A genomic screen identifies Dsk2p and Rad23p as essential components of ER‐associated degradation

Medicherla, Balasubrahmanyam; Kostova, Zlatka; Schaefer, Antje; Wolf, Dieter H.

doi:10.1038/sj.embor.7400164

Cited by 205 publications

(192 citation statements)

References 29 publications

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“…Moreover, CD3␦ is highly ubiquitinated in gp78⌬C49-transfected cells, suggesting an absence of the necessary step to target ubiquitinated CD3␦ for degradation. It is known that multiple pathways operate to degrade unwanted proteins from the ER (45,(47)(48)(49)(50). In agreement with this, a recent study describes the discovery of a novel ER membrane-anchored protein VIMP that interacts with p97/VCP and recruits p97/VCP to Derlin1, 4 Y. Shen, P. Ballar, and S. Fang, unpublished observation.…”

Section: Discussionsupporting

confidence: 54%

AAA ATPase p97/Valosin-containing Protein Interacts with gp78, a Ubiquitin Ligase for Endoplasmic Reticulum-associated Degradation

Zhong

Shen

Ballar

et al. 2004

Journal of Biological Chemistry

194

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Endoplasmic reticulum-associated degradation (ERAD)is a protein quality control mechanism that eliminates unwanted proteins from the endoplasmic reticulum (ER) through a ubiquitin-dependent proteasomal degradation pathway. gp78 is a previously described ER membrane-anchored ubiquitin ligase (E3) involved in ubiquitination of ER proteins. AAA ATPase (ATPase associated with various cellular activities) p97/valosincontaining protein (VCP) subsequently dislodges the ubiquitinated proteins from the ER and chaperones them to the cytosol, where they undergo proteasomal degradation. We now report that gp78 physically interacts with p97/VCP and enhances p97/VCP-polyubiquitin association. The enhanced association correlates with decreases in ER stress-induced accumulation of polyubiquitinated proteins. This effect is abolished when the p97/VCP-interacting domain of gp78 is removed. Further, using ERAD substrate CD3␦, gp78 consistently enhances p97/VCP-CD3␦ binding and facilitates CD3␦ degradation. Moreover, inhibition of endogenous gp78 expression by RNA interference markedly increases the levels of total polyubiquitinated proteins, including CD3␦, and abrogates VCP-CD3␦ interactions. The gp78 mutant with deletion of its p97/VCP-interacting domain fails to increase CD3␦ degradation and leads to accumulation of polyubiquitinated CD3␦, suggesting a failure in delivering ubiquitinated CD3␦ for degradation. These data suggest that gp78-p97/VCP interaction may represent one way of coupling ubiquitination with retrotranslocation and degradation of ERAD substrates.

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Section: Discussionsupporting

confidence: 54%

AAA ATPase p97/Valosin-containing Protein Interacts with gp78, a Ubiquitin Ligase for Endoplasmic Reticulum-associated Degradation

Zhong

Shen

Ballar

et al. 2004

Journal of Biological Chemistry

194

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“…CPY ‡ was fused to GFP, yielding the optically detectable misfolded cytoplasmic protein CPY ‡ -GFP (13,14) (Fig. S1A).…”

Section: Resultsmentioning

confidence: 99%

Cytoplasmic protein quality control degradation mediated by parallel actions of the E3 ubiquitin ligases Ubr1 and San1

Heck

Cheung

Hampton

2009

Proc. Natl. Acad. Sci. U.S.A.

