Functional Characterization of TbMCP5, a Conserved and Essential ADP/ATP Carrier Present in the Mitochondrion of the Human Pathogen Trypanosoma brucei

Peña-Diaz, Priscila; Pélosi, Ludovic; Ebikeme, Charles; Colasante, Claudia; Gao, Fei; Bringaud, Frédéric; Voncken, Frank

doi:10.1074/jbc.m112.404699

Cited by 32 publications

(42 citation statements)

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Section: Discussionsupporting

confidence: 87%

“…Furthermore, TbAAC biochemical properties and ADP/ATP exchange kinetics are similar to those of the yeast AAC2 carrier (26). Similar to the study done by Pena-Diaz and coworkers (26), our in vitro data assaying ADP influx into the mitochondrion for oxidative and substrate phosphorylation support these data, as ADP import into the organelle ceases upon an efficient TbAAC knockdown. Interestingly, when we measured the reverse function of this transporter, mainly the ATP import into the mitochondrion, we observed only a 51% decrease in the ATP-induced ATPase activity, suggesting the existence of another mt import route for ATP.…”

Section: Discussionsupporting

confidence: 87%

“…8.1310 and TbAAC, with the latter being represented by three identical and consecutively arranged genes (Tb927.10.14820, -30, and -40) and showing significant amino acid sequence similarity to functionally characterized AACs from other eukaryotes (25). Importantly, TbAAC exhibits biochemical properties and ADP/ATP exchange kinetics similar to the main yeast carrier, AAC2, while Tb927.8.1310 lacks the canonical sequence features required for ADP/ATP exchange activity (26). Furthermore, RNA interference (RNAi) studies demonstrated that TbAAC is essential for the growth of PF cells and therefore must function as the main ADP/ATP carrier in PF mitochondria (26).…”

mentioning

confidence: 99%

“…Importantly, TbAAC exhibits biochemical properties and ADP/ATP exchange kinetics similar to the main yeast carrier, AAC2, while Tb927.8.1310 lacks the canonical sequence features required for ADP/ATP exchange activity (26). Furthermore, RNA interference (RNAi) studies demonstrated that TbAAC is essential for the growth of PF cells and therefore must function as the main ADP/ATP carrier in PF mitochondria (26).…”

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confidence: 99%

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The ADP/ATP Carrier and Its Relationship to Oxidative Phosphorylation in Ancestral Protist Trypanosoma brucei

et al. 2015

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The highly conserved ADP/ATP carrier (AAC) is a key energetic link between the mitochondrial (mt) and cytosolic compartments of all aerobic eukaryotic cells, as it exchanges the ATP generated inside the organelle for the cytosolic ADP. Trypanosoma brucei, a parasitic protist of medical and veterinary importance, possesses a single functional AAC protein (TbAAC) that is related to the human and yeast ADP/ATP carriers. However, unlike previous studies performed with these model organisms, this study showed that TbAAC is most likely not a stable component of either the respiratory supercomplex III؉IV or the ATP synthasome but rather functions as a physically separate entity in this highly diverged eukaryote. Therefore, TbAAC RNA interference (RNAi) ablation in the insect stage of T. brucei does not impair the activity or arrangement of the respiratory chain complexes. Nevertheless, RNAi silencing of TbAAC caused a severe growth defect that coincides with a significant reduction of mt ATP synthesis by both substrate and oxidative phosphorylation. Furthermore, TbAAC downregulation resulted in a decreased level of cytosolic ATP, a higher mt membrane potential, an elevated amount of reactive oxygen species, and a reduced consumption of oxygen in the mitochondria. Interestingly, while TbAAC has previously been demonstrated to serve as the sole ADP/ATP carrier for ADP influx into the mitochondria, our data suggest that a second carrier for ATP influx may be present and active in the T. brucei mitochondrion. Overall, this study provides more insight into the delicate balance of the functional relationship between TbAAC and the oxidative phosphorylation (OXPHOS) pathway in an early diverged eukaryote. The origination event of an ADP/ATP carrier (AAC) was crucial in the evolution of the present-day mitochondrion that enabled it to become the powerhouse of the eukaryotic cell. Due to the activity of AAC, the energy-producing mitochondria can supply the cytosol with ATP molecules, which fuel most cellular reactions. AAC belongs to the well-defined family of mitochondrial (mt) carrier proteins that are located in the inner mt membrane and are involved in the transport of a wide range of metabolites (1). Under physiological aerobic conditions, AAC is responsible for the 1:1 counterexchange of mt ATP for cytosolic ADP (2). mt ATP is produced mainly by the evolutionarily conserved biochemical pathway of oxidative phosphorylation (OXPHOS), which employs respiratory complexes I through IV to convert the redox energy of various mt substrates into an electrochemical proton gradient (⌬m) across the mt inner membrane. Respiratory complexes I (NADH-ubiquinone oxidoreductase) and II (succinate-ubiquinone oxidoreductase) are responsible for the oxidation of reduced NADH and FADH 2 , respectively. The electrons derived from these biochemical processes continue along the electron transport chain as they are sequentially passed to coenzyme Q, complex III (ubiquinol-cytochrome c oxidoreductase), cytochrome c, complex IV (cytochrome c-O 2 oxid...

