Functional analyses of Toxoplasma gondii dihydroorotase reveal a promising anti‐parasitic target

Toxoplasma gondii relies heavily on the de novo pyrimidine biosynthesis pathway for fueling the high uridine‐5′‐monophosphate (UMP) demand during parasite growth. The third step of de novo pyrimidine biosynthesis is catalyzed by dihydroorotase (DHO), a metalloenzyme that catalyzes the reversible con...

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Bibliographic Details
Published in:The FASEB journal 2024-01, Vol.38 (1), p.e23397-n/a
Main Authors: Pan, Ming, Ge, Ceng‐Ceng, Niu, Shui‐Zhu, Duan, Yin‐Yan, Fan, Yi‐Min, Jin, Qi‐Wang, Chen, Xiang, Tao, Jian‐Ping, Huang, Si‐Yang
Format: Article
Language:English
Subjects:
Animals
Dihydroorotase
growth
Mammals
Mice
Parasites
pyrimidine biosynthesis pathway
Pyrimidines - pharmacology
Toxoplasma
Toxoplasma gondii
Uracil
Uridine Monophosphate
uridine‐5′‐monophosphate
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Summary:Toxoplasma gondii relies heavily on the de novo pyrimidine biosynthesis pathway for fueling the high uridine‐5′‐monophosphate (UMP) demand during parasite growth. The third step of de novo pyrimidine biosynthesis is catalyzed by dihydroorotase (DHO), a metalloenzyme that catalyzes the reversible condensation of carbamoyl aspartate to dihydroorotate. Here, functional analyses of TgDHO reveal that tachyzoites lacking DHO are impaired in overall growth due to decreased levels of UMP, and the noticeably growth restriction could be partially rescued after supplementation with uracil or high concentrations of L‐dihydroorotate in vitro. When pyrimidine salvage pathway is disrupted, both DHOH35A and DHOD284E mutant strains proliferated much slower than DHO‐expressing parasites, suggesting an essential role of both TgDHO His35 and Asp284 residues in parasite growth. Additionally, DHO deletion causes the limitation of bradyzoite growth under the condition of uracil supplementation or uracil deprivation. During the infection in mice, the DHO‐deficient parasites are avirulent, despite the generation of smaller tissue cysts. The results reveal that TgDHO contributes to parasite growth both in vitro and in vivo. The significantly differences between TgDHO and mammalian DHO reflect that DHO can be exploited to produce specific inhibitors targeting apicomplexan parasites. Moreover, potential DHO inhibitors exert beneficial effects on enzymatic activity of TgDHO and T. gondii growth in vitro. In conclusion, these data highlight the important role of TgDHO in parasite growth and reveal that it is a promising anti‐parasitic target for future control of toxoplasmosis. Toxoplasma gondii lacking DHO are impaired in intracellular replication, due to decreased levels of UMP. The noticeably growth restriction could be partially restored after supplementation with uracil or high concentrations of L‐dihydroorotate. Disruption of pyrimidine salvage pathway leads to a severely growth limitation in DHO mutant strains compared to a normal proliferation of DHO complementation strain. Due to the differences between TgDHO and mammalian DHO, DHO inhibitors PLU exerts beneficial effects on TgDHO activity and T. gondii growth.
ISSN:0892-6638
1530-6860
DOI:10.1096/fj.202301493R