Faculty

Pathogenesis of parasitic infections: Mechanisms of cell invasion and survival during infection
    

    The Carruthers lab seeks to understand invasion and survival strategies employed by microbial pathogens during infection. We use the protozoan Toxoplasma gondii as a model pathogen because of its genetic and biochemical tractability, well defined cellular structure, and the availability of excellent rodent models of disease. Toxoplasma replicates in a remarkable variety of cells and organs, causing encephalitis, pneumonia, myocarditis, ocular disease, and congenital birth defects during acute infection. While disease is seen only in a small fraction of the ~2 billion people infected worldwide, the diagnosis and treatment of toxoplasmosis are suboptimal and the disease remains a significant public health problem. In people with healthy immune status, the parasite typically remains in a chronic, encysted state, but the infection can erupt when immune function is compromised such as individuals with HIV/AIDS, organ transplant recipients, or cancer patients undergoing chemotherapy.

    As an obligate intracellular pathogen, Toxoplasma critically relies on cell invasion as a major survival strategy to avoid host antibody defense and phagocytic clearance. Cell invasion also initiates the parasite’s lytic cycle that ultimately destroys the infected cell, causing direct tissue pathology and indirect inflammatory damage. We and our collaborators have shown that Toxoplasma uses a battery of adhesive protein complexes to recognize and bind host cells prior to invasion. Many of these adhesive proteins reside in specialized secretory organelles called micronemes (Greek: small threads) that are discharged when the parasite has identified a suitable site for cell invasion. We have shown that parasites conditionally deficient in one particular complex (TgMIC2-M2AP) are invasion incompetent, partially defective in gliding motility, and fail to kill mice even at high doses. The deficient strain also acts as a live-attenuated vaccine since surviving animals are protected from subsequent challenge with a lethal parasite strain. Our team is actively investigating how adhesive complexes such as TgMIC2-M2AP are assembled and shuttled to invasion organelles, in addition to identifying novel invasion-related proteins that may contribute to the parasite’s broad host range.

    From another prospective, we also seek to understand what roles the host cell plays in cell invasion by Toxoplasma. Although the motive force for cell invasion is clearly parasite-derived, we propose that a number of host molecules are recruited and manipulated by the parasite to rapidly traverse the host actin cytoskeletal system and moments later pinch off the host plasma membrane to create the intracellular compartment in which replication ensues. Indeed, an astonishing degree of molecular coordination must be employed during the 20 seconds or so the parasite takes to invade a target cell. Our goal is to understand the orchestration of parasite and host proteins involved in this uncommonly rapid entry phenomenon.

    The team has also identified parasite proteases that act upon adhesive complexes both within the parasite in preparation for cell invasion and on the parasite surface during parasite entry. In addition to facilitating invasion by proteolytically modifying adhesins, our findings imply that one such protease (TgCPL) may also function during intracellular replication since it occupies a novel and dynamic digestive compartment that is likely part of the parasite’s endocytic system. We are using selective inhibitors of these proteases to determine substrate range, their specific role in invasion and replication, and to test the efficacy of inhibitory compounds for ameliorating infection.

    To navigate the above research avenues, we use a diverse array of approaches including molecular genetics, proteomics, bioinformatics, video and fluorescence microscopy, structural biology, mathematical modeling, and bioluminescent imaging.

Selected Publications: 

    Brydges S. D., Harper, J. M., Parussini, F. Coppens, I., Carruthers V.B. A transient forward targeting element for microneme regulated secretion in Toxoplasma gondii. Biol. Cell (submitted for publication)

Kafsack, B.F.C, Carruthers, V.B., Pineda F. (2007) A mathematical model for cell invasion by Toxoplasma gondii. J. Theoretical Biol. (in press).

Teo, C.F., Zhou, X.W., Bogyo, M., Carruthers, V.B., (2007) Cysteine protease inhibitors block Toxoplasma microneme secretion and cell invasion. Antimicrob. Agents Chemotherapy 51;679-688.

Brydges S. D., Zhou X. W., Carruthers V.B. (2006) Genetic ablation of TgMIC5 enhances proteolytic processing on the surface of Toxoplasma gondii tachyzoites. Euk. Cell 5;2174-2183. [Cover illustration]

Starnes, G.L., Travis J. Jewett, T.J., Carruthers, V.B., Sibley, L.D. (2006) Two separate, conserved acidic amino acid domains within the Toxoplasma gondii MIC2 cytoplasmic tail are required for parasite survival. J. Biol. Chem. 281;30745-30754.

Harper, J., Huynh, M.-H., Coppens, I., Parussini, F., Moreno, S., Carruthers, V.B. (2006) A cleavable propeptide influences Toxoplasma infection by facilitating the trafficking and secretion of the TgMIC2-M2AP invasion complex. Mol. Biol. Cell. 17;4551-4563.

Carruthers, V.B., Boothroyd, J.C. (2006) Pulling together: An integrated model of Toxoplasma cell invasion. Curr. Opin. Microbiol. 9:1-7.

Huynh, M.-H., Carruthers, V.B. (2006) Toxoplasma MIC2 is a major determinant of invasion and virulence. PLoS Pathogens Aug 18;2(8).

Carruthers, V.B., Proteolysis and Toxoplasma invasion. (2006) Int. J. Parasitol. 36;494-500.

Huynh, M.-H., Harper, J.M., Carruthers, V.B. (2006) Preparing for an invasion: Charting the pathway of adhesion proteins to Toxoplasma micronemes. Parasite Res. 98;389-395.

Carruthers, V.B., Blackman, M.J. (2005) A new release on life: Emerging concepts in proteolysis and parasite invasion. Mol. Microbiol..55; 1617-1630. [Cover Illustration].

Zhou, X.W., Kafsack, B.F., Cole, R.N., Beckett, P., Shen, R.F., Carruthers, V.B. (2005) The opportunistic pathogen Toxoplasma gondiideploys a diverse legion of invasion and survival proteins. J. Biol. Chem. 280; 34233-34244.

Howell, S.A., Hackett, F., Jongco, A.M., Withers-Martinez, C., Kim, K., Carruthers, V.B., Blackman, M.J. (2005) Distinct mechanisms govern proteolytic shedding of a key invasion protein in apicomplexan pathogens. Mol. Microbiol. 57; 1342-1356.

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