A yeast Ubc9 mutant protein with temperature-sensitive in vivo function is subject to conditional proteolysis by a ubiquitin- and proteasome-dependent pathway.
The UBC9 gene of the yeast Saccharomyces cerevisiae is essential for cell viability and encodes a soluble protein of the nucleus that is metabolically stable. Products of mutant alleles selected to confer temperature-sensitive in vivo function were found to be extremely short-lived at the restrictive but long-lived at the permissive condition. An extragenic suppressor mutation was isolated which increased thermoresistance of a ubc9-1 strain. This suppressor turned out to stabilize the mutated gene product, indicating that the physiological activity of ubc9-1 protein is primarily controlled by conditional proteolysis. The labile ubc9-1 protein appears to be a substrate for ubiquitination, and its turnover was substantially reduced by expression of a ubiquitin derivative that interferes with formation of multi-ubiquitin chains. Stabilization resulted also from competitive inhibition of Ubc4-related ubiquitin-conjugating enzymes. Activity of the proteasome complex was crucial to rapid breakdown, whereas vacuolar proteases were dispensable. Thus, the heat-denatured ubc9-1 protein is targeted for proteolysis by the ubiquitin-proteasome pathway and may serve as a useful tool to further define the process by which a misfolded polypeptide is recognized.
Betting
J
J
Institut für Genetik und Mikrobiologie, Universität München, Maria-Ward-Strasse 1a, D-80638 München, Germany.
Seufert
W
W
D000483:Alleles; D064173:Anaphase-Promoting Complex-Cyclosome; D017489:CDC28 Protein Kinase, S cerevisiae / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D017931:DNA Primers / Q000378:metabolism; D004269:DNA, Bacterial / Q000378:metabolism; D008025:Ligases / Q000235:genetics / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D013329:Structure-Activity Relationship; D013489:Suppression, Genetic; D013696:Temperature; D044763:Ubiquitin-Conjugating Enzymes; D043743:Ubiquitin-Protein Ligase Complexes; D044767:Ubiquitin-Protein Ligases; D014452:Ubiquitins / Q000378:metabolism*; D000098843:Ubiquitin-Conjugating Enzyme UBC9
D017931:DNA Primers; D004269:DNA, Bacterial; D009097:Multienzyme Complexes; D014452:Ubiquitins; D044763:Ubiquitin-Conjugating Enzymes; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D044767:Ubiquitin-Protein Ligases; D017489:CDC28 Protein Kinase, S cerevisiae; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D008025:Ligases; D000098843:Ubiquitin-Conjugating Enzyme UBC9