title: N-Terminal Presequence-Independent Import of Phosphofructokinase into Hydrogenosomes of Trichomonas vaginalis.
abstract: Mitochondrial evolution entailed the origin of protein import machinery that allows nuclear-encoded proteins to be targeted to the organelle, as well as the origin of cleavable N-terminal targeting sequences (NTS) that allow efficient sorting and import of matrix proteins. In hydrogenosomes and mitosomes, reduced forms of mitochondria with reduced proteomes, NTS-independent targeting of matrix proteins is known. Here, we studied the cellular localization of two glycolytic enzymes in the anaerobic pathogen Trichomonas vaginalis: PPi-dependent phosphofructokinase (TvPPi-PFK), which is the main glycolytic PFK activity of the protist, and ATP-dependent PFK (TvATP-PFK), the function of which is less clear. TvPPi-PFK was detected predominantly in the cytosol, as expected, while all four TvATP-PFK paralogues were imported into T. vaginalis hydrogenosomes, although none of them possesses an NTS. The heterologous expression of TvATP-PFK in Saccharomyces cerevisiae revealed an intrinsic capability of the protein to be recognized and imported into yeast mitochondria, whereas yeast ATP-PFK resides in the cytosol. TvATP-PFK consists of only a catalytic domain, similarly to "short" bacterial enzymes, while ScATP-PFK includes an N-terminal extension, a catalytic domain, and a C-terminal regulatory domain. Expression of the catalytic domain of ScATP-PFK and short Escherichia coli ATP-PFK in T. vaginalis resulted in their partial delivery to hydrogenosomes. These results indicate that TvATP-PFK and the homologous ATP-PFKs possess internal structural targeting information that is recognized by the hydrogenosomal import machinery. From an evolutionary perspective, the predisposition of ancient ATP-PFK to be recognized and imported into hydrogenosomes might be a relict from the early phases of organelle evolution.
authors.affiliation: Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Rada
authors.affiliation: Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic.
authors.forename: Abhijith Radhakrishna
authors.initials: AR
authors.lastname: Makki
authors.affiliation: Institute for Molecular Evolution, Heinrich-Heine-University Düsseldorf, Düsseldorf, Germany.
authors.forename: Verena
authors.initials: V
authors.lastname: Zimorski
authors.affiliation: Institute for Molecular Evolution, Heinrich-Heine-University Düsseldorf, Düsseldorf, Germany.
authors.forename: Sriram
authors.initials: S
authors.lastname: Garg
authors.affiliation: Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic.
authors.forename: Vladimír
authors.initials: V
authors.lastname: Hampl
authors.affiliation: Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic.
authors.forename: Ivan
authors.initials: I
authors.lastname: Hrdý
authors.affiliation: Institute for Molecular Evolution, Heinrich-Heine-University Düsseldorf, Düsseldorf, Germany.
authors.forename: Sven B
authors.initials: SB
authors.lastname: Gould
authors.affiliation: Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic tachezy@natur.cuni.cz.
authors.forename: Jan
authors.initials: J
authors.lastname: Tachezy
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000595:Amino Acid Sequence; D011756:Diphosphates / Q000378:metabolism; D005288:Ferredoxins / Q000378:metabolism; D006859:Hydrogen / Q000378:metabolism*; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D015388:Organelles / Q000187:drug effects / Q000378:metabolism*; D025221:Phosphofructokinases / Q000737:chemistry* / Q000378:metabolism*; D010802:Phylogeny; D011401:Promoter Regions, Genetic / Q000235:genetics; D021381:Protein Transport / Q000187:drug effects; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000378:metabolism; D016415:Sequence Alignment; D014246:Trichomonas vaginalis / Q000187:drug effects / Q000201:enzymology*
chemical_list: D011756:Diphosphates; D005288:Ferredoxins; D006859:Hydrogen; D000255:Adenosine Triphosphate; D025221:Phosphofructokinases