9 research outputs found
Metabolic selection of a homologous recombination-mediated gene loss protects Trypanosoma brucei from ROS production by glycosomal fumarate reductase
The genome of trypanosomatids rearranges by using repeated sequences as platforms for amplification or deletion of genomic segments. These stochastic recombination events have a direct impact on gene dosage and foster the selection of adaptive traits in response to environmental pressure. We provide here such an example by showing that the phosphoenolpyruvate carboxykinase (PEPCK) gene knockout (Îpepck) leads to the selection of a deletion event between two tandemly arranged fumarate reductase (FRDg and FRDm2) genes to produce a chimeric FRDg-m2 gene in the Îpepckâ cell line. FRDg is expressed in peroxisome-related organelles, named glycosomes, expression of FRDm2 has not been detected to date, and FRDg-m2 is nonfunctional and cytosolic. Re-expression of FRDg significantly impaired growth of the Îpepckâ cells, but FRD enzyme activity was not required for this negative effect. Instead, glycosomal localization as well as the covalent flavinylation motif of FRD is required to confer growth retardation and intracellular accumulation of reactive oxygen species (ROS). The data suggest that FRDg, similar to Escherichia coli FRD, can generate ROS in a flavin-dependent process by transfer of electrons from NADH to molecular oxygen instead of fumarate when the latter is unavailable, as in the Îpepck background. Hence, growth retardation is interpreted as a consequence of increased production of ROS, and rearrangement of the FRD locus liberates Îpepckâ cells from this obstacle. Interestingly, intracellular production of ROS has been shown to be required to complete the parasitic cycle in the insect vector, suggesting that FRDg may play a role in this proces
PLoS Biol
Microorganisms must make the right choice for nutrient consumption to adapt to their changing environment. As a consequence, bacteria and yeasts have developed regulatory mechanisms involving nutrient sensing and signaling, known as "catabolite repression," allowing redirection of cell metabolism to maximize the consumption of an energy-efficient carbon source. Here, we report a new mechanism named "metabolic contest" for regulating the use of carbon sources without nutrient sensing and signaling. Trypanosoma brucei is a unicellular eukaryote transmitted by tsetse flies and causing human African trypanosomiasis, or sleeping sickness. We showed that, in contrast to most microorganisms, the insect stages of this parasite developed a preference for glycerol over glucose, with glucose consumption beginning after the depletion of glycerol present in the medium. This "metabolic contest" depends on the combination of 3 conditions: (i) the sequestration of both metabolic pathways in the same subcellular compartment, here in the peroxisomal-related organelles named glycosomes; (ii) the competition for the same substrate, here ATP, with the first enzymatic step of the glycerol and glucose metabolic pathways both being ATP-dependent (glycerol kinase and hexokinase, respectively); and (iii) an unbalanced activity between the competing enzymes, here the glycerol kinase activity being approximately 80-fold higher than the hexokinase activity. As predicted by our model, an approximately 50-fold down-regulation of the GK expression abolished the preference for glycerol over glucose, with glucose and glycerol being metabolized concomitantly. In theory, a metabolic contest could be found in any organism provided that the 3 conditions listed above are met
Procyclic trypanosomes recycle glucose catabolites and TCA cycle intermediates to stimulate growth in the presence of physiological amounts of proline
