Differential ubiquitin-dependent degradation of the yeast apo-cytochrome c isozymes.
The yeast Saccharomyces cerevisiae contains two forms of cytochrome c, iso-1- and iso-2-cytochrome c, which are encoded by the nuclear genes CYC1 and CYC7, respectively. The cytochromes c are synthesized in the cytosol, imported into mitochondria, and subsequently modified by the covalent attachment of heme through the action of cytochrome c heme lyase, which is encoded by CYC3. Apo-iso-2-cytochrome c but not apo-iso-1-cytochrome c was observed in cyc3(-) mutants. Furthermore, pulse-chase experiments previously demonstrated that the lack of apo-iso-1-cytochrome c was due to its rapid degradation. We report herein that this degradation of apo-iso-1-cytochrome c is dependent on ubiquitination and on the action of the proteasome. Diminished degradation of apo-iso-1-cytochrome c was observed in pre2-2 and pre1-1 mutants having altered proteasome subunits; in ubc1, ubc4, and ubc5 strains lacking one or more of the ubiquitin-conjugating enzymes; and in strains blocked in multi-ubiquitination by overproduction of the abnormal ubiquitin-K48R ubiquitin. In addition, we have used epitope-tagged ubiquitin to demonstrate that apo-iso-1-cytochrome c but not apo-iso-2-cytochrome c is ubiquitinated. Furthermore, the degradation of apo-iso-1-cytochrome c was diminished when the N-terminal region was replaced with the N-terminal region of apo-iso-2-cytochrome c, indicating that this region may be the target for degradation. We suggest that ubiquitin-dependent degradation of apo-iso-1-cytochrome c is part of the regulatory process controlling the preferential expression of the iso-cytochromes c.
Pearce
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DA
Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA.
Sherman
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F
D000595:Amino Acid Sequence; D001059:Apoproteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism; D003574:Cytochrome c Group / Q000235:genetics / Q000378:metabolism*; D045304:Cytochromes c; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D046988:Proteasome Endopeptidase Complex; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D014452:Ubiquitins / Q000378:metabolism*
D001059:Apoproteins; D003574:Cytochrome c Group; D007527:Isoenzymes; D009097:Multienzyme Complexes; D014452:Ubiquitins; D045304:Cytochromes c; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex