Phosphoenolpyruvate carboxylase (PEPC) is essential for the glycolytic pathway and parasite proliferation in Babesia gibsoni
Abstract Apicomplexan parasites predominantly generate ATP and lactic acid through glycolysis and anaerobic glucose metabolism, incorporating CO2 into glycolysis via a stage-dependent phosphoenolpyruvate carboxylase (PEPC) mechanism. Although the role of PEPC in plant and bacterial carbon fixation i...
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BMC
2024-12-01
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| Series: | Animal Diseases |
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| Online Access: | https://doi.org/10.1186/s44149-024-00148-5 |
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| author | Dong-Fang Li Ling-Na Wang Yi-Dan Bai Yu-Xin Yu Xing Lu Xing-Ai Guan Fang-Jie Li Sen Wang Lan He Jun-Long Zhao |
| author_facet | Dong-Fang Li Ling-Na Wang Yi-Dan Bai Yu-Xin Yu Xing Lu Xing-Ai Guan Fang-Jie Li Sen Wang Lan He Jun-Long Zhao |
| author_sort | Dong-Fang Li |
| collection | DOAJ |
| description | Abstract Apicomplexan parasites predominantly generate ATP and lactic acid through glycolysis and anaerobic glucose metabolism, incorporating CO2 into glycolysis via a stage-dependent phosphoenolpyruvate carboxylase (PEPC) mechanism. Although the role of PEPC in plant and bacterial carbon fixation is well documented, its function within Babesia remains largely unexplored. This study employs reverse genetics to probe the biological role of PEPC in Babesia gibsoni, noting its conservation across similar protozoa, suggesting a pivotal and conserved biological function. Western blotting and immunofluorescence (IFA) experiments using the BgPEPC-3 × Flag strain revealed that the BgPEPC protein has a molecular weight of 105 kDa and localizes predominantly to the cytoplasm. Attempts to knock out the PEPC gene in BgPEPC-3 × Flag strains failed under standard media conditions, succeeded only with the addition of 5 mM malate, an upstream metabolite of oxaloacetic acid (OAA). In addition to malate, the downstream metabolite of OAA can also partially compensate for the phenotypic defects caused by PEPC deficiency. This intervention alleviated severe growth deficits, underscoring the critical role of aspartate in the parasite lifecycle. Moreover, metabolic inhibitors such as L-cycloserine and triazamidine, which target aspartate aminotransferase and mitochondrial functions, respectively, demonstrated increased efficacy against BgPEPC knockout strains. The lack of a compensatory response to malic acid supplementation underscores the integral role of BgPEPC in intermediary carbon metabolism and its necessity in providing aspartate as a precursor to pyrimidine synthesis. Collectively, these findings suggest that PEPC could be a potential target for future drug development against B. gibsoni infections. Graphical Abstract |
| format | Article |
| id | doaj-art-55a123763bf74127aab22c249bc18186 |
| institution | OA Journals |
| issn | 2731-0442 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | BMC |
| record_format | Article |
| series | Animal Diseases |
| spelling | doaj-art-55a123763bf74127aab22c249bc181862025-08-20T02:31:02ZengBMCAnimal Diseases2731-04422024-12-014111210.1186/s44149-024-00148-5Phosphoenolpyruvate carboxylase (PEPC) is essential for the glycolytic pathway and parasite proliferation in Babesia gibsoniDong-Fang Li0Ling-Na Wang1Yi-Dan Bai2Yu-Xin Yu3Xing Lu4Xing-Ai Guan5Fang-Jie Li6Sen Wang7Lan He8Jun-Long Zhao9National Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityNational Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural UniversityAbstract Apicomplexan parasites predominantly generate ATP and lactic acid through glycolysis and anaerobic glucose metabolism, incorporating CO2 into glycolysis via a stage-dependent phosphoenolpyruvate carboxylase (PEPC) mechanism. Although the role of PEPC in plant and bacterial carbon fixation is well documented, its function within Babesia remains largely unexplored. This study employs reverse genetics to probe the biological role of PEPC in Babesia gibsoni, noting its conservation across similar protozoa, suggesting a pivotal and conserved biological function. Western blotting and immunofluorescence (IFA) experiments using the BgPEPC-3 × Flag strain revealed that the BgPEPC protein has a molecular weight of 105 kDa and localizes predominantly to the cytoplasm. Attempts to knock out the PEPC gene in BgPEPC-3 × Flag strains failed under standard media conditions, succeeded only with the addition of 5 mM malate, an upstream metabolite of oxaloacetic acid (OAA). In addition to malate, the downstream metabolite of OAA can also partially compensate for the phenotypic defects caused by PEPC deficiency. This intervention alleviated severe growth deficits, underscoring the critical role of aspartate in the parasite lifecycle. Moreover, metabolic inhibitors such as L-cycloserine and triazamidine, which target aspartate aminotransferase and mitochondrial functions, respectively, demonstrated increased efficacy against BgPEPC knockout strains. The lack of a compensatory response to malic acid supplementation underscores the integral role of BgPEPC in intermediary carbon metabolism and its necessity in providing aspartate as a precursor to pyrimidine synthesis. Collectively, these findings suggest that PEPC could be a potential target for future drug development against B. gibsoni infections. Graphical Abstracthttps://doi.org/10.1186/s44149-024-00148-5Babesia gibsoniPhosphoenolpyruvatePhosphoenolpyruvate carboxylaseMalateAspartate |
| spellingShingle | Dong-Fang Li Ling-Na Wang Yi-Dan Bai Yu-Xin Yu Xing Lu Xing-Ai Guan Fang-Jie Li Sen Wang Lan He Jun-Long Zhao Phosphoenolpyruvate carboxylase (PEPC) is essential for the glycolytic pathway and parasite proliferation in Babesia gibsoni Animal Diseases Babesia gibsoni Phosphoenolpyruvate Phosphoenolpyruvate carboxylase Malate Aspartate |
| title | Phosphoenolpyruvate carboxylase (PEPC) is essential for the glycolytic pathway and parasite proliferation in Babesia gibsoni |
| title_full | Phosphoenolpyruvate carboxylase (PEPC) is essential for the glycolytic pathway and parasite proliferation in Babesia gibsoni |
| title_fullStr | Phosphoenolpyruvate carboxylase (PEPC) is essential for the glycolytic pathway and parasite proliferation in Babesia gibsoni |
| title_full_unstemmed | Phosphoenolpyruvate carboxylase (PEPC) is essential for the glycolytic pathway and parasite proliferation in Babesia gibsoni |
| title_short | Phosphoenolpyruvate carboxylase (PEPC) is essential for the glycolytic pathway and parasite proliferation in Babesia gibsoni |
| title_sort | phosphoenolpyruvate carboxylase pepc is essential for the glycolytic pathway and parasite proliferation in babesia gibsoni |
| topic | Babesia gibsoni Phosphoenolpyruvate Phosphoenolpyruvate carboxylase Malate Aspartate |
| url | https://doi.org/10.1186/s44149-024-00148-5 |
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