Trypanosoma brucei, a protist responsible for human African trypanosomiasis (sleeping sickness), is transmitted by the tsetse fly where the procyclic forms of the parasite develop in the proline-rich (1â2 mM) and glucose-depleted digestive tract. Proline is essential for the midgut colonization of the parasite in the insect vector, however other carbon sources could be available and used to feed its central metabolism. Here we show that procyclic trypanosomes can consume and metabolize metabolic intermediates, including those excreted from glucose catabolism (succinate, alanine and pyruvate), with the exception of acetate, which is the ultimate end-product excreted by the parasite. Among the tested metabolites, tricarboxylic acid (TCA) cycle intermediates (succinate, malate and α-ketoglutarate) stimulated growth of the parasite in the presence of 2 mM proline. The pathways used for their metabolism were mapped by proton-NMR metabolic profiling and phenotypic analyses of thirteen RNAi and/or null mutants affecting central carbon metabolism. We showed that (i) malate is converted to succinate by both the reducing and oxidative branches of the TCA cycle, which demonstrates that procyclic trypanosomes can use the full TCA cycle, (ii) the enormous rate of α-ketoglutarate consumption (15-times higher than glucose) is possible thanks to the balanced production and consumption of NADH at the substrate level and (iii) α-ketoglutarate is toxic for trypanosomes if not appropriately metabolized as observed for an α-ketoglutarate dehydrogenase null mutant. In addition, epimastigotes produced from procyclics upon overexpression of RBP6 showed a growth defect in the presence of 2 mM proline, which is rescued by α-ketoglutarate, suggesting that physiological amounts of proline are not sufficient per se for the development of trypanosomes in the fly. In conclusion, these data show that trypanosomes can metabolize multiple metabolites, in addition to proline, which allows them to confront challenging environments in the fly
UDP-glucose pyrophosphorylase (UGP) : import into glycosomes and implication in glycosomal and cytosolic nucleotide sugar biosynthetic pathways in Trypanosoma brucei
Trypanosoma brucei, un protiste responsable de la Trypanosomose Humaine Africaine, Ă©galement connue sous le nom de la maladie du sommeil, est transmis par la mouche tsĂ©-tsĂ© (Glossina sp.). La dĂ©couverte d'organites de type peroxysome spĂ©cialisĂ©s dans la glycolyse, appelĂ©s glycosomes, a soulevĂ© un certain nombre de questions sur le rĂŽle de cet organite dans la biologie des trypanosomes. Plusieurs voies mĂ©taboliques prĂ©sentes dans le cytosol d'autres eucaryotes, comme la glycolyse et la biosynthĂšse des sucres nuclĂ©otidiques, sont compartimentĂ©es dans les glycosomes. Les raisons et les avantages de la prĂ©sence des enzymes glycolytiques dans l'organite ont Ă©tĂ© largement discutĂ©s, mais la fonctionnalitĂ© et le rĂŽle des voies de biosynthĂšse des sucres nuclĂ©otidiques glycosomales ne sont pas connus. Notre Ă©tude s'est focalisĂ©e sur l'UDP-glucose pyrophosphorylase (UGP), une enzyme impliquĂ©e dans la synthĂšse de l'UDP-glucose (UDP-Glc). Sur la base de la double localisation glycosomale et cytosolique de l'UGP mise en Ă©vidence ici Ă l'aide de plusieurs techniques de localisation subcellulaire, nous avons abordĂ© deux questions en utilisant comme modĂšle les formes procycliques de T. brucei prĂ©sentes dans l'insecte vecteur. La premiĂšre est liĂ©e au mĂ©canisme d'import de l'UGP dans les glycosomes, car cette protĂ©ine ne possĂšde aucun signal d'adressage aux peroxysomes de type PTS1 ou PTS2. Nous avons montrĂ© que l'UGP est importĂ©e dans les glycosomes par "piggybacking" en s'associant Ă la phosphoĂ©nolpyruvate dĂ©carboxylase (PEPCK) possĂ©dant un signal dâadressage PTS1. Les interactions entre l'UGP et la PEPCK ont Ă©tĂ© montrĂ©es in situ et l'identification les rĂ©gions impliquĂ©es dans ces interactions ont Ă©tĂ© identifiĂ©es. Nos rĂ©sultats suggĂšrent que le complexe UGP-PEPCK est formĂ© de maniĂšre transitoire lors de son import dans les glycosomes nouvellement produits et compĂ©tents pour l'import des protĂ©ines. La seconde question concerne le rĂŽle de l'UGP dans les glycosomes. Nous avons montrĂ© que l'UGP est essentielle Ă la croissance des trypanosomes et que les voies mĂ©taboliques glycosomales et cytosoliques dont l'UGP fait partie sont fonctionnelles. En effet, des mutants viables contenant l'UGP exclusivement dans les glycosomes ou dans le cytosol sont viables et produisent des quantitĂ©s similaires d'UDP-Glc. La raison d'ĂȘtre de la production glycosomale d'UDP-Glc par l'UGP reste inconnue, mais n'est probablement pas liĂ©e aux rĂ©actions de glycosylation, Ă©tant donnĂ© qu'aucune glycosyltransfĂ©rase n'a Ă©tĂ© dĂ©tectĂ©e dans l'organite.Un autre aspect de ce travail concerne le rĂŽle des intermĂ©diaires du cycle de l'acide tricarboxylique (TCA) dans le mĂ©tabolisme mitochondrial des formes procycliques. Dans le tractus digestif de son insecte vecteur, les