Targeting capacity and conservation of PreP homologues localization in mitochondria of different species |
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Authors: | Alikhani Nyosha Berglund Anna-Karin Engmann Tanja Spånning Erika Vögtle F-Nora Pavlov Pavel Meisinger Chris Langer Thomas Glaser Elzbieta |
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Institution: | 1 Department of Biochemistry and Biophysics, Arrhenius Laboratories for Natural Science, Stockholm University, SE-10691 Stockholm, Sweden2 Institute for Genetics, Center for Molecular Medicine, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, 50674 Cologne, Germany3 Institut für Biochemie und Molekularbiologie, ZBMZ, Universität Freiburg, 79104 Freiburg, Germany4 Fakultät für Biologie, Universität Freiburg, Freiburg, Germany5 BIOSS, Center for Biological Signalling Studies, Universität Freiburg, Freiburg, Germany6 Max Planck Institute for Biology of Aging, 50931 Cologne, Germany |
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Abstract: | Mitochondrial presequences and other unstructured peptides are degraded inside mitochondria by presequence proteases (PrePs) identified in Arabidopsis thaliana (AtPreP), humans (hPreP), and yeast (Cym1/Mop112). The presequences of A. thaliana and human PreP are predicted to consist of 85 and 29 amino acids, respectively, whereas the Saccharomyces cerevisiae Cym1/Mop112 presequence contains only 7 residues. These differences may explain the reported targeting of homologous proteins to different mitochondrial subcompartments. Here we have investigated the targeting capacity of the PreP homologues' presequences. We have produced fusion constructs containing N-terminal portions of AtPreP(1-125), hPreP(1-69), and Cym1(1-40) coupled to green fluorescent protein (GFP) and studied their import into isolated plant, mammalian, and yeast mitochondria, followed by mitochondrial subfractionation. Whereas the AtPreP presequence has the capacity to target GFP into the mitochondrial matrix of all three species, the hPreP presequence only targets GFP to the matrix of mammalian and yeast mitochondria. The Cym1/Mop112 presequence has an overall much weaker targeting capacity and only ensures mitochondrial sorting in its host species yeast. Revisiting the submitochondrial localization of Cym1 revealed that endogenous Cym1/Mop112 is localized to the matrix space, as has been previously reported for the plant and human homologues. Moreover, complementation studies in yeast show that native AtPreP restores the growth phenotype of yeast cells lacking Cym1, demonstrating functional conservation. |
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Keywords: | PreP presequence protease GFP green fluorescent protein MPP mitochondrial processing peptidase PK proteinase K o-ph ortho-phenanthroline EDTA ethylenediaminetetraacetic acid Mops 4-morpholinepropanesulfonic acid BSA bovine serum albumin WT wild type |
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