The Doa4 Deubiquitinating Enzyme Is Functionally Linked to the Vacuolar Protein-sorting and Endocytic Pathways

Amerik, Alexander Y.; Nowak, Jonathan A.; Swaminathan, Sowmya; Hochstrasser, Mark

doi:10.1091/mbc.11.10.3365

Cited by 280 publications

(281 citation statements)

References 66 publications

Supporting

Mentioning

257

Contrasting

Order By: Relevance

“…5D). This is consistent with previous findings in yeast that deubiquitination by Doa4 is not essential for cargo sorting into vacuoles (Amerik et al, 2000). Instead, in Doa4 mutants, intracellular free ubiquitins are depleted by the insufficiency of Doa4-function, which cleaves ubiquitin chains to produce free ubiquitins (Swaminathan et al, 1999).…”

Section: Discussionsupporting

confidence: 93%

“…Indeed, there is no evidence from the database that yeast possesses an AMSH orthologue. Instead, yeast has a DUB called Doa4 that functions in MVB sorting proximal to the last steps (Swaminathan et al, 1999;Amerik et al, 2000;Katzmann et al, 2001;Babst et al, 2002;Luhtala and Odorizzi, 2004). Thus, it has been unclear whether a DUB is involved in MVB sorting in mammals.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

AMSH, an ESCRT-III Associated Enzyme, Deubiquitinates Cargo on MVB/Late Endosomes

Kyuuma

Kikuchi

Kojima

et al. 2006

Cell Struct. Funct.

View full text Add to dashboard Cite

and 6 These two authors contributed equally to this work.ABSTRACT. The appropriate sorting of vesicular cargo, including cell-surface proteins, is critical for many cellular functions. Ubiquitinated cargo is targeted to endosomes and digested by lysosomal enzymes. We previously identified AMSH, a deubiquitination enzyme (DUB), to be involved in vesicular transport. Here, we purified an AMSH-binding protein, CHMP3, which is an ESCRT-III subunit. ESCRT-III functions on maturing endosomes, indicating AMSH might also play a role in MVB/late endosomes. Expression of an AMSH mutant lacking CHMP3-binding ability resulted in aberrant endosomes with accumulations of ubiquitinated cargo. Nevertheless, CHMP3-binding capability was not essential for AMSH's in vitro DUB activity or its endosomal localization, suggesting that, in vivo, the deubiquitination of endosomal cargo is CHMP3-dependent. Ubiquitinated cargo also accumulated on endosomes when catalytically inactive AMSH was expressed or AMSH was depleted. These results suggest that both the DUB activity of AMSH and its CHMP3-binding ability are required to clear ubiquitinated cargo from endosomes.

show abstract

Section: Discussionsupporting

confidence: 93%

Section: Introductionmentioning

confidence: 99%

AMSH, an ESCRT-III Associated Enzyme, Deubiquitinates Cargo on MVB/Late Endosomes

Kyuuma

Kikuchi

Kojima

et al. 2006

Cell Struct. Funct.

View full text Add to dashboard Cite

show abstract

“…In yeast, the deubiquitinating enzyme Doa4 is associated with the 26 S proteasome (45,46). Yeast cells lacking Doa4 are significantly depleted of ubiquitin, and genetic evidence shows that the proteasome is at least partially responsible for the degradation of ubiquitin (45,46).…”

Section: Discussionmentioning

confidence: 99%

Concurrent Translocation of Multiple Polypeptide Chains through the Proteasomal Degradation Channel

Lee

Prakash

Matouschek

2002

Journal of Biological Chemistry

View full text Add to dashboard Cite

The proteasome can actively unfold proteins by sequentially unraveling their substrates from the attachment point of the degradation signal. To investigate the steric constraints imposed on substrate proteins during their degradation by the proteasome, we constructed a model protein in which specific parts of the polypeptide chain were covalently connected through disulfide bridges. The cross-linked model proteins were fully degraded by the proteasome, but two or more cross-links retarded the degradation slightly. These results suggest that the pore of the proteasome allows the concurrent passage of at least three stretches of a polypeptide chain. A degradation channel that can tolerate some steric bulk may reconcile the two opposing needs for degradation that is compartmentalized to avoid aberrant proteolysis yet able to handle a range of substrates of various sizes.Protein degradation is a critical part of cellular regulation (1). In eukaryotic cells, a multicomponent protease called the proteasome is responsible for the turnover of short-lived regulatory proteins, the removal of abnormal polypeptides, and the production of peptides for antigen presentation (2). Degradation of short-lived regulatory proteins is essential for a wide range of cellular functions including cell cycle control and signal transduction (3). Failure to degrade aggregates of misfolded proteins can lead to disease such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and Huntington's disease (4). Degradation by the proteasome usually involves two consecutive steps: targeting of the substrate for degradation by the attachment of polyubiquitin chains and degradation of the tagged protein by the proteasome with concomitant release of ubiquitin (5).The active sites of proteolysis of the proteasome by themselves show little substrate specificity. Specificity of degradation is achieved by sequestering the proteolytic sites within the structure and tightly controlling access. The three-dimensional structure of the proteasome has been determined by x-ray crystallography and electron microscopy (6 -11). The proteasome consists of a central proteolytic core particle with regulatory caps at either end of it. The core particle is made of two copies of seven different ␣-subunits and seven different ␤-subunits. The subunits are arranged in four heptameric rings, which are stacked on top of each other to form a cylindrical particle. The rings form a central ␤-chamber and two ␣-chambers, one at each end of the particle (2). The proteolytic sites are located in the ␤-chamber and are accessible only through a central channel that runs along the long axis of the particle. The channel has narrow constrictions at the entrance and exit of the ␣-chamber (2). In the isolated yeast core particle, the entrance to the degradation channel is blocked by the N termini of the ␣-subunits (7). The channel opens when the regulatory caps bind to the core (12). The caps consist of 17 subunits, six of which have ATPase activity, and contact the co...

