To study the degradation requirements of unassembled immunoglobulin (Ig) chains, we heterologously expressed a cDNA encoding the secretory form of murine mu in the yeast S. cerevisiae. We found that mu chains were translocated into and retained in the endoplasmic reticulum (ER) as they were N-glycosylated and bound to the yeast homolog of BiP, Kar2p. Similar to mutant yeast carboxypeptidase Y (CPY*), known to undergo cytosolic degradation, mu protein is stabilized in yeast mutants lacking the ubiquitinating enzymes Ubc6p and Ubc7p or in cells overexpressing mutant ubiquitin. Unexpectedly, the translation inhibitor cycloheximide (CHX), but not puromycin, led to the accumulation of polyubiquitinated mu chains that were still glycosylated. By contrast, degradation of CPY* was not impaired by CHX, indicating that the drug affects a substrate-specific degradation step. In contrast to the situation for CPY*, the ER-transmembrane protein Der1p is not essential for mu degradation. Strikingly, however, the CHX-induced accumulation of polyubiquitinated Igmu chains was stronger in deltader1-mutants as compared to wild-type cells, indicating an additive effect of two inhibitory conditions. The results support a previously unknown activity of CHX, i.e. impairing the degradation of transport-incompetent secretory mu chains. Moreover, this activity will allow to dissect substrate-specific steps in ER associated protein degradation.
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Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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