A deubiquitinase negatively regulates retro-translocation of nonubiquitinated substrates.
Endoplasmic reticulum (ER) membrane-bound E3 ubiquitin ligases promote ER-associated degradation (ERAD) by ubiquitinating a retro-translocated substrate that reaches the cytosol from the ER, targeting it to the proteasome for destruction. Recent findings implicate ERAD-associated deubiquitinases (DUBs) as positive and negative regulators during ERAD, reflecting the different consequences of deubiquitinating a substrate prior to proteasomal degradation. These observations raise the question of whether a DUB can control the fate of a nonubiquitinated ERAD substrate. In this study, we probed the role of the ERAD-associated DUB, YOD1, during retro-translocation of the nonubiquitinated cholera toxin A1 (CTA1) peptide, a critical intoxication step. Through combining knockdown, overexpression, and binding studies, we demonstrated that YOD1 negatively controls CTA1 retro-translocation, likely by deubiquitinating and inactivating ubiquitinated ERAD components that normally promote toxin retro-translocation. YOD1 also antagonizes the proteasomal degradation of nonglycosylated pro-α factor, a postulated nonubiquitinated yeast ERAD substrate, in mammalian cells. Our findings reveal that a cytosolic DUB exerts a negative function during retro-translocation of nonubiquitinated substrates, potentially by acting on elements of the ERAD machinery.
Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48103 Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, MI 48109.
Kaleena M
KM
Bernardi
Jeffrey M
JM
Williams
Takamasa
T
Inoue
Aric
A
Schultz
Billy
B
Tsai
D002772:Cholera Toxin / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000201:enzymology / Q000235:genetics*; D060746:Endoplasmic Reticulum-Associated Degradation / Q000235:genetics*; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D010455:Peptides / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D021381:Protein Transport; D059748:Proteolysis; D015398:Signal Transduction; D013869:Thiolester Hydrolases / Q000235:genetics / Q000378:metabolism*; D025801:Ubiquitin / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism; D054875:Ubiquitination
D010455:Peptides; D011498:Protein Precursors; D025801:Ubiquitin; D002772:Cholera Toxin; D044767:Ubiquitin-Protein Ligases; C545663:YOD1 protein, human; D013869:Thiolester Hydrolases; D010450:Endopeptidases; D046988:Proteasome Endopeptidase Complex