Servicenavigation

Project Details

Back

Projekt Print View

Genetically encoded biosensors for monitoring redox changes in the trypanothione-based thiol metabolism of trypanosomes

Subject Area Biochemistry
Term from 2014 to 2020
Project identifier Deutsche Forschungsgemeinschaft (DFG) - Project number 251665161
 
Final Report Year 2021

Final Report Abstract

Trypanosomes rely on a unique trypanothione-based system to maintain cellular redox balance. The enzymes involved in the biosynthesis and reduction of trypanothione are located in the cytosol. The thiols and oxidoreductases that maintain the redox homeostasis in the single mitochondrion of the parasites have remained largely unknown. In this work we show that upon exposure to diamide or hydrogen peroxide, bloodstream Trypanosoma brucei undergo protein S-thiolation. Remarkably, not only glutathione but also the dithiol trypanothione is able to form protein-mixed disulfides. A cytosolic glutaredoxin, which contributes to about half of the deglutathionylation capacity of the cells, facilitates reversion of the stress-induced protein modification. S- Glutathionylation of the oxidoreductase itself lowers this activity. Thus, under conditions where protein S-glutathionylation is favored, the deglutathionylation capacity of the cell may be diminished by modifying a glutaredoxin, acting as a thiol redox switch. By fusing to tryparedoxin (Tpx), the central small oxidoreductase of trypanosomes, we generated Tpx-roGFP2, a novel and superior probe for the trypanothione redox couple. Notably, in the context of trypanosomes which harbor both glutathione and trypanothione, also roGFP2-hGrx1 proved to be a sensor for trypanothione. By expressing the sensors in either the cytosol or mitochondrion of procyclic insect stage T. brucei, we provide strong evidence that the mitochondrion harbors a trypanothionebased redox system. Our data indicate that a small portion of cytosolic Tpx resides in the mitochondrion together with another, so-far unknown, oxidoreductase that is able to transfer electrons between trypanothione and the mitochondrial thiol peroxidases. Inhibition of trypanothione biosynthesis by the anti-trypanosomal drug Eflornithine affects the oxidative stress response in the cytosol and mitochondrion demonstrating a close link between the redox systems in the two compartments. Very recently we could show that the mitochondrion of bloodstream T. brucei undergoes a shift to more oxidizing conditions when the cells are cultured at 39°C mimicking a fever situation in the mammalian host. In bloodstream stage parasites, the mitochondrial peroxiredoxin and glutathione peroxidase-type peroxidase are both dispensable whereas in the insect stage, the latter enzyme is essential but the peroxidase activity is not the crucial physiological function.

Publications

 
 

Additional Information

© 2024 DFG Disclaimer / Copyright / Privacy Policy

Textvergrößerung und Kontrastanpassung