Abstract
Most mitochondrial proteins are synthesized in the cytosol, imported into mitochondria via the TOM40 (translocase of the mitochondrial outer membrane 40) complex, and follow several distinct sorting pathways to reach their destination submitochondrial compartments. Phosphate carrier (PiC) is an inner membrane protein with 6 transmembrane segments (TM1-TM6) and requires, after translocation across the outer membrane, the Tim9-Tim10 complex and the TIM22 complex to be inserted into the inner membrane. Here we analyzed an in vitro import of fusion proteins between various PiC segments and mouse dihydrofolate reductase. The fusion protein without TM1 and TM2 was translocated across the outer membrane but was not inserted into the inner membrane. The fusion proteins without TM1-TM4 were not inserted into the inner membrane but instead translocated across the inner membrane. Functional defects of Tim50 of the TIM23 complex caused either by depletion of the protein or the addition of anti-Tim50 antibodies blocked translocation of the fusion proteins without TM1-TM4 across the inner membrane, suggesting that lack of TM1-TM4 led to switch of its sorting pathway from the TIM22 pathway to the TIM23 pathway. PiC thus appears to have a latent signal for sorting to the TIM23 pathway, which is exposed by reduced interactions with the Tim9-Tim10 complex and maintenance of the import competence.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Motifs
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Animals
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Biological Transport
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Cell Membrane / metabolism
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Chromatography, High Pressure Liquid
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Cytosol / metabolism
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Dose-Response Relationship, Drug
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Electrophoresis, Polyacrylamide Gel
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Fungal Proteins / chemistry*
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Fungal Proteins / metabolism
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Membrane Potentials
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Membrane Proteins / metabolism
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Membrane Transport Proteins / chemistry*
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Membrane Transport Proteins / metabolism
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Mice
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Mitochondria / metabolism*
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Mitochondrial Membrane Transport Proteins / metabolism
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Mitochondrial Precursor Protein Import Complex Proteins
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Mitochondrial Proteins / metabolism
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Models, Biological
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Phosphate Transport Proteins / metabolism
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Phosphate Transport Proteins / physiology*
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Phosphates / chemistry*
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Plasmids / metabolism
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Promoter Regions, Genetic
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Protein Structure, Tertiary
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Protein Transport
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / metabolism
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Saccharomyces cerevisiae / metabolism
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Saccharomyces cerevisiae Proteins / chemistry*
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Saccharomyces cerevisiae Proteins / metabolism
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Signal Transduction
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Tetrahydrofolate Dehydrogenase / metabolism
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Time Factors
Substances
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Fungal Proteins
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MIR1 protein, S cerevisiae
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Membrane Proteins
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Membrane Transport Proteins
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Mitochondrial Membrane Transport Proteins
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Mitochondrial Precursor Protein Import Complex Proteins
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Mitochondrial Proteins
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Phosphate Transport Proteins
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Phosphates
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Recombinant Fusion Proteins
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Saccharomyces cerevisiae Proteins
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TIM10 protein, S cerevisiae
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TIM22 protein, S cerevisiae
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TIM23 protein, S cerevisiae
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TIM50 protein, S cerevisiae
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Tim9 protein, S cerevisiae
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Tetrahydrofolate Dehydrogenase