In vivo disassembly of free polyubiquitin chains by yeast Ubp14 modulates rates of protein degradation by the proteasome.
Degradation of many eukaryotic proteins requires their prior ligation to polyubiquitin chains, which target substrates to the 26S proteasome, an abundant cellular protease. We describe a yeast deubiquitinating enzyme, Ubp14, that specifically disassembles unanchored ('free') ubiquitin chains in vitro, a specificity shared by mammalian isopeptidase T. Correspondingly, deletion of the UBP14 gene from yeast cells results in a striking accumulation of free ubiquitin chains, which correlates with defects in ubiquitin-dependent proteolysis. Increasing the steady-state levels of ubiquitin chains in wild-type cells (by expressing a derivative of ubiquitin with an altered C-terminus) inhibits protein degradation to a degree comparable with that observed in ubp14delta cells. Inhibition of degradation is also seen when an active site mutant of Ubp14 is overproduced in vivo. Surprisingly, overproduction of wild-type Ubp14 can inhibit degradation of some proteins as well. Finally, Ubp14 and human isopeptidase T are shown to be functional homologs by complementation analysis. We propose that Ubp14 and isopeptidase T facilitate proteolysis in vivo by preventing unanchored ubiquitin chains from competitively inhibiting polyubiquitin-substrate binding to the 26S proteasome.
Amerik AYu
University of Chicago, Department of Biochemistry and Molecular Biology, IL 60637, USA.
Swaminathan
S
S
Krantz
B A
BA
Wilkinson
K D
KD
Hochstrasser
M
M
D000595:Amino Acid Sequence; D001665:Binding Sites; D019759:Carbon-Nitrogen Lyases; D010450:Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D006801:Humans; D015151:Immunoblotting; D008190:Lyases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010447:Peptide Hydrolases / Q000737:chemistry / Q000378:metabolism*; D010641:Phenotype; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D013379:Substrate Specificity; D014452:Ubiquitins / Q000737:chemistry / Q000378:metabolism*
D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; D014452:Ubiquitins; D010450:Endopeptidases; D010447:Peptide Hydrolases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; C108364:UBP14 protein, S cerevisiae; D008190:Lyases; D019759:Carbon-Nitrogen Lyases; C034300:isopeptidase