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The mitochondrial protein import machinery of parasitic protists

     

    Mitochondria are double membrane-bounded organelles that exert essential metabolic functions such as the synthesis of iron-sulfur clusters. Since most of the mitochondrial proteins are encoded in the nucleus, they have to be imported into one of the four mitochondrial compartments (Fig. 1).

    Fig. 1: Current simplified model of six different mitochondrial protein import pathways. The model is mainly based on numerous studies in yeast, bread mold and mammals but needs to be revised for parasitic protists (Eckers et al. 2012a).

     

    Regardless whether the compartment is the outer membrane (OM), the intermembrane space (IMS), the inner membrane (IM) or the matrix, cytosolic proteins enter the mitochondria of yeast and other opisthokonts via the TOM complex. From there they use different protein import machineries including the TOB/SAM complex, the small Tim proteins, the Mia40/Erv1 system, the TIM23 complex or the components Tim22 and Oxa1. We are interested in various aspects of the mitochondrial protein import of parasitic protists. The following questions are addressed:

     

    - How conserved is the mitochondrial protein import throughout evolution?

     

    - Which of the components are present and isofunctional in parasites?

     

    - Do some parasites have alternative components or transport systems?

     

    - Which residues are essential in isofunctional homologues?

     

    - What are the exact functions of these residues?

     

    Our favourite pet is the unicellular parasite Leishmania tarentolae, which can be easily cultured and manipulated (Fig. 2). Although L. tarentolae is absolutely harmless (unless you are a gecko from Africa), it is closely related to other kinetoplastida, including important human pathogens such as L. donovani (causing kala azar), Trypanosoma brucei (sleeping sickness) and T. cruzi (Chagas disease). One advantage and interesting feature of the L. tarentolae mitochondrion is its strong autofluorescence, which can be used, for example, for co-localization studies (Eckers et al. 2012b). Furthermore, when we established L. tarentolae as a non-opisthokont model organism for mitochondrial import, we were able to demonstrate that the different mitochondrial import signals from Fig. 1 are functionally conserved among eukaryotes despite significant compositional differences of the protein import machineries (see Eckers et al. 2012a).

    Fig. 2: Liquid cultures and agar plates with L. tarentolae insect stages in our lab.

     

    We also work with selected candidate genes of the mitochondrial transport machinery from the human malaria parasite Plasmodium falciparum (reviewed in Deponte et al. 2012 and Deponte 2014). The components form L. tarentolae and P. falciparum are analyzed in vitro and by complementation studies in yeast. For example, we could show that parasite homologues of Erv1/ALR (Fig. 1, Deponte and Hell 2009) are functional sulfhydryl:cytochrome c oxidoreductases that localize to the mitochondrial intermembrane space. We also showed that Erv homologues from kinetoplastid parasites have an altered catalytic mechanism (Eckers et al. 2013). However, it is not the altered catalytic mechanism but the presence of a single cysteine residue that renders L. tarentolae Erv incompatible with Mia40 from yeast (Specht et al. 2018). The physiological role of this partially conserved cysteine residue and the identity of the protein that fulfills the functions of Mia40 in parasitic protists still remain unknown.

     

    The project is currently funded by the DFG(grant DE 1431/10). Previous funding was provided by the DFG (grant DE 1431/2), the Friedrich-Baur-Stiftung, and the Bavaria California Technology Center.

     

Further projects

Mitochondria

GFP-tagging sheds light on protein translocation

     

Thiol-dependent redox catalysis

The glyoxalase system of the malaria parasite Plasmodium falciparum

Hydroperoxide and thiol-disulfide metabolism - new enzymes, new lessons

Quantitative and mechanistic assessment of thiol switches