The Catalytic Activity of Ubp6 Enhances Maturation of the Proteasomal Regulatory Particle

Sakata, Eri; Stengel, Florian; Fukunaga, Kimitoshi; Zhou, Min; Saeki, Yasushi; Förster, Friedrich; Baumeister, Wolfgang; Tanaka, Keiji; Robinson, Carol V.

doi:10.1016/j.molcel.2011.04.021

Cited by 61 publications

(67 citation statements)

References 51 publications

(111 reference statements)

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“…Proteasome function is modulated by transiently binding cofactors, the proteasome-interacting proteins (PIPs) (16)(17)(18)(19), which typically are found in substoichiometric amounts in purified proteasomes (20). Of these, ubiquitin C-terminal hydrolase 6 [Ubp6, human ubiquitin-specific protease 14 (Usp14)] is most abundant.…”

Section: Significancementioning

confidence: 99%

Structural characterization of the interaction of Ubp6 with the 26S proteasome

Aufderheide

Beck

Stengel

et al. 2015

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

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In eukaryotic cells, the 26S proteasome is responsible for the regulated degradation of intracellular proteins. Several cofactors interact transiently with this large macromolecular machine and modulate its function. The deubiquitylating enzyme ubiquitin C-terminal hydrolase 6 [Ubp6; ubiquitin-specific protease (USP) 14 in mammals] is the most abundant proteasome-interacting protein and has multiple roles in regulating proteasome function. Here, we investigate the structural basis of the interaction between Ubp6 and the 26S proteasome in the presence and absence of the inhibitor ubiquitin aldehyde. To this end we have used single-particle electron cryomicroscopy in combination with cross-linking and mass spectrometry. Ubp6 binds to the regulatory particle non-ATPase (Rpn) 1 via its N-terminal ubiquitin-like domain, whereas its catalytic USP domain is positioned variably. Addition of ubiquitin aldehyde stabilizes the binding of the USP domain in a position where it bridges the proteasome subunits Rpn1 and the regulatory particle triple-A ATPase (Rpt) 1. The USP domain binds to Rpt1 in the immediate vicinity of the Ubp6 active site, which may effect its activation. The catalytic triad is positioned in proximity to the mouth of the ATPase module and to the deubiquitylating enzyme Rpn11, strongly implying their functional linkage. On the proteasome side, binding of Ubp6 favors conformational switching of the 26S proteasome into an intermediateenergy conformational state, in particular upon the addition of ubiquitin aldehyde. This modulation of the conformational space of the 26S proteasome by Ubp6 explains the effects of Ubp6 on the kinetics of proteasomal degradation.conformational switching | proteolysis | proteostasis | quality control D egradation of proteins that are misfolded, damaged, or no longer needed is an essential element of cellular homeostasis. In eukaryotic cells, the ubiquitin-proteasome system (UPS) is the major pathway for regulated protein degradation (1). Proteins that are processed by the UPS are marked for destruction by polyubiquitin chains, which are recognized as a degradation signal by the 26S proteasome.The 26S proteasome consists of the core particle (CP), which degrades substrates into short peptides, and one or two 19S regulatory particles (RP), which associate with the ends of the cylinder-shaped CP to recruit substrates and prepare them for degradation (2, 3). Although the structure of the CP has been known for more than two decades (4, 5), the molecular architecture of the RP was unraveled by cryo-electron microscope (EM)-based approaches only recently (6-9). It comprises six RP triple A (AAA) ATPases (Rpt), 1-6, and 13 RP non-ATPases (Rpn), 1-3, 5-13, and 15. Similar to AAA-ATPases in prokaryotic ATP-dependent proteases, the Rpts form a hexameric ring that binds to the ends of the CP and is responsible for substrate unfolding and translocation into the CP. Unlike their prokaryotic counterparts, the Rpts are surrounded by non-ATPases. Apart from Rpn1, all Rpns form a cohesive struct...

