Roles of triosephosphate isomerase and aerobic metabolism in Trypanosoma brucei

Abstract

Kinetoplastid protozoa compartmentalize the first seven enzymes of glycolysis and two enzymes of glycerol metabolism in a microbody, the glycosome. While in its mammalian host, Trypanosoma brucei depends entirely on glucose for ATP generation. Under aerobic conditions, most of the glucose is metabolized to pyruvate. Aerobic metabolism depends on the activities of glycosomal triosephosphate isomerase and a mitochondrial glycerophosphate oxidase, and on glycerophosphate<-->dihydroxyacetone phosphate exchange across the glycosomal membrane. Using a combination of genetics and computer modelling, we show that triosephosphate isomerase is probably essential for bloodstream trypanosome survival, but not for the insect-dwelling procyclics, which preferentially use amino acids as an energy source. When the enzyme level decreased to about 15% of that of the wild-type, the growth rate was halved. Below this level, a lethal rise in dihydroxyacetone phosphate was predicted. Expression of cytosolic triosephosphate isomerase inhibited cell growth. Attempts to knockout the trypanosome alternative oxidase genes (which are needed for glycerophosphate oxidase activity) were unsuccessful, but when we lowered the level of the corresponding mRNA by expressing a homologous double-stranded RNA, oxygen consumption was reduced fourfold and the rate of trypanosome growth was halved.

Glucose is toxic to glycosome-deficient trypanosomes

“…Metabolic interference likely contributes to cell death in the bloodstream stage, as it has been previously shown that mislocalization of the glycosomal isozyme of PGK or triose-phosphate isomerase to the cytosol in this stage is toxic (40,41). Furthermore, hexokinase and phosphofructokinase, typically key regulators of glycolysis, are not subject to regulation in trypanosomes (42).…”

Trypanosome Letm1 Protein Is Essential for Mitochondrial Potassium Homeostasis

“…Among the few active pathways in the BS organelle requiring the preservation of mt membrane potential are the maintenance of the glycosomal redox balance via the glycerol-3-phosphatase/dihydroxyacetone phosphate shuttle, fatty acid synthesis, and thymidylate production for DNA synthesis (31,32,(37)(38)(39). Although Ca 2ϩ uptake activity has previously been described in the BS mitochondrion, the essentiality of this pathway was not explored, and we are now therefore able to add ion homeostasis to the list of indispensable functions of the BS organelle.…”

“…Membrane potential is maintained by the remaining F O F 1 -ATP synthase, which hydrolyzes ATP to pump H ϩ out of the matrix (31,32). However, the BS mitochondrion is not dormant because organellar gene expression is still needed for cell viability (33)(34)(35)(36), and a handful of essential mt biochemical pathways have been revealed (31,32,(37)(38)(39).…”

Iron metabolism in trypanosomatids, and its crucial role in infection

“…However, computer modelling of metabolic flux and RNAi-mediated depletion of TAO suggests, in vitro, that the TAO protein would have to be inhibited by more than 95 % for there to be a significant effect on trypanosome growth (Helfert et al 2001). In T. brucei, TAO inhibition is only lethal when coupled with administration of glycerol to block glycerol-phosphate dehydrogenase or with repeated administration of the inhibitor (Yabu et al 1998(Yabu et al , 2003.…”