trypanosomes dĂ©pendent de la proline pour alimenter leur mĂ©tabolisme Ă©nergĂ©tique. Cependant, la disponibilitĂ© d'Ă©ventuelles autres sources de carbone pouvant ĂȘtre utilisĂ©es par le parasite est actuellement inconnue. Nous avons montrĂ© que les intermĂ©diaires du cycle TCA, i.e. succinate, malate et a-cĂ©toglutarate, stimulent la croissance des formes procycliques incubĂ©es dans un milieu contenant 2 mM de proline, concentration se situant dans la gamme des quantitĂ©s mesurĂ©es dans l'intestin de la mouche. De plus, le dĂ©veloppement de nouvelles approches ont permis d'Ă©tudier une branche peu explorĂ©e du cycle TCA convertissant le malate en a-cĂ©toglutarate, prĂ©cĂ©demment dĂ©crite comme peu ou pas utilisĂ©e par le parasite, quellles que soient les quantitĂ©s de glucose disponibles. L'activitĂ© de cette branche suggĂšre qu'un cycle TCA complet peut ĂȘtre mis en Ćuvre dans les formes procycliques et probablement dans les autres formes parasitaires de l'insecte. Nos donnĂ©es Ă©largissent le potentiel mĂ©tabolique des trypanosomes et ouvrent la voie vers une meilleure comprĂ©hension du mĂ©tabolisme de ce parasite dans divers organes de la mouche tsĂ©-tsĂ©, oĂč il Ă©volue.Trypanosoma brucei, a protist responsible for human African trypanosomiasis, also known as sleeping sickness, is transmitted by the tsetse fly (Glossina sp.). The discovery of peroxisome-like organelles specialized in glycolysis called glycosomes, has raised a number of questions about the role of this organelle in the biology of trypanosomes. Several metabolic pathways present in the cytosol of eukaryotes, like glycolysis and sugar nucleotides biosynthesis, are compartmentalized within glycosomes. While the reasons and advantages of having glycolytic enzymes compartmentalized in the organelle have been extensively discussed, little is proposed for sugar nucleotides biosynthetic pathways. This study is focused on the UDP-glucose pyrophosphorylase (UGP), an enzyme involved in the synthesis of UDP-glucose (UDP-Glc). Based on the UGP's dual glycosomal and cytosolic localization evidenced here using several subcellular localization techniques, we addressed two questions using as a model the procyclic forms of T. brucei present in the insect vector. The first one is related to the mechanism of UGP import into glycosomes, since this protein lacks any known peroxisomal targeting signal (PTS1 and PTS2). We demonstrated that UGP is imported into the organelle by piggybacking on the glycosomal PTS1-containing phosphoenolpyruvate decarboxylase (PEPCK). Interactions between UGP and PEPCK have been showed in situ and the interacting regions have been identified. Our data suggest that the complex UGP-PEPCK is formed transiently to facilitate the import of UGP and that it is detected in newly formed import-competent glycosomes. The second question concerns the role of UGP in glycosomes. We demonstrated that UGP is essential for the growth of trypanosomes and that mutants containing UGP exclusively in glycosomes or in the cytosol still produce UDP-Glc at similar levels and are viable, which implies that the glycosomal and cytosolic metabolic pathways involving UGP are functional. The glycosomal function of UDP-Glc is currently unknown and probably not related to glycosylation reactions, since no glycosyltransferases have been detected in the organelle.Another aspect of this work concerns the role of tricarboxylic acid (TCA) cycle intermediates in the mitochondrial metabolism of the procyclic trypanosomes. In the midgut of its insect vector, trypanosomes rely on proline to feed their energy metabolism. However, the availability of other potential carbon sources that can be used by the parasite is currently unknown. We showed that TCA cycle intermediates, i.e. succinate, malate and a-ketoglutarate, stimulate growth of procyclic trypanosomes incubated in medium containing 2 mM proline, which is in the range of the amounts measured in the midgut of the fly. In addition, we have implemented new approaches to study cell growth and metabolic pathways in order to investigate mitochondrial metabolism. These new tools have allowed us to study a poorly explored branch of the TCA cycle converting malate to a-ketoglutarate, which was previously described as non-functional or little used in the parasite, regardless of the glucose levels available. The discovery of this branch reveals that a full TCA cycle can operate in procyclic trypanosomes and probably in the other trypanosome forms present in the fly. Our data broaden the metabolic potential of trypanosomes and pave the way for a better understanding of the parasite's metabolism in various organ systems of the tsetse fly, where it evolves
