End products and enzyme levels of aerobic glucose fermentation in trypanosomatids

Cazzulo, Juán José; Cazzulo, Berta M. Franke de; Engel, Juan C.; Cannata, Joaquı́n J.B.

doi:10.1016/0166-6851(85)90074-x

Cited by 198 publications

(120 citation statements)

References 23 publications

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“…Briefly, epimastigotes were grown axenically in a medium containing brain-heart infusiontryptose medium and fetal calf serum (BHT) as described (20). Metacyclic trypomastigotes were obtained by spontaneous differentiation of epimastigotes at 28°C, followed by purification by DEAE-cellulose chromatography (21).…”

Section: Methodsmentioning

confidence: 99%

SUMOylation Pathway in Trypanosoma cruzi: Functional Characterization and Proteomic Analysis of Target Proteins

Bayona

Nakayasu

Laverrière

et al. 2011

Molecular & Cellular Proteomics

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SUMOylation is a relevant protein post-translational modification in eukaryotes. The C terminus of proteolytically activated small ubiquitin-like modifier (SUMO) is covalently linked to a lysine residue of the target protein by an isopeptide bond, through a mechanism that includes an E1-activating enzyme, an E2-conjugating enzyme, and transfer to the target, sometimes with the assistance of a ligase. The modification is reversed by a protease, also responsible for SUMO maturation. A number of proteins have been identified as SUMO targets, participating in the regulation of cell cycle progression, transcription, translation, ubiquitination, and DNA repair. In this study, we report that orthologous genes corresponding to the SUMOylation pathway are present in the etiological agent of Chagas disease, Trypanosoma cruzi. Furthermore, the SUMOylation system is functionally active in this protozoan parasite, having the requirements for SUMO maturation and conjugation. Immunofluorescence analysis showed that T. cruzi SUMO (TcSUMO) is predominantly found in the nucleus. To identify SUMOylation targets and get an insight into their physiological roles we generated transfectant T. cruzi epimastigote lines expressing a double-tagged T. cruzi SUMO, and SUMOylated proteins were enriched by tandem affinity chromatography. By two-dimensional liquid chromatography-mass spectrometry a total of 236 proteins with diverse biological functions were identified as potential T. cruzi SUMO targets. Of these, metacaspase-3 was biochemically validated as a bona fide SUMOylation substrate. Proteomic studies in other organisms have reported that orthologs of putative T.

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Section: Methodsmentioning

confidence: 99%

SUMOylation Pathway in Trypanosoma cruzi: Functional Characterization and Proteomic Analysis of Target Proteins

Bayona

Nakayasu

Laverrière

et al. 2011

Molecular & Cellular Proteomics

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“…The Tc45 coding DNA sequence was amplified by PCR (using the primers described above) and a radiolabeled probe was ob- tained according to standard procedures. 29 Tulahuén and CL Brener epimastigote chromosomes 33 were separated by Pulse Field Gel Electrophoresis (PFGE) (CHEF DR II Bio-Rad) using standard procedures. [34][35][36] The DNA was transferred onto nylon membranes.…”

mentioning

confidence: 99%

Tc45, a dimorphic Trypanosoma cruzi immunogen with variable chromosomal localization, is calreticulin.

Aguillón

Ferreira²,

Pérez³

et al. 2000

The American Journal of Tropical Medicine and Hygiene

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Abstract. We demonstrate that Tc45, a polypeptide described as an immunogenetically restricted Trypanosoma cruzi antigen in mice, is calreticulin, a dimorphic molecule encoded by genes with variable chromosomal distribution. Previously we showed that IgG from A.SW (H2 s ) mice immunized with T. cruzi trypomastigotes or epimastigotes and sera from infected humans recognize Tc45, a 45 kD parasite polypeptide. Herein we describe the cloning, sequencing, and expression of the Tc45 gene. A 98% homology in the deduced amino acid sequence was found with a T. cruzi calreticulin-like molecule and 41% with Leishmania donovani and human calreticulin. In the T. cruzi CL Brener clone and in the Tulahuén strain, the gene is located in two and four chromosomes, respectively. Calreticulin was detected in several T. cruzi clones, in the Tulahuén strain, and in T. rangeli, displaying alternative 43 and 46 kD forms.