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Section: Discussionsupporting

confidence: 87%

Section: Discussionsupporting

confidence: 87%

mentioning

confidence: 99%

mentioning

confidence: 99%

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The ADP/ATP Carrier and Its Relationship to Oxidative Phosphorylation in Ancestral Protist Trypanosoma brucei

et al. 2015

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“…The majority of the ATP produced in PCF T. brucei is derived from its mitochondrion [13], [16]. The produced ATP is exported from the mitochondrion to the cytosol via the ATP/ADP-carrier TbMCP5 [44], [45]. In the cytosol, part of the ATP could be converted to phosphoarginine by the cytoplasmic TbAK3 to serve as a highly mobile cellular energy carrier and local energy buffer.…”

Section: Discussionmentioning

confidence: 99%

The Phosphoarginine Energy-Buffering System of Trypanosoma brucei Involves Multiple Arginine Kinase Isoforms with Different Subcellular Locations

et al. 2013

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Phosphagen energy-buffering systems play an essential role in regulating the cellular energy homeostasis in periods of high-energy demand or energy supply fluctuations. Here we describe the phosphoarginine/arginine kinase system of the kinetoplastid parasite Trypanosoma brucei, consisting of three highly similar arginine kinase isoforms (TbAK1-3). Immunofluorescence microscopy using myc-tagged protein versions revealed that each isoform is located in a specific subcellular compartment: TbAK1 is exclusively found in the flagellum, TbAK2 in the glycosome, and TbAK3 in the cytosol of T. brucei. The flagellar location of TbAK1 is dependent on a 22 amino acid long N-terminal sequence, which is sufficient for targeting a GFP-fusion protein to the trypanosome flagellum. The glycosomal location of TbAK2 is in agreement with the presence of a conserved peroxisomal targeting signal, the C-terminal tripeptide ‘SNL’. TbAK3 lacks any apparent targeting sequences and is accordingly located in the cytosol of the parasite. Northern blot analysis indicated that each TbAK isoform is differentially expressed in bloodstream and procyclic forms of T. brucei, while the total cellular arginine kinase activity was 3-fold higher in bloodstream form trypanosomes. These results suggest a substantial change in the temporal and spatial energy requirements during parasite differentiation. Increased arginine kinase activity improved growth of procyclic form T. brucei during oxidative challenges with hydrogen peroxide. Elimination of the total cellular arginine kinase activity by RNA interference significantly decreased growth (>90%) of procyclic form T. brucei under standard culture conditions and was lethal for this life cycle stage in the presence of hydrogen peroxide. The putative physiological roles of the different TbAK isoforms in T. brucei are further discussed.

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Protein Expression-Yeast

Nielsen

2014

Methods in Enzymology

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Functional Characterization of TbMCP5, a Conserved and Essential ADP/ATP Carrier Present in the Mitochondrion of the Human Pathogen Trypanosoma brucei

Cited by 32 publications

References 56 publications

The ADP/ATP Carrier and Its Relationship to Oxidative Phosphorylation in Ancestral Protist Trypanosoma brucei

The ADP/ATP Carrier and Its Relationship to Oxidative Phosphorylation in Ancestral Protist Trypanosoma brucei

The Phosphoarginine Energy-Buffering System of Trypanosoma brucei Involves Multiple Arginine Kinase Isoforms with Different Subcellular Locations

Protein Expression-Yeast

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