UDP-glucose pyrophosphorylase (UGP) : import dans les glycosomes et implication dans la biosynthÚse glycosomale et cytosolique des sucres nucléotidiques chez Trypanosoma brucei
Trypanosoma brucei, a protist responsible for human African trypanosomiasis, also known as sleeping sickness, is transmitted by the tsetse fly (Glossina sp.). The discovery of peroxisome-like organelles specialized in glycolysis called glycosomes, has raised a number of questions about the role of this organelle in the biology of trypanosomes. Several metabolic pathways present in the cytosol of eukaryotes, like glycolysis and sugar nucleotides biosynthesis, are compartmentalized within glycosomes. While the reasons and advantages of having glycolytic enzymes compartmentalized in the organelle have been extensively discussed, little is proposed for sugar nucleotides biosynthetic pathways. This study is focused on the UDP-glucose pyrophosphorylase (UGP), an enzyme involved in the synthesis of UDP-glucose (UDP-Glc). Based on the UGP's dual glycosomal and cytosolic localization evidenced here using several subcellular localization techniques, we addressed two questions using as a model the procyclic forms of T. brucei present in the insect vector. The first one is related to the mechanism of UGP import into glycosomes, since this protein lacks any known peroxisomal targeting signal (PTS1 and PTS2). We demonstrated that UGP is imported into the organelle by piggybacking on the glycosomal PTS1-containing phosphoenolpyruvate decarboxylase (PEPCK). Interactions between UGP and PEPCK have been showed in situ and the interacting regions have been identified. Our data suggest that the complex UGP-PEPCK is formed transiently to facilitate the import of UGP and that it is detected in newly formed import-competent glycosomes. The second question concerns the role of UGP in glycosomes. We demonstrated that UGP is essential for the growth of trypanosomes and that mutants containing UGP exclusively in glycosomes or in the cytosol still produce UDP-Glc at similar levels and are viable, which implies that the glycosomal and cytosolic metabolic pathways involving UGP are functional. The glycosomal function of UDP-Glc is currently unknown and probably not related to glycosylation reactions, since no glycosyltransferases have been detected in the organelle.Another aspect of this work concerns the role of tricarboxylic acid (TCA) cycle intermediates in the mitochondrial metabolism of the procyclic trypanosomes. In the midgut of its insect vector, trypanosomes rely on proline to feed their energy metabolism. However, the availability of other potential carbon sources that can be used by the parasite is currently unknown. We showed that TCA cycle intermediates, i.e. succinate, malate and a-ketoglutarate, stimulate growth of procyclic trypanosomes incubated in medium containing 2 mM proline, which is in the range of the amounts measured in the midgut of the fly. In addition, we have implemented new approaches to study cell growth and metabolic pathways in order to investigate mitochondrial metabolism. These new tools have allowed us to study a poorly explored branch of the TCA cycle converting malate to a-ketoglutarate, which was previously described as non-functional or little used in the parasite, regardless of the glucose levels available. The discovery of this branch reveals that a full TCA cycle can operate in procyclic trypanosomes and probably in the other trypanosome forms present in the fly. Our data broaden the metabolic potential of trypanosomes and pave the way for a better understanding of the parasite's metabolism in various organ systems of the tsetse fly, where it evolves.Trypanosoma brucei, un protiste responsable de la Trypanosomose Humaine Africaine, Ă©galement connue sous le nom de la maladie du sommeil, est transmis par la mouche tsĂ©-tsĂ© (Glossina sp.). La dĂ©couverte d'organites de type peroxysome spĂ©cialisĂ©s dans la glycolyse, appelĂ©s glycosomes, a soulevĂ© un certain nombre de questions sur le rĂŽle de cet organite dans la biologie des