukaryotes Study Section (PTHE)
Program Officer
Mcgugan, Glen C
Project Start
1999-12-01
Project End
2016-04-30
Budget Start
2013-05-01
Budget End
2014-04-30
Support Year
14
Fiscal Year
2013
Total Cost
$321,981
Indirect Cost
$110,481
Name
University of Michigan Ann Arbor
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Huynh, My-Hang; Boulanger, Martin J; Carruthers, Vern B (2014) A conserved apicomplexan microneme protein contributes to Toxoplasma gondii invasion and virulence. Infect Immun 82:4358-68
Roiko, Marijo S; Carruthers, Vern B (2013) Functional dissection of Toxoplasma gondii perforin-like protein 1 reveals a dual domain mode of membrane binding for cytolysis and parasite egress. J Biol Chem 288:8712-25
Tomavo, Stanislas; Slomianny, Christian; Meissner, Markus et al. (2013) Protein trafficking through the endosomal system prepares intracellular parasites for a home invasion. PLoS Pathog 9:e1003629
Blackman, Michael J; Carruthers, Vern B (2013) Recent insights into apicomplexan parasite egress provide new views to a kill. Curr Opin Microbiol 16:459-64
Pszenny, Viviana; Davis, Paul H; Zhou, Xing W et al. (2012) Targeted disruption of Toxoplasma gondii serine protease inhibitor 1 increases bradyzoite cyst formation in vitro and parasite tissue burden in mice. Infect Immun 80:1156-65
Gaji, Rajshekhar Y; Behnke, Michael S; Lehmann, Margaret M et al. (2011) Cell cycle-dependent, intercellular transmission of Toxoplasma gondii is accompanied by marked changes in parasite gene expression. Mol Microbiol 79:192-204
Gaji, Rajshekhar Y; Flammer, Halley P; Carruthers, Vern B (2011) Forward targeting of Toxoplasma gondii proproteins to the micronemes involves conserved aliphatic amino acids. Traffic 12:840-53
Laliberte, Julie; Carruthers, Vern B (2011) Toxoplasma gondii toxolysin 4 is an extensively processed putative metalloproteinase secreted from micronemes. Mol Biochem Parasitol 177:49-56
Lagal, Vanessa; Binder, Emily M; Huynh, My-Hang et al. (2010) Toxoplasma gondii protease TgSUB1 is required for cell surface processing of micronemal adhesive complexes and efficient adhesion of tachyzoites. Cell Microbiol 12:1792-808
Beck, Josh R; Rodriguez-Fernandez, Imilce A; de Leon, Jessica Cruz et al. (2010) A novel family of Toxoplasma IMC proteins displays a hierarchical organization and functions in coordinating parasite division. PLoS Pathog 6:e1001094

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