A point mutation in the core subunit gene of yeast mitochondrial RNA polymerase is suppressed by a high level of specificity factor MTF1.
The temperature-sensitive yeast mutant pet-ts798 is characterized by an altered mitochondrial transcription apparatus. The mutation has previously been shown to map in the RPO41 gene encoding the core enzyme of mitochondrial RNA polymerase. In the present study the rpo41/pet-ts798 allele was cloned and sequenced, demonstrating that the mutant phenotype is caused by a single amino acid change in a conserved region of the core polymerase. The nuclear gene MTF1, previously isolated as a high copy suppressor of mutant rpo41/pet-ts798, and its gene product were characterized in more detail. Import of a MTF1-COXIV fusion protein in vivo and also import studies with in vitro synthesized MTF1 precursors indicate that MTF1 is a mitochondrial protein and that no apparent cleavage occurs during its import into mitochondria. DNA-binding assays demonstrate that the MTF1 protein alone interacts with DNA in a non-specific manner. An antibody directed against specificity factor MTF1 was raised and used for immunological quantification experiments. The results indicate that suppression is mediated by an increased level of MTF1 protein in mitochondria of the rpo41/pet-ts798 mutant. Possible implications of this finding for the mechanism of suppression are discussed.
Riemen
G
G
Botanisches Institut, Universität Düsseldorf, FRG.
Michaelis
G
G
D000483:Alleles; D000595:Amino Acid Sequence; D001692:Biological Transport; D002451:Cell Compartmentation; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000096:biosynthesis / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D006358:Hot Temperature; D008928:Mitochondria / Q000201:enzymology*; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D017354:Point Mutation / Q000235:genetics; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D013329:Structure-Activity Relationship; D013489:Suppression, Genetic; D014157:Transcription Factors
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C071720:MTF1 protein, S cerevisiae; D024101:Mitochondrial Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D003576:Electron Transport Complex IV; D012321:DNA-Directed RNA Polymerases