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Updated: Mar 17, 2026

Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii
Published on: July 12, 2013
Pyrimidine Pathway-Dependent and -Independent Functions of the Toxoplasma gondii Mitochondrial Dihydroorotate
Miryam Andrea Hortua Triana1, Daniela Cajiao Herrera2, Barbara H Zimmermann2
1Department of Microbiology and Immunology, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire, USA.
Abstract:
Dihydroorotate dehydrogenase (DHODH) mediates the fourth step of de novo pyrimidine biosynthesis and is a proven drug target for inducing immunosuppression in therapy of human disease as well as a rapidly emerging drug target for treatment of malaria. In Toxoplasma gondii, disruption of the first, fifth, or sixth step of de novo pyrimidine biosynthesis induced uracil auxotrophy. However, previous attempts to generate uracil auxotrophy by genetically deleting the mitochondrion-associated DHODH of T. gondii (TgDHODH) failed. To further address the essentiality of TgDHODH, mutant gene alleles deficient in TgDHODH activity were designed to ablate the enzyme activity. Replacement of the endogenous DHODH gene with catalytically deficient DHODH gene alleles induced uracil auxotrophy. Catalytically deficient TgDHODH localized to the mitochondria, and parasites retained mitochondrial membrane potential. These results show that TgDHODH is essential for the synthesis of pyrimidines and suggest that TgDHODH is required for a second essential function independent of its role in pyrimidine biosynthesis.
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