show abstract

“…Doa4 deubiquitinates ubiquitinated cargo proteins on the endosomal membrane before they are incorporated into the lumenal vesicles of MVBs (Dupre and Haguenauer-Tsapis, 2001;Katzmann et al, 2001;Losko et al, 2001). In doa4 mutant yeast cells, the cytoplasmic free Ub pool is depleted, suggesting that Doa4 retrieves Ub molecules to the cytoplasm before they are cotransported with cargo proteins to and degraded in vacuoles (Swaminathan et al, 1999;Amerik et al, 2000). In cells in which UBPY was depleted or overexpressed, however, the level of free Ub was unchanged compared with that in normal cells (Figure 7).…”

Section: Role Of Ubpy In Egfr Down-regulationmentioning

confidence: 98%

“…At 3 h after stimulation, the level of EGFR in UBPY-depleted cells was consistently reduced to ϳ50% of that in mock-transfected cells, suggesting that the EGFR down-regulation is accelerated in the absence of UBPY. Doa4, a yeast deubiquitinating enzyme homologous to mammalian UBPY, is implicated in maintaining the level of free Ub in the cytoplasm by recycling Ub molecules from proteasome-and vacuole-targeted ubiquitinated proteins (Swaminathan et al, 1999;Amerik et al, 2000). We therefore examined the effect of depleting or overexpressing UBPY on the cytoplasmic free Ub level in unstimulated cells as well as in cells stimulated with EGF for 15 min.…”

Section: Ubpy Depletion Elevates the Level Of Egfr Ubiquitination Andmentioning

confidence: 99%

Regulation of Epidermal Growth Factor Receptor Down-Regulation by UBPY-mediated Deubiquitination at Endosomes

Mizuno

Iura

Mukai

et al. 2005

MBoC

254

264

View full text Add to dashboard Cite

Ligand-activated receptor tyrosine kinases undergo endocytosis and are transported via endosomes to lysosomes for degradation. This "receptor down-regulation" process is crucial to terminate the cell proliferation signals produced by activated receptors. During the process, ubiquitination of the receptors serves as a sorting signal for their trafficking from endosomes to lysosomes. Here, we describe the role of a deubiquitinating enzyme UBPY/USP8 in the down-regulation of epidermal growth factor (EGF) receptor (EGFR). Overexpression of UBPY reduced the ubiquitination level of EGFR and delayed its degradation in EGF-stimulated cells. Immunopurified UBPY deubiquitinated EGFR in vitro. In EGF-stimulated cells, UBPY underwent ubiquitination and bound to EGFR. Overexpression of Hrs or a dominant-negative mutant of SKD1, proteins that play roles in the endosomal sorting of ubiquitinated receptors, caused the accumulation of endogenous UBPY on exaggerated endosomes. A catalytically inactive UBPY mutant clearly localized on endosomes, where it overlapped with EGFR when cells were stimulated with EGF. Finally, depletion of endogenous UBPY by RNA interference resulted in elevated ubiquitination and accelerated degradation of EGF-activated EGFR. We conclude that UBPY negatively regulates the rate of EGFR down-regulation by deubiquitinating EGFR on endosomes.

show abstract

The Doa4 Deubiquitinating Enzyme Is Functionally Linked to the Vacuolar Protein-sorting and Endocytic Pathways

Cited by 280 publications

References 66 publications

AMSH, an ESCRT-III Associated Enzyme, Deubiquitinates Cargo on MVB/Late Endosomes

AMSH, an ESCRT-III Associated Enzyme, Deubiquitinates Cargo on MVB/Late Endosomes

Concurrent Translocation of Multiple Polypeptide Chains through the Proteasomal Degradation Channel

Regulation of Epidermal Growth Factor Receptor Down-Regulation by UBPY-mediated Deubiquitination at Endosomes

Contact Info

Product

Resources

About