trypanosomes. Plusieurs voies mĂ©taboliques prĂ©sentes dans le cytosol d'autres eucaryotes, comme la glycolyse et la biosynthĂšse des sucres nuclĂ©otidiques, sont compartimentĂ©es dans les glycosomes. Les raisons et les avantages de la prĂ©sence des enzymes glycolytiques dans l'organite ont Ă©tĂ© largement discutĂ©s, mais la fonctionnalitĂ© et le rĂŽle des voies de biosynthĂšse des sucres nuclĂ©otidiques glycosomales ne sont pas connus. Notre Ă©tude s'est focalisĂ©e sur l'UDP-glucose pyrophosphorylase (UGP), une enzyme impliquĂ©e dans la synthĂšse de l'UDP-glucose (UDP-Glc). Sur la base de la double localisation glycosomale et cytosolique de l'UGP mise en Ă©vidence ici Ă l'aide de plusieurs techniques de localisation subcellulaire, nous avons abordĂ© deux questions en utilisant comme modĂšle les formes procycliques de T. brucei prĂ©sentes dans l'insecte vecteur. La premiĂšre est liĂ©e au mĂ©canisme d'import de l'UGP dans les glycosomes, car cette protĂ©ine ne possĂšde aucun signal d'adressage aux peroxysomes de type PTS1 ou PTS2. Nous avons montrĂ© que l'UGP est importĂ©e dans les glycosomes par "piggybacking" en s'associant Ă la phosphoĂ©nolpyruvate dĂ©carboxylase (PEPCK) possĂ©dant un signal dâadressage PTS1. Les interactions entre l'UGP et la PEPCK ont Ă©tĂ© montrĂ©es in situ et l'identification les rĂ©gions impliquĂ©es dans ces interactions ont Ă©tĂ© identifiĂ©es. Nos rĂ©sultats suggĂšrent que le complexe UGP-PEPCK est formĂ© de maniĂšre transitoire lors de son import dans les glycosomes nouvellement produits et compĂ©tents pour l'import des protĂ©ines. La seconde question concerne le rĂŽle de l'UGP dans les glycosomes. Nous avons montrĂ© que l'UGP est essentielle Ă la croissance des trypanosomes et que les voies mĂ©taboliques glycosomales et cytosoliques dont l'UGP fait partie sont fonctionnelles. En effet, des mutants viables contenant l'UGP exclusivement dans les glycosomes ou dans le cytosol sont viables et produisent des quantitĂ©s similaires d'UDP-Glc. La raison d'ĂȘtre de la production glycosomale d'UDP-Glc par l'UGP reste inconnue, mais n'est probablement pas liĂ©e aux rĂ©actions de glycosylation, Ă©tant donnĂ© qu'aucune glycosyltransfĂ©rase n'a Ă©tĂ© dĂ©tectĂ©e dans l'organite.Un autre aspect de ce travail concerne le rĂŽle des intermĂ©diaires du cycle de l'acide tricarboxylique (TCA) dans le mĂ©tabolisme mitochondrial des formes procycliques. Dans le tractus digestif de son insecte vecteur, les trypanosomes dĂ©pendent de la proline pour alimenter leur mĂ©tabolisme Ă©nergĂ©tique. Cependant, la disponibilitĂ© d'Ă©ventuelles autres sources de carbone pouvant ĂȘtre utilisĂ©es par le parasite est actuellement inconnue. Nous avons montrĂ© que les intermĂ©diaires du cycle TCA, i.e. succinate, malate et a-cĂ©toglutarate, stimulent la croissance des formes procycliques incubĂ©es dans un milieu contenant 2 mM de proline, concentration se situant dans la gamme des quantitĂ©s mesurĂ©es dans l'intestin de la mouche. De plus, le dĂ©veloppement de nouvelles approches ont permis d'Ă©tudier une branche peu explorĂ©e du cycle TCA convertissant le malate en a-cĂ©toglutarate, prĂ©cĂ©demment dĂ©crite comme peu ou pas utilisĂ©e par le parasite, quellles que soient les quantitĂ©s de glucose disponibles. L'activitĂ© de cette branche suggĂšre qu'un cycle TCA complet peut ĂȘtre mis en Ćuvre dans les formes procycliques et probablement dans les autres formes parasitaires de l'insecte. Nos donnĂ©es Ă©largissent le potentiel mĂ©tabolique des trypanosomes et ouvrent la voie vers une meilleure comprĂ©hension du mĂ©tabolisme de ce parasite dans divers organes de la mouche tsĂ©-tsĂ©, oĂč il Ă©volue
Carbohydrate metabolism in trypanosomatids:New insights revealing novel complexity, diversity and species-unique features
The Trypanosome UDP-Glucose Pyrophosphorylase Is Imported by Piggybacking into Glycosomes, Where Unconventional Sugar Nucleotide Synthesis Takes Place
