An essential thioredoxin-type protein of Trypanosoma brucei acts as redox-regulated mitochondrial chaperone

Abstract

Most known thioredoxin-type proteins (Trx) participate in redox pathways, using two highly conserved cysteine residues to catalyze thiol-disulfide exchange reactions. Here we demonstrate that the so far unexplored Trx2 from African trypanosomes (Trypanosoma brucei) lacks protein disulfide reductase activity but functions as an effective temperature-activated and redox-regulated chaperone. Immunofluorescence microscopy and fractionated cell lysis revealed that Trx2 is located in the mitochondrion of the parasite. RNA-interference and gene knock-out approaches showed that depletion of Trx2 impairs growth of both mammalian bloodstream and insect stage procyclic parasites. Procyclic cells lacking Trx2 stop proliferation under standard culture conditions at 27°C and are unable to survive prolonged exposure to 37°C, indicating that Trx2 plays a vital role that becomes augmented under heat stress. Moreover, we found that Trx2 contributes to the in vivo infectivity of T. brucei. Remarkably, a Trx2 version, in which all five cysteines were replaced by serine residues, complements for the wildtype protein in conditional knock-out cells and confers parasite infectivity in the mouse model. Characterization of the recombinant protein revealed that Trx2 can coordinate an iron sulfur cluster and is highly sensitive towards spontaneous oxidation. Moreover, we discovered that both wildtype and mutant Trx2 protect other proteins against thermal aggregation and preserve their ability to refold upon return to non-stress conditions. Activation of the chaperone function of Trx2 appears to be triggered by temperature-mediated structural changes and inhibited by oxidative disulfide bond formation. Our studies indicate that Trx2 acts as a novel chaperone in the unique single mitochondrion of T. brucei and reveal a new perspective regarding the physiological function of thioredoxin-type proteins in trypanosomes.

Genome-scale RNA interference profiling of Trypanosoma brucei cell cycle progression defects

“…Thus, we propose a role for the current putative nucleoredoxin in reducing thiols and potentially disrupting disulphide bonds in one or more key cell cycle regulators. This role may be conserved amongst trypanosomatids and indeed, redox signalling may play further roles in cell cycle control, since our screen linked two additional thioredoxin-like proteins to specific and distinct cell cycle defects; TRX2, a redox-regulated mitochondrial chaperone 103 , was linked to accumulation in G 1 , while the putative thioredoxin encoded by Tb927.9.12330 was linked to accumulation in G 2 M (Supplementary data 1 ). Further work will be required to explore how thiol-based redox switch 104 or sensing mechanisms choreograph cell cycle progression in the trypanosomatids.…”

A mitochondrial SET domain protein is essential inT. bruceiand required for mitochondrial morphology and function

“…The growth defect was most pronounced on days 2-5 when BF TbSETD.KD cells had a doubling time of ∼19 h compared to that of parental 90-13 BF cells (∼9 h). After day 6, the growth rate of RNAi cell lines increased, likely because the cells had become refractory to silencing of essential proteins (22, 23). These results indicate that TbSETD is essential in BF parasites and PF parasites grown in low glucose and suggests that TbSETD activity may be less important when PF are grown in high glucose.…”

Transcriptome Analysis of Intracellular Amastigotes of Clinical Leishmania infantum Lines from Therapeutic Failure Patients after Infection of Human Macrophages

“…The reducing substrate for Trx is T(SH) 2 [ 78 ]. Recently, a new Trx has been described in T. brucei (TbTrx2) located in the mitochondria and proposed to be essential for parasite growth in both mammalian and insect stages, as well as relevant for in vivo infectivity [ 79 ]. Furthermore, glutaredoxins (Grxs, formerly called thiol transferases) are ubiquitous small thiol–disulfide proteins that catalyze the reduction of proteins that are thiolated by GSH (glutathionylated, PSSG).…”