Trypanosoma cruzi epimastigotes lack ornithine decarboxylase but can express a foreign gene encoding this enzyme

Carrillo, Carolina; Cejas, Silvina; González, Nélida S.; Algranati, Israel D.

doi:10.1016/s0014-5793(99)00804-2

Cited by 58 publications

(42 citation statements)

References 31 publications

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“…However, T. cruzi is one of the extremely few eukaryotic organisms auxotrophic for polyamines. We have previously demonstrated that T. cruzi epimastigotes are unable to synthesize putrescine de novo because they contain neither ODC nor ADC enzymatic activity, due to the absence of ODC and ADC genes [38, 41, 42]. This conclusion is also supported by data from the T. cruzi genome project [30].…”

Section: Introductionmentioning

confidence: 88%

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Trypanosoma cruzi Coexpressing Ornithine Decarboxylase and Green Fluorescence Proteins as a Tool to Study the Role of Polyamines in Chagas Disease Pathology

et al. 2011

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Polyamines are essential for Trypanosoma cruzi, the causative agent of Chagas disease. As T. cruzi behaves as a natural auxotrophic organism, it relies on host polyamines biosynthesis. In this paper we obtained a double-transfected T. cruzi parasite that expresses the green fluorescent protein (GFP) and a heterologous ornithine decarboxylase (ODC), used itself as a novel selectable marker. These autotrophic and fluorescent parasites were characterized; the ODC presented an apparent Km for ornithine of 0.51 ± 0.16 mM and an estimated Vmax value of 476.2 nmoles/h/mg of protein. These expressing ODC parasites showed higher metacyclogenesis capacity than the auxotrophic counterpart, supporting the idea that polyamines are engaged in this process. This double-transfected T. cruzi parasite results in a powerful tool—easy to follow by its fluorescence—to study the role of polyamines in Chagas disease pathology and in related processes such as parasite survival, invasion, proliferation, metacyclogenesis, and tissue spreading.

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Section: Introductionmentioning

confidence: 88%

“…The latter is encoded by the ODC gene, an auxotrophy rescuing gene used here as a selectable marker. Then we selected a Y-GFP-ODC clone with high expression of both GFP [7] and ODC activity [38]. These autotrophic and fluorescent parasites were characterized as well as the heterologous ODC enzyme activity.…”

Section: Introductionmentioning

confidence: 99%

Trypanosoma cruzi Coexpressing Ornithine Decarboxylase and Green Fluorescence Proteins as a Tool to Study the Role of Polyamines in Chagas Disease Pathology

et al. 2011

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“…The supply of essential polyamines in T . cruzi is exclusively achieved through transport processes, a clear case of metabolic-transport replacement in the evolutionary adaptation to parasitism [15]. Interestingly, Tc PAT12 (also known as Tc POT1) is the only polyamine transporter present in T .…”

Section: Introductionmentioning

confidence: 99%

Trypanocidal Effect of Isotretinoin through the Inhibition of Polyamine and Amino Acid Transporters in Trypanosoma cruzi

et al. 2017

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Polyamines are essential compounds to all living organisms and in the specific case of Trypanosoma cruzi, the causative agent of Chagas disease, they are exclusively obtained through transport processes since this parasite is auxotrophic for polyamines. Previous works reported that retinol acetate inhibits Leishmania growth and decreases its intracellular polyamine concentration. The present work describes a combined strategy of drug repositioning by virtual screening followed by in vitro assays to find drugs able to inhibit TcPAT12, the only polyamine transporter described in T. cruzi. After a screening of 3000 FDA-approved drugs, 7 retinoids with medical use were retrieved and used for molecular docking assays with TcPAT12. From the docked molecules, isotretinoin, a well-known drug used for acne treatment, showed the best interaction score with TcPAT12 and was selected for further in vitro studies. Isotretinoin inhibited the polyamine transport, as well as other amino acid transporters from the same protein family (TcAAAP), with calculated IC50 values in the range of 4.6–10.3 μM. It also showed a strong inhibition of trypomastigote burst from infected cells, with calculated IC50 of 130 nM (SI = 920) being significantly less effective on the epimastigote stage (IC50 = 30.6 μM). The effect of isotretinoin on the parasites plasma membrane permeability and on mammalian cell viability was tested, and no change was observed. Autophagosomes and apoptotic bodies were detected as part of the mechanisms of isotretinoin-induced death indicating that the inhibition of transporters by isotretinoin causes nutrient starvation that triggers autophagic and apoptotic processes. In conclusion, isotretinoin is a promising trypanocidal drug since it is a multi-target inhibitor of essential metabolites transporters, in addition to being an FDA-approved drug largely used in humans, which could reduce significantly the requirements for its possible application in the treatment of Chagas disease.

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“…T. cruzi epimastigotes are auxotrophic for diamines, such as putrescine since T. cruzi , unlike other trypanosomatids, lacks the genes encoding either ornithine decarboxylase or arginine decarboxylase (ODC and ADC), enzymes responsible for putrescine biosynthesis [21–23]. Diamines are essential for cell proliferation, differentiation and macromolecular synthesis [24].…”

Section: Resultsmentioning

confidence: 99%

LM14 defined medium enables continuous growth of Trypanosoma cruzi

et al. 2014

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BackgroundTrypanosoma cruzi, the etiologic agent of Chagas disease, alternates between distinct morphological and functional forms during its life cycle. Axenic multiplication and differentiation processes of this protozoan parasite can be reproduced in vitro, enabling the isolation and study of the different evolutionary forms. Although there are several publications attempting the cultivation of T. cruzi under chemically defined conditions, in our experience none of the published media are capable of maintaining T. cruzi in continuous growth.ResultsIn this work we modified a known chemically defined medium for Trypanosoma brucei growth. The resulting LM14 and LM14B defined media enabled cultivation of five different strains of T. cruzi for more than forty passages until now. The parasite’s biological characteristics such as morphology and differentiation to metacyclic trypomastigotes were maintained when defined media is used.ConclusionsThe establishment of a defined medium for T. cruzi cultivation is an important tool for basic biological research allowing several different approaches, providing new perspectives for further studies related to cell biology of this parasite.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-014-0238-y) contains supplementary material, which is available to authorized users.

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Trypanosoma cruzi epimastigotes lack ornithine decarboxylase but can express a foreign gene encoding this enzyme

Cited by 58 publications

References 31 publications

Trypanosoma cruzi Coexpressing Ornithine Decarboxylase and Green Fluorescence Proteins as a Tool to Study the Role of Polyamines in Chagas Disease Pathology

Trypanosoma cruzi Coexpressing Ornithine Decarboxylase and Green Fluorescence Proteins as a Tool to Study the Role of Polyamines in Chagas Disease Pathology

Trypanocidal Effect of Isotretinoin through the Inhibition of Polyamine and Amino Acid Transporters in Trypanosoma cruzi

LM14 defined medium enables continuous growth of Trypanosoma cruzi

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