Genetically encoded biosensors for monitoring redox changes in the trypanothione-based thiol metabolism of trypanosomes
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
- (2015) Detection of thiol-based switch processes in parasites – facts and future. Biol. Chem. 396, 445-463
Rahbari, M., Diederich, K., Becker, K., Krauth-Siegel, R. L. and Jortzik, E
(See online at https://doi.org/10.1515/hsz-2014-0279) - (2015) Glutaredoxin-deficiency confers bloodstream Trypanosoma brucei with improved thermotolerance. Mol. Biochem. Parasitol. 204, 93-105
Musunda, B., Benítez, D., Dirdjaja, N., Comini, M. A. and Krauth-Siegel, R. L.
(See online at https://doi.org/10.1016/j.molbiopara.2016.02.001) - (2016) The cytosolic or the mitochondrial glutathione peroxidase-type tryparedoxin peroxidase is sufficient to protect procyclic Trypanosoma brucei from iron-mediated mitochondrial damage and lysis. Mol. Microbiol. 99, 172-187
Schaffroth, C., Bogacz, M., Dirdjaja, N., Nissen, A. and Krauth-Siegel, R. L.
(See online at https://doi.org/10.1111/mmi.13223) - (2016) Thiol redox biology of trypanosomatids and potential targets for chemotherapy. Mol. Biochem. Parasitol. 206, 67-74
Leroux, A. and Krauth-Siegel, R. L.
(See online at https://doi.org/10.1016/j.molbiopara.2015.11.003) - (2017) Stress-induced protein S-glutathionylation and S- trypanothiolation in African trypanosomes – a quantitative redox proteome and thiol analysis. Antiox. Redox Sign. 27, 517-533
Ulrich, K., Finkenzeller, C., Merker, S., Rojas, F., Matthews, K., Ruppert, T. and Krauth-Siegel, R. L.
(See online at https://doi.org/10.1089/ars.2016.6947) - (2018) A glutaredoxin in the mitochondrial intermembrane space has stage-specific functions in the thermotolerance and proliferation of African trypanosomes. Redox Biology 15, 532-547
Ebersoll, S., Musunda, B., Schmenger, T., Dirdjaja, N., Bonilla, M., Manta, B., Ulrich, K., Comini, M. A., and Krauth-Siegel, R. L.
(See online at https://doi.org/10.1016/j.redox.2018.01.011) - (2019) An essential thioredoxin-type protein of Trypanosoma brucei acts as redox-regulated mitochondrial chaperone. PLoS Pathogens, e1008065
Currier, R.B., Ulrich, K., Leroux, A., Dirdjaja, N., Deambrosi, M., Bonilla, M., Ahmed, Y.L., Adrian, L., Antelmann, H., Jakob, U., Comini, M.A, and Krauth-Siegel, R.L.
(See online at https://doi.org/10.1371/journal.ppat.1008065) - (2020) A tryparedoxin-coupled biosensor reveals a mitochondrial trypanothione metabolism in trypanosomes. eLife: e53227
Ebersoll, S., Bogacz, M., Günter, L.M., Dick, T. P., and Krauth-Siegel, R.L.
(See online at https://doi.org/10.7554/elife.53227) - (2020) The mitochondrial peroxiredoxin displays distinct roles in different developmental stages of African trypanosomes. Redox Biology 34, 101547
Bogacz, M., Dirdjaja, N., Wimmer, B., Habich, C., and Krauth-Siegel, R.L.
(See online at https://doi.org/10.1016/j.redox.2020.101547)