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Section: Significancementioning

confidence: 99%

Structural characterization of the interaction of Ubp6 with the 26S proteasome

Aufderheide

Beck

Stengel

et al. 2015

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

Self Cite

100

135

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“…C, model of the base assembly. Four assembly intermediates are formed from nine free base subunits and a deubiquitylating enzyme Ubp6 (42). This study suggests that Hsm3 plays a scaffolding role in the formation of the Rpt1 and Rpt2 complex.…”

Section: Structure Of the Complex Of Hsm3 With The C-terminal Domain mentioning

confidence: 76%

“…6C). According to our results and recent studies (42,43), we update a model for the base assembly (Fig. 6C).…”

Section: Structure Of the Complex Of Hsm3 With The C-terminal Domain mentioning

confidence: 91%

Structural Basis for Specific Recognition of Rpt1p, an ATPase Subunit of 26 S Proteasome, by Proteasome-dedicated Chaperone Hsm3p

Takagi

Kim

Yukii

et al. 2012

Journal of Biological Chemistry

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Background: Hsm3 is a proteasome-dedicated chaperone that forms a base precursor, Hsm3-Rpt1-Rpt2-Rpn1. Results: Crystal structures of Hsm3 and Hsm3-Rpt1 were determined. Conclusion: The Hsm3-Rpt1 interface is formed by a hydrophobic core and complementary charged interactions and is important for the proteasome assembly. Significance: The study provides the structural basis for the assembly mechanism of the base subcomplex of the 26 S proteasome.

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“…The ax J / ax J mice had signifi cantly low levels of the USP14 protein, which were associated with defective synaptic transmission in the central and peripheral nervous systems, though this was not associated with any detectable neuronal cell loss (Wilson et al , 2002 ). By a quantitative proteomics approach, we found near stoichiometric binding of Ubp6 to the base precursors of the RP as well as the mature 26S proteasome (Sakata et al , 2011 ). Ubp6 facilitates proteasomal assembly by clearing ubiquitylated substrates from assembly precursors through its deubiquitylating activity.…”

Section: Deubiquitylating Enzymesmentioning

confidence: 88%

“…Intriguingly, recent follow-up studies using more refi ned mass technology to characterize the intact yeast proteasomes, e.g., electrospray ionization mass spectrometry (ESI-MS) of whole complexes, revealed that molecular sizes of the CP, RP, RP-CP, and RP-CP-RP are ∼ 730, ∼ 930, ∼ 1660, and ∼ 2590 kDa, respectively (Sakata et al , 2011 ). The results of subsequent extensive studies indicate that the numbers of subunits in the CP and RP are 14 and 19, respectively.…”

Section: Introductionmentioning

confidence: 99%

The proteasome: molecular machinery and pathophysiological roles

Tanaka

Mizushima

Saeki

2012

BCHM

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108

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The 26S proteasome, in collaboration with ubiquitin, operates the energy-dependent regulated proteolysis process in eukaryotic cells. Over the past 30 years, several studies have comprehensively characterized the structure and molecular/ physiological functions of the 26S proteasome. It is a sophisticated 2.5-MDa protein degradation machine comprising a proteolytic core particle (CP) and one or two terminal regulatory particle(s) (RP). The CP consists of two outer α rings and two inner β rings, which are made up of seven structurally similar α and β subunits, respectively. The CP contains catalytic threonine residues ( β 1, β 2, and β 5; caspase-like, trypsin-like, and chymotrypsin-like activities, respectively) on the inner surface of the chamber formed by two abutting β rings. Intriguingly, the immunotype proteasomes, named ' immunoproteasome ' and ' thymoproteasome ' , whose catalytic subunits are replaced by the related counterparts, were discovered lately. Both unique isoenzymes essentially contribute to the acquisition of adaptive immunity in vertebrates. The RP, which serves to recognize polyubiquitylated substrate proteins and plays a role in their deubiquitylating, unfolding, and translocation into the interior of the CP for destruction, forms two subcomplexes: the base and the lid. On another front, the PA28 and PA200, alternative CP activator proteins discovered biochemically, both play independent roles in proteolysis of the 26S proteasome. Several studies have highlighted the importance of the proteasome in various intractable diseases that have been increasing in the aged society of the 21st century.

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The Catalytic Activity of Ubp6 Enhances Maturation of the Proteasomal Regulatory Particle

Cited by 61 publications

References 51 publications

Structural characterization of the interaction of Ubp6 with the 26S proteasome

Structural characterization of the interaction of Ubp6 with the 26S proteasome

Structural Basis for Specific Recognition of Rpt1p, an ATPase Subunit of 26 S Proteasome, by Proteasome-dedicated Chaperone Hsm3p

The proteasome: molecular machinery and pathophysiological roles

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