title: Endoplasmic Reticulum-associated Degradation of Pca1p, a Polytopic Protein, via Interaction with the Proteasome at the Membrane.
abstract: Endoplasmic reticulum-associated degradation (ERAD) plays a critical role in the destruction of terminally misfolded proteins at the secretory pathway. The system also regulates expression levels of several proteins such as Pca1p, a cadmium exporter in yeast. To gain better insight into the mechanisms underlying ERAD of Pca1p and other polytopic proteins by the proteasome in the cytosol, our study determined the roles for the molecular factors of ERAD in dislodging Pca1p from the endoplasmic reticulum (ER). Inactivation of the 20S proteasome leads to accumulation of ubiquitinated Pca1p in the ER membrane, suggesting a role for the proteasome in extraction of Pca1p from the ER. Pca1p formed a complex with the proteasome at the membrane in a Doa10p E3 ligase-dependent manner. Cdc48p is required for recruiting the proteasome to Pca1p. Although the Ufd2p E4 ubiquitin chain extension enzyme is involved in efficient degradation of Pca1p, Ufd2p-deficient cells did not affect the formation of a complex between Pca1p and the proteasome. Two other polytopic membrane proteins undergoing ERAD, Ste6*p and Hmg2p, also displayed the same outcomes observed for Pca1p. However, poly-ubiquitinated Cpy1*p, a luminal ERAD substrate, was detected in the cytosol independent of proteolytic activities of the proteasome. These results indicate that extraction and degradation of polytopic membrane proteins at the ER is a coupled event. This mechanism would relieve the cost of exposed hydrophobic domains in the cytosol during ERAD.
authors.affiliation: From the Department of Biochemistry and Redox Biology Center, University of Nebraska, Lincoln, Nebraska, 68588-0664 and.
authors.forename: Nathan
authors.initials: N
authors.lastname: Smith
authors.affiliation: From the Department of Biochemistry and Redox Biology Center, University of Nebraska, Lincoln, Nebraska, 68588-0664 and.
authors.forename: David J
authors.initials: DJ
authors.lastname: Adle
authors.affiliation: From the Department of Biochemistry and Redox Biology Center, University of Nebraska, Lincoln, Nebraska, 68588-0664 and.
authors.forename: Miaoyun
authors.initials: M
authors.lastname: Zhao
authors.affiliation: From the Department of Biochemistry and Redox Biology Center, University of Nebraska, Lincoln, Nebraska, 68588-0664 and the College of Life and Environmental Sciences, Shanghai Normal University, Shanghai, China 200234.
authors.forename: Xiaojuan
authors.initials: X
authors.lastname: Qin
authors.affiliation: From the Department of Biochemistry and Redox Biology Center, University of Nebraska, Lincoln, Nebraska, 68588-0664 and.
authors.forename: Heejeong
authors.initials: H
authors.lastname: Kim
authors.affiliation: From the Department of Biochemistry and Redox Biology Center, University of Nebraska, Lincoln, Nebraska, 68588-0664 and jlee7@unl.edu.
authors.forename: Jaekwon
authors.initials: J
authors.lastname: Lee
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D027682:Cation Transport Proteins / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D060746:Endoplasmic Reticulum-Associated Degradation; D006903:Hydroxymethylglutaryl CoA Reductases / Q000235:genetics / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D061988:Proteasome Inhibitors / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D054875:Ubiquitination; D000074405:Valosin Containing Protein
chemical_list: D018528:ATP-Binding Cassette Transporters; D027682:Cation Transport Proteins; D018797:Cell Cycle Proteins; D007976:Leupeptins; C093603:PCA1 protein, S cerevisiae; D061988:Proteasome Inhibitors; D011993:Recombinant Fusion Proteins; C489277:STE6 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C541571:HMG2 protein, S cerevisiae; D006903:Hydroxymethylglutaryl CoA Reductases; D044763:Ubiquitin-Conjugating Enzymes; D044767:Ubiquitin-Protein Ligases; D046988:Proteasome Endopeptidase Complex; D000251:Adenosine Triphosphatases; C000614177:CDC48 protein, S cerevisiae; D000074405:Valosin Containing Protein; C114683:UFD2 protein, S cerevisiae; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde
keywords: ATPase; cadmium transporter; degron; endoplasmic reticulum-associated protein degradation (ERAD); membrane protein; metal homeostasis; proteasome; ubiquitin ligase; yeast