Alg13p, the catalytic subunit of the endoplasmic reticulum UDP-GlcNAc glycosyltransferase, is a target for proteasomal degradation.
The second step of dolichol-linked oligosaccharide synthesis in the N-linked glycosylation pathway at the endoplasmic reticulum (ER) membrane is catalyzed by an unusual hetero-oligomeric UDP-N-acetylglucosamine transferase that in most eukaryotes is comprised of at least two subunits, Alg13p and Alg14p. Alg13p is the cytosolic and catalytic subunit that is recruited to the ER by the membrane protein Alg14p. We show that in Saccharomyces cerevisiae, cytosolic Alg13p is very short-lived, whereas membrane-associated Alg13 is relatively stable. Cytosolic Alg13p is a target for proteasomal degradation, and the failure to degrade excess Alg13p leads to glycosylation defects. Alg13p degradation does not require ubiquitin but instead, requires a C-terminal domain whose deletion results in Alg13p stability. Conversely, appending this sequence onto normally long-lived beta-galactosidase causes it to undergo rapid degradation, demonstrating that this C-terminal domain represents a novel and autonomous degradation motif. These data lead to the model that proteasomal degradation of excess unassembled Alg13p is an important quality control mechanism that ensures proper protein complex assembly and correct N-linked glycosylation.
Averbeck
Nicole
N
Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, NY 11794-5215, USA.
Gao
Xiao-Dong
XD
Nishimura
Shin-Ichiro
S
Dean
Neta
N
D000251:Adenosine Triphosphatases; D020134:Catalytic Domain; D016923:Cell Death; D002462:Cell Membrane / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D004721:Endoplasmic Reticulum / Q000201:enzymology*; D004795:Enzyme Stability; D006031:Glycosylation; D009154:Mutation / Q000235:genetics; D017351:N-Acetylglucosaminyltransferases / Q000737:chemistry / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D012097:Repressor Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism
D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; C506082:Alg13 protein, S cerevisiae; C506083:Alg14 protein, S cerevisiae; D017351:N-Acetylglucosaminyltransferases; C023227:UDP-N-acetylglucosamine transferase; D046988:Proteasome Endopeptidase Complex; D000251:Adenosine Triphosphatases; C069670:RPT6 protein, S cerevisiae