Import of yeast mitochondrial transcription factor (Mtf1p) via a nonconventional pathway.
The yeast mitochondrial (mt) transcription factor Mtf1p is imported into the mitochondria from the cytoplasm without a conventional mt-targeting presequence. To understand its import the mt translocation of wild type and mutant Mtf1p constructs was investigated in vitro under various assay conditions. We report here that Mtf1p, unlike most mt matrix proteins hitherto studied, is translocated into the mitochondria independent of membrane potential, ATP hydrolysis, and membrane receptor. This unusual import of Mtf1p was also observed on ice (3 degrees C). Sub-mitochondrial fractionation demonstrated that Mtf1p was translocated in vitro to one or more of the same mt sites as the endogenous protein that includes the matrix. To identify the mt-targeting sequence of Mtf1p, various N-terminal, C-terminal, or internally deleted Mtf1p derivatives were generated. The full-length and C-terminal deletions but not the N-terminal truncated Mtf1p were imported into mitochondria, indicating the importance of its N-terminal sequence for mt targeting. However, the internal deletion of Mtf1p revealed that the first 150-amino acid N-terminal sequence alone was not sufficient for mt targeting of Mtf1p, suggesting that an extended rather than a short N-terminal sequence is required for import. We favor a model in which Mtf1p adopts an import-competent conformation during translation. Consistent with this model are three findings: most of the protein sequence appears to be required for optimal import, urea denaturation eliminates its import competence, and the import-competent form of the protein is more resistant to tryptic hydrolysis than is the denatured protein. This represents a novel mechanism for mitochondrial protein import.
Biswas
Tapan K
TK
Department of Pathology, University of Chicago, Chicago, Illinois 60637, USA. tbiswas@midway.uchicago.edu
Getz
Godfrey S
GS
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002458:Cell Fractionation; D019286:Endopeptidase K / Q000378:metabolism; D006801:Humans; D008564:Membrane Potentials / Q000502:physiology; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011489:Protein Denaturation; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013696:Temperature; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism
C071720:MTF1 protein, S cerevisiae; D024101:Mitochondrial Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D000255:Adenosine Triphosphate; D013762:Tetrahydrofolate Dehydrogenase; D014357:Trypsin; D019286:Endopeptidase K