(provide by the applicant): Toxoplasma gondii is a widespread Apicomplexan parasite that causes devastating disease in immunocompromised persons and the congenitally infected fetus. The pathology of toxoplasmosis is due to repeated cycles of host cell invasion and lysis by the actively dividing form of T. gondii, the tachyzoite. Despite the importance of invasion to the life cycle of the parasite and the pathology of toxoplasmosis, little is known about the tachyzoite proteins that mediate invasion. Because T. gondii is a haploid, obligate intracellular parasite, forward and reverse genetic approaches to studying invasion are problematic. Small molecules will be used as a means to circumvent this difficulty and identify gene products that play an important role in invasion. A collection of over 14000 structurally diverse small molecules has been screened in a high-throughput invasion assay, and 28 invasion inhibitors have been identified. Unexpectedly, 7 invasion enhancers were also discovered. In secondary assays, several of the bioactive small molecules were shown to affect motility of the parasite and/or secretion from its apical organelles. The goals of the proposal are to: (I) Test the invasion inhibitors and enhancers against several other related and unrelated parasites, to determine whether they target conserved components of the Apicomplexan invasion machinery; (II) Test whether any of the motility inhibitors/enhancers exert their effects through parasite myosin A or the multi-protein complex to which it belongs; (III) Test whether an inhibitor of cGMP phosphodiesterase identified in the screen affects parasite cGMP levels during invasion; (IV) Screen a comprehensive collection of >500 small molecules of known biological function for an effect on invasion; and (V) Use synthetic, biochemical and genetic methods to determine the in vivo targets of 3-5 of the highest priority invasion inhibitors/enhancers, where prioritization is based on the results of I-IV. Target identification will be greatly facilitated by the recently released sequence of the Toxoplasma genome. A strength of the proposal is the level to which biological experiments and synthetic chemistry will be integrated to address the project's goals. This work has the potential to provide important new insights into the proteins and pathways involved in host cell invasion by T. gondii and related parasites, and it may simultaneously identify lead compounds for the development of new and urgently needed anti-parasitic drugs.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI054961-03
Application #
6857131
Study Section
Tropical Medicine and Parasitology Study Section (TMP)
Program Officer
Coyne, Philip Edward
Project Start
2003-03-01
Project End
2008-02-29
Budget Start
2005-03-01
Budget End
2006-02-28
Support Year
3
Fiscal Year
2005
Total Cost
$560,724
Indirect Cost
Name
University of Vermont & St Agric College
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
066811191
City
Burlington
State
VT
Country
United States
Zip Code
05405
Powell, Cameron J; Ramaswamy, Raghavendran; Kelsen, Anne et al. (2018) Structural and mechanistic insights into the function of the unconventional class XIV myosin MyoA from Toxoplasma gondii. Proc Natl Acad Sci U S A 115:E10548-E10555
Whitelaw, Jamie A; Latorre-Barragan, Fernanda; Gras, Simon et al. (2017) Surface attachment, promoted by the actomyosin system of Toxoplasma gondii is important for efficient gliding motility and invasion. BMC Biol 15:1
Gras, Simon; Jackson, Allison; Woods, Stuart et al. (2017) Parasites lacking the micronemal protein MIC2 are deficient in surface attachment and host cell egress, but remain virulent in vivo. Wellcome Open Res 2:32
Powell, Cameron J; Jenkins, Meredith L; Parker, Michelle L et al. (2017) Dissecting the molecular assembly of the Toxoplasma gondii MyoA motility complex. J Biol Chem 292:19469-19477
Odell, Anahi V; Tran, Fanny; Foderaro, Jenna E et al. (2015) Yeast three-hybrid screen identifies TgBRADIN/GRA24 as a negative regulator of Toxoplasma gondii bradyzoite differentiation. PLoS One 10:e0120331
Leung, Jacqueline M; Rould, Mark A; Konradt, Christoph et al. (2014) Disruption of TgPHIL1 alters specific parameters of Toxoplasma gondii motility measured in a quantitative, three-dimensional live motility assay. PLoS One 9:e85763
Leung, Jacqueline M; Tran, Fanny; Pathak, Ravindra B et al. (2014) Identification of T. gondii myosin light chain-1 as a direct target of TachypleginA-2, a small-molecule inhibitor of parasite motility and invasion. PLoS One 9:e98056
Tilley, Lucas D; Krishnamurthy, Shruthi; Westwood, Nicholas J et al. (2014) Identification of TgCBAP, a novel cytoskeletal protein that localizes to three distinct subcompartments of the Toxoplasma gondii pellicle. PLoS One 9:e98492
Bookwalter, Carol S; Kelsen, Anne; Leung, Jacqueline M et al. (2014) A Toxoplasma gondii class XIV myosin, expressed in Sf9 cells with a parasite co-chaperone, requires two light chains for fast motility. J Biol Chem 289:30832-41
Tang, Qing; Andenmatten, Nicole; Hortua Triana, Miryam A et al. (2014) Calcium-dependent phosphorylation alters class XIVa myosin function in the protozoan parasite Toxoplasma gondii. Mol Biol Cell 25:2579-91

Showing the most recent 10 out of 23 publications