Role of the putative structural protein Sed1p in mitochondrial genome maintenance.
The nuclear gene MIP1 encodes the mitochondrial DNA polymerase responsible for replicating the mitochondrial genome in Saccharomyces cerevisiae. A number of other factors involved in replicating and segregating the mitochondrial genome are yet to be identified. Here, we report that a bacterial two-hybrid screen using the mitochondrial polymerase, Mip1p, as bait identified the yeast protein Sed1p. Sed1p is a cell surface protein highly expressed in the stationary phase. We find that several modified forms of Sed1p are expressed and the largest of these forms interacts with the mitochondrial polymerase in vitro. Deletion of SED1 causes a 3.5-fold increase in the rate of mitochondrial DNA point mutations as well as a 4.3-fold increase in the rate of loss of respiration. In contrast, we see no change in the rate of nuclear point mutations indicating the specific role of Sed1p function in mitochondrial genome stability. Indirect immunofluorescence analysis of Sed1p localization shows that Sed1p is targeted to the mitochondria. Moreover, Sed1p is detected in purified mitochondrial fractions and the localization to the mitochondria of the largest modified form is insensitive to the action of proteinase K. Deletion of the sed1 gene results in a reduction in the quantity of Mip1p and also affects the levels of a mitochondrially-expressed protein, Cox3p. Our results point towards a role for Sed1p in mitochondrial genome maintenance.
Phadnis
Naina
N
Department of Biology, University of Rochester, Rochester, NY 14627-0211, USA.
Ayres Sia
Elaine
E
D001483:Base Sequence; D004272:DNA, Mitochondrial / Q000235:genetics; D005455:Fluorescent Antibody Technique; D016681:Genome, Fungal; D008562:Membrane Glycoproteins / Q000378:metabolism / Q000502:physiology*; D008928:Mitochondria / Q000235:genetics*; D017354:Point Mutation; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism / Q000502:physiology*; D020798:Two-Hybrid System Techniques
D004272:DNA, Mitochondrial; D008562:Membrane Glycoproteins; C077394:SED1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins