Competition and protease sensitivity assays provide evidence for the existence of a hydrogenosomal protein import machinery in Trichomonas vaginalis.
Hydrogenosomes are double membrane bounded redox organelles found in a number of amitochondriate protists and fungi. They are involved in carbohydrate metabolism and ATP synthesis and thus resemble mitochondria. Molecular analysis of the hydrogenosomal heat shock proteins Hsp70, Hsp60 and Hsp10 in Trichomonas vaginalis, one of the deepest-branching eukaryotes known to date, has revealed that these group exclusively with mitochondrial heat shock proteins. This finding indicates strongly that a progenitor organelle which gave rise to contemporary mitochondria and hydrogenosomes existed early in eukaryotic life. This hypothesis is further supported by similarities of hydrogenosomal and mitochondrial biogenesis. It was shown that T. vaginalis hydrogenosomal proteins are synthesized on free ribosomes in the cytosol with an N-terminal presequence that carries targeting information and is cleaved upon import into the organelle. Furthermore, as in mitochondrial import, hydrogenosomal protein import requires ATP, an electrochemical transmembrane potential and cytosolic protein factor(s). Here we demonstrate that inhibition of hydrogenosomal protein import occurs (i) in the presence of a synthetic presequence peptide and (ii) after pretreatment of hydrogenosomes with the protease trypsin. Trypsin pretreatment affects two hydrogenosomal membrane proteins of 31 and 70 kDa, respectively. Thus, we present evidence that import is saturable and that proteinaceous hydrogenosomal import receptor(s) exist. These results are a first step towards a characterization of the hydrogenosomal import machinery which should provide further insights into the relationship of hydrogenosomes and mitochondria and the evolution of protein targeting into organelles of endosymbiotic origin.
Plümper
E
E
Department of Microbiology and Immunology, University of California, Los Angeles 90095-1489, USA.
Bradley
P J
PJ
Johnson
P J
PJ
D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005288:Ferredoxins / Q000378:metabolism; D017125:Genes, Protozoan; D008565:Membrane Proteins / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D015388:Organelles / Q000187:drug effects / Q000378:metabolism; D011498:Protein Precursors / Q000378:metabolism; D015800:Protozoan Proteins / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors; D014246:Trichomonas vaginalis / Q000378:metabolism*; D014357:Trypsin / Q000494:pharmacology
D005288:Ferredoxins; D008565:Membrane Proteins; D011498:Protein Precursors; D015800:Protozoan Proteins; D014357:Trypsin