Toxoplasma gondii is an important opportunistic infection of AIDS patients. Improved strategies and approaches are urgently needed to more effectively prevent and treat recurrent infections during AIDS. Due to the ease of in vitro culture and cloning, efficiency of transformation, abundance of established genetic tools, and availability of murine models of infection, Toxoplasma gondii is increasingly recognized as a model apicomplexan parasite. T. gondii is also being adopted as a surrogate parasite model to investigate biology of Cryptosporidium parvum, another very important opportunistic pathogen in the context of the AIDS pandemic, and other important apicomplexan parasites such as Plasmodium and Eimeria. To more effectively utilize T. gondii as a surrogate model organism for other parasites as well as to accelerate the development of improved prevention and treatment strategies for Toxoplasmosis, we propose to develop genetically improved model organism T gondii parasites with an enhanced efficiency of homologous recombination. Such model T. gondii parasites could be used to improve model systems for heterologous expression of genes from C. parvum or other pathogens and to develop genetically defined vaccine strains. T. gondii strains with enhanced homologous recombination can potentially accelerate functional genomics to more effectively identify and validate new drug targets, as well as to enhance aspects of biological discovery in the post-genomic era of T. gondii. To construct these novel T. gondii parasites, we propose to develop genetically defined parasite strains that are deficient in nonhomologous end-joining recombination repair based on the genetic construction of defined knockouts in the T. gondii gene homologs corresponding to KU70 and KU80. We predict that KU70 as well as KU80 knockouts in T. gondii will result in a phenotype of significantly enhanced homologous recombination. These studies will develop and validate strains that are likely to be useful model organisms for the more efficient construction of defined gene knockouts and gene manipulations in Toxoplasma gondii. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21AI073142-01A1
Application #
7275872
Study Section
Special Emphasis Panel (ZRG1-AARR-C (03))
Program Officer
Mcgugan, Glen C
Project Start
2007-03-15
Project End
2009-02-28
Budget Start
2007-03-15
Budget End
2008-02-29
Support Year
1
Fiscal Year
2007
Total Cost
$199,875
Indirect Cost
Name
Dartmouth College
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
041027822
City
Hanover
State
NH
Country
United States
Zip Code
03755
Baird, Jason R; Byrne, Katelyn T; Lizotte, Patrick H et al. (2013) Immune-mediated regression of established B16F10 melanoma by intratumoral injection of attenuated Toxoplasma gondii protects against rechallenge. J Immunol 190:469-78
Rommereim, Leah M; Hortua Triana, Miryam A; Falla, Alejandra et al. (2013) Genetic manipulation in ?ku80 strains for functional genomic analysis of Toxoplasma gondii. J Vis Exp :e50598
Hortua Triana, Miryam Andrea; Huynh, My-Hang; Garavito, Manuel F et al. (2012) Biochemical and molecular characterization of the pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase from Toxoplasma gondii. Mol Biochem Parasitol 184:71-81
Denkers, Eric Y; Bzik, David J; Fox, Barbara A et al. (2012) An inside job: hacking into Janus kinase/signal transducer and activator of transcription signaling cascades by the intracellular protozoan Toxoplasma gondii. Infect Immun 80:476-82
Fox, Barbara A; Falla, Alejandra; Rommereim, Leah M et al. (2011) Type II Toxoplasma gondii KU80 knockout strains enable functional analysis of genes required for cyst development and latent infection. Eukaryot Cell 10:1193-206
Butcher, Barbara A; Fox, Barbara A; Rommereim, Leah M et al. (2011) Toxoplasma gondii rhoptry kinase ROP16 activates STAT3 and STAT6 resulting in cytokine inhibition and arginase-1-dependent growth control. PLoS Pathog 7:e1002236
Charles, Elizabeth; Joshi, Sunil; Ash, John D et al. (2010) CD4 T-cell suppression by cells from Toxoplasma gondii-infected retinas is mediated by surface protein PD-L1. Infect Immun 78:3484-92
Fentress, Sarah J; Behnke, Michael S; Dunay, Ildiko R et al. (2010) Phosphorylation of immunity-related GTPases by a Toxoplasma gondii-secreted kinase promotes macrophage survival and virulence. Cell Host Microbe 8:484-95
Fox, Barbara A; Bzik, David J (2010) Avirulent uracil auxotrophs based on disruption of orotidine-5'-monophosphate decarboxylase elicit protective immunity to Toxoplasma gondii. Infect Immun 78:3744-52
Fox, Barbara A; Ristuccia, Jessica G; Bzik, David J (2009) Genetic identification of essential indels and domains in carbamoyl phosphate synthetase II of Toxoplasma gondii. Int J Parasitol 39:533-9

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