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“…In L. mexicana parasites, the enzymatic activities of the key enzymes in gluconeogenesis PEPCK and FBPase have been detected (41,42), and PEPCK has been purified and partially characterized (43); however, reports concerning pyruvate carboxylase activity are contradictory (41,42,44,45). Searching the genome data base from L. major using the BLAST algorithm, we found homologous genes for only FBPase (LmjF04.1160) and PEPCK (LmjF27.1810 and LmjF27.1805), the same enzymes whose activities have been clearly detected experimentally, but not for pyruvate carboxylase or glucose-6-phosphatase.…”

Section: Discussionmentioning

confidence: 99%

Metabolic Changes in Glucose Transporter-deficient Leishmania mexicana and Parasite Virulence

Rodriguez‐Contreras

Landfear

2006

Journal of Biological Chemistry

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Leishmania mexicana are parasitic protozoa that express a variety of glycoconjugates that play important roles in their biology as well as the storage carbohydrate ␤-mannan, which is an essential virulence factor for survival of intracellular amastigote forms in the mammalian host. Glucose transporter null mutants, which are viable as insect form promastigotes but not as amastigotes, do not take up glucose and other hexoses but are still able to synthesize these glycoconjugates and ␤-mannan, although at reduced levels. Synthesis of these carbohydratecontaining macromolecules could be accounted for by incorporation of non-carbohydrate precursors into carbohydrates by gluconeogenesis. However, the significantly reduced level of the virulence factor ␤-mannan in the glucose transporter null mutants compared with wild-type parasites may contribute to the non-viability of these null mutants in the disease-causing amastigote stage of the life cycle.Leishmaniae are pathogenic protozoa of the order Kinetoplastida that cause important diseases in humans and other vertebrates (1). There are two principal stages in the parasite life cycle, (i) extracellular flagellated promastigotes that live within the alimentary tract of the sand fly vector and (ii) nonflagellated amastigotes that reside within the acidified phagolysosomal vesicles of vertebrate host macrophages. Both life cycle stages of Leishmania mexicana can be cultivated in vitro (2) facilitating biochemical and genetic studies. Uptake and metabolism of glucose in Leishmania species has been of considerable interest, as this sugar provides a major source of carbon and energy to these parasites (3). In L. mexicana, glucose catabolism is 10 -20-fold higher in the promastigotes compared with amastigotes (4). Furthermore, amastigotes transport glucose at a much lower rate than promastigotes and derive metabolic energy primarily from fatty acid oxidation (4, 5). The higher rates of glucose uptake and utilization by promastigotes in relation to amastigotes is consistent with the high concentrations of sugars that promastigotes experience from the plant sap ingested by the sand fly vector (6) compared with the apparently sugar-limited environment that amastigotes experience (7) in the macrophage phagolysosome.The L. mexicana genome encompasses a cluster of three glucose transporter genes, LmGT1, LmGT2, and LmGT3, which encode closely related isoforms (8). A L. mexicana glucose transporter "knock-out" line (⌬lmgt) (9), in which the entire cluster of glucose transporter genes was eliminated by targeted gene replacement, exhibited no detectable glucose transport activity. The ⌬lmgt promastigotes were able to grow, although at a reduced rate compared with that of wild-type cells. In contrast, ⌬lmgt cells exhibited dramatically reduced viability inside macrophages compared with wild-type parasites and were unable to grow as axenic amastigotes, demonstrating that glucose transporters are essential for amastigote viability. Furthermore, axenically cultured wild-type amastigotes re...

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End products and enzyme levels of aerobic glucose fermentation in trypanosomatids

Cited by 198 publications

References 23 publications

SUMOylation Pathway in Trypanosoma cruzi: Functional Characterization and Proteomic Analysis of Target Proteins

SUMOylation Pathway in Trypanosoma cruzi: Functional Characterization and Proteomic Analysis of Target Proteins

Tc45, a dimorphic Trypanosoma cruzi immunogen with variable chromosomal localization, is calreticulin.

Metabolic Changes in Glucose Transporter-deficient Leishmania mexicana and Parasite Virulence

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