Glycosomes are peroxisome-related organelles of trypanosomatid parasites containing metabolic pathways, such as glycolysis and biosynthesis of sugar nucleotides, usually present in the cytosol of other eukaryotes. UDP-glucose pyrophosphorylase (UGP), the enzyme responsible for the synthesis of the sugar nucleotide UDP-glucose, is localized in the cytosol and glycosomes of the bloodstream and procyclic trypanosomes, despite the absence of any known peroxisome-targeting signal (PTS1 and PTS2). The questions that we address here are (i) is the unusual glycosomal biosynthetic pathway of sugar nucleotides functional and (ii) how is the PTS-free UGP imported into glycosomes? We showed that UGP is imported into glycosomes by piggybacking on the glycosomal PTS1-containing phosphoenolpyruvate carboxykinase (PEPCK) and identified the domains involved in the UGP/PEPCK interaction. Proximity ligation assays revealed that this interaction occurs in 3 to 10% of glycosomes, suggesting that these correspond to organelles competent for protein import. We also showed that UGP is essential for the growth of trypanosomes and that both the glycosomal and cytosolic metabolic pathways involving UGP are functional, since the lethality of the knockdown UGP mutant cell line (RNAi UGP, where RNAi indicates RNA interference) was rescued by expressing a recoded UGP (rUGP) in the organelle (RNAi UGP/ EXP rUGP-GPDH, where GPDH is glycerol-3-phosphate dehydrogenase). Our conclusion was supported by targeted metabolomic analyses (ion chromatography-high-resolution mass spectrometry [IC-HRMS]) showing that UDPglucose is no longer detectable in the RNAi UGP mutant, while it is still produced in cells expressing UGP exclusively in the cytosol (PEPCK null mutant) or glycosomes (RNAi UGP/ EXP rUGP-GPDH). Trypanosomatids are the only known organisms to have selected functional peroxisomal (glycosomal) sugar nucleotide biosynthetic pathways in addition to the canonical cytosolic ones. IMPORTANCE Unusual compartmentalization of metabolic pathways within organelles is one of the most enigmatic features of trypanosomatids. These unicellular eukaryotes are the only organisms that sequestered glycolysis inside peroxisomes (glycosomes), although the selective advantage of this compartmentalization is still no
Gluconeogenesis is essential for trypanosome development in the tsetse fly vector
In the glucose-free environment that is the midgut of the tsetse fly vector, the procyclic form of Trypanosoma brucei primarily uses proline to feed its central carbon and energy metabolism. In these conditions, the parasite needs to produce glucose 6-phosphate (G6P) through gluconeogenesis from metabolism of non-glycolytic carbon source(s). We showed here that two phosphoenolpyruvate-producing enzymes, PEP carboxykinase (PEPCK) and pyruvate phosphate dikinase (PPDK) have a redundant function for the essential gluconeogenesis from proline. Indeed, incorporation of 13C-enriched proline into G6P was abolished in the PEPCK/PPDK null double mutant (Îppdk/Îpepck), but not in the single Îppdk and Îpepck mutant cell lines. The procyclic trypanosome also uses the glycerol conversion pathway to feed gluconeogenesis, since the death of the Îppdk/Îpepck double null mutant in glucose-free conditions is only observed after RNAi-mediated down-regulation of the expression of the glycerol kinase, the first enzyme of the glycerol conversion pathways. Deletion of the gene encoding fructose-1,6-bisphosphatase (Îfbpase), a key gluconeogenic enzyme irreversibly producing fructose 6-phosphate from fructose 1,6-bisphosphate, considerably reduced, but not abolished, incorporation of 13C-enriched proline into G6P. In addition, the Îfbpase cell line is viable in glucose-free conditions, suggesting that an alternative pathway can be used for G6P production in vitro. However, FBPase is essential in vivo, as shown by the incapacity of the Îfbpase null mutant to colonise the fly vector salivary glands, while the parental phenotype is restored in the Îfbpase rescued cell line re-expressing FBPase. The essential role of FBPase for the development of T. brucei in the tsetse was confirmed by taking advantage of an in vitro differentiation assay based on the RNA-binding protein 6 over-expression, in which the procyclic forms differentiate into epimastigote forms but not into mammalian-infective metacyclic parasites. In total, morphology, immunofluorescence and cytometry analyses showed that the differentiation of the epimastigote stages into the metacyclic forms is abolished in the Îfbpase mutant.Voies mĂ©taboliques glycosomales non glycolytiques: nouvelles fonctions pour le dĂ©veloppement et la virulence des trypanosomesMetabolisme de l'acetyl-CoA et de l'acetate chez les trypanosomes: identification de nouvelles voies mĂ©taboliques spĂ©cifiques aux parasitesAlliance française contre les maladies parasitaire