The obligate intracellular protozoan Toxoplasma gondii is a leading cause of posterior uveitis in the general population, encephalitis in immunodeficient individuals, and cognitive impairment in congenitally infected children. We seek to understand how Toxoplasma prepares for and achieves cell invasion. Although it has been well recognized that many Toxoplasma invasion proteins undergo proteolytic maturation en route to apical secretory organelles, little is known about the proteases involved or where and why maturation occurs. We hypothesize that T. gondii uses a cathepsin proteases to proteolytically activate and stabilize invasion proteins within a novel multivesicular endosome (MVE), based on preliminary immunolocalization, gene knockout, and inhibitor studies, along with protease specificity analysis and in vitro processing of a putative substrate. To test this hypothesis we will: (1) Characterize defects and compensatory mechanisms in CPL knockout parasites;(2) Confirm the overlapping roles of cathepsins in proMIC maturation by genetic and inhibitor studies;and (3) Test the "lock and load" hypothesis that proteolytic maturation stabilizes proMIC protein complexes. This work will illuminate basic mechanisms of secretory protein trafficking and proteolytic modulation of protein function that may be shared with related apicomplexan parasites and other eukaryotic cells that utilize regulated secretion.

Public Health Relevance

Our goal is to find new ways of interfering with Toxoplasma invasion of human cells as a strategy of controlling infections caused by this parasite. Toxoplasma proteases may be good targets for new anti-parasitic drugs;this work is designed to test the function and significance of two such proteases.

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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Rogers, Martin J
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University of Michigan Ann Arbor
Schools of Medicine
Ann Arbor
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Dou, Zhicheng; McGovern, Olivia L; Di Cristina, Manlio et al. (2014) Toxoplasma gondii ingests and digests host cytosolic proteins. MBio 5:e01188-14
Liu, Jing; Pace, Douglas; Dou, Zhicheng et al. (2014) A vacuolar-H(+) -pyrophosphatase (TgVP1) is required for microneme secretion, host cell invasion, and extracellular survival of Toxoplasma gondii. Mol Microbiol 93:698-712
Warring, Sally D; Dou, Zhicheng; Carruthers, Vern B et al. (2014) Characterization of the chloroquine resistance transporter homologue in Toxoplasma gondii. Eukaryot Cell 13:1360-70
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
Dou, Zhicheng; Coppens, Isabelle; Carruthers, Vern B (2013) Non-canonical maturation of two papain-family proteases in Toxoplasma gondii. J Biol Chem 288:3523-34
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
Gaji, Rajshekhar Y; Huynh, My-Hang; Carruthers, Vern B (2013) A novel high throughput invasion screen identifies host actin regulators required for efficient cell entry by Toxoplasma gondii. PLoS One 8:e64693
Dou, Zhicheng; Carruthers, Vern B (2011) Cathepsin proteases in Toxoplasma gondii. Adv Exp Med Biol 712:49-61
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
Parussini, Fabiola; Coppens, Isabelle; Shah, Parag P et al. (2010) Cathepsin L occupies a vacuolar compartment and is a protein maturase within the endo/exocytic system of Toxoplasma gondii. Mol Microbiol 76:1340-57

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