249

383

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Eukaryotic cells maintain proteostasis by quality control (QC) degradation. These pathways can specifically target a wide variety of distinct misfolded proteins, and so are important for management of cellular stress. Although a number of conserved QC pathways have been described in yeast, the E3 ligases responsible for cytoplasmic QC are unknown. We now show that Ubr1 and San1 mediate chaperone-dependent ubiquitination of numerous misfolded cytoplasmic proteins. This action of Ubr1 is distinct from its role in the "N-end rule." In this capacity, Ubr1 functions to protect cells from proteotoxic stresses. Our phenotypic and biochemical studies of Ubr1 and San1 indicate that two strategies are employed for cytoplasmic QC: chaperone-assisted ubiquitination by Ubr1 and chaperone-dependent delivery to nuclear San1. The broad conservation of Ubr ligases and the relevant chaperones indicates that these mechanisms will be important in understanding both basic and biomedical aspects of cellular proteostasis.chaperone | proteostasis | misfolding P rotein quality control (QC) functions to ensure that damaged and misfolded proteins are maintained at acceptable levels to limit their stress-causing, or proteotoxic, effects. One strategy of protein QC is the selective degradation of misfolded proteins. For degradative QC pathways to be effective, they must be specific for aberrant proteins; sufficiently general to recognize selectively common structural hallmarks shared by numerous unrelated proteins; and physiologically important, better allowing the cell to survive proteotoxic stress. Because protein QC underlies many pressing maladies, such as parkinsonism, cystic fibrosis, and aging, discovery of the rules of substrate selectivity and destruction is a key step in understanding these conditions and designing appropriate therapeutical interventions to combat them.In eukaryotes, the ubiquitin proteasome system is employed in the selective degradation of many proteins (1). A substrate protein is marked for degradation by assembly of a polyubiquitin chain, initiated by covalent addition of the small (7.6 kDa) protein ubiquitin to a lysine in an isopeptide bond, followed by iterative addition of the next ubiquitin to the previously added one to create a polyubiquitin chain that is uniquely recognized by the 26S proteasome. Protein ubiquitination is catalyzed by a three-enzyme cascade. The single E1 ubiquitin-activating enzyme hydrolyzes ATP to acquire ubiquitin in labile thioester linkage, which is then transferred in thioester linkage to one of a small group of E2s or ubiquitin-conjugating enzymes (UBCs). E2-bound ubiquitin is finally transferred to an isopeptide linkage on the target protein or the growing polyubiquitin chain by the action of the E3 ubiquitin ligase. It is the E3 ubiquitin ligase that determines the specificity of a given ubiquitination process; identifying and understanding the E3s involved in a degradative pathway are thus key parts of understanding the mechanisms of substrate selection and modification.E3s ...

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“…Dsk2 functions in spindle body duplication 24 and, like hHR23a's ortholog Rad23, has been implicated in ER-associated degradation of certain substrates. 25 Our studies indicate that the UBA domain of hPLIC2 binds to hHR23a's UBL domain, and that its UBL domain binds hHR23a's C-terminal UBA domain. In contrast to these interactions, hPLIC2's UBL domain does not demonstrate binding to the internal UBA domain of hHR23a, even when at 2-fold molar excess and mM protein concentrations.…”

Section: Introductionmentioning

confidence: 98%

Ubiquitin Receptor Proteins hHR23a and hPLIC2 Interact

Yang¹,

Zhang²,

Koepp³

et al. 2007

Journal of Molecular Biology

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SummaryUbiquitin receptor proteins play an important role in delivering ubiquitylated protein substrates to the proteasome for degradation. HHR23a and hPLIC2 are two such ubiquitin receptors that contain ubiquitin-like (UBL) domains, which interact with the proteasome, and ubiquitin-associated (UBA) domains, which interact with ubiquitin. Depending on their abundance UBL/UBA family members can either promote or inhibit the degradation of other proteins, which suggests their participation in the delivery of substrates to the proteasome is highly regulated. In previous work, we determined UBL/UBA domain interactions to promote intramolecular interactions in hHR23a that are abrogated with the addition of either ubiquitin or the proteasome component S5a. In yeast, we determined the hHR23a ortholog (Rad23) to interact with another UBL/UBA family member (Ddi1) and to bind a common tetraubiquitin chain. Here, we use NMR spectroscopy to reveal that hHR23a interacts with hPLIC2 via UBL/UBA domain interactions and to map their binding surfaces. In addition, we demonstrate that these two proteins associate in mammalian cells. Intriguingly, inhibition of the proteasome mitigates hHR23a/hPLIC2 interaction.

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A genomic screen identifies Dsk2p and Rad23p as essential components of ER‐associated degradation

Cited by 205 publications

References 29 publications

AAA ATPase p97/Valosin-containing Protein Interacts with gp78, a Ubiquitin Ligase for Endoplasmic Reticulum-associated Degradation

AAA ATPase p97/Valosin-containing Protein Interacts with gp78, a Ubiquitin Ligase for Endoplasmic Reticulum-associated Degradation

Cytoplasmic protein quality control degradation mediated by parallel actions of the E3 ubiquitin ligases Ubr1 and San1

Ubiquitin Receptor Proteins hHR23a and hPLIC2 Interact

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