The JAMM motif of human deubiquitinase Poh1 is essential for cell viability

Gallery, Melissa; Blank, Jonathan L.; Lin, Yuan; Gutierrez, Juan A.; Pulido, Jacqueline C.; Rappoli, David M.; Badola, Sunita; Rolfe, Mark; MacBeth, Kyle J.

doi:10.1158/1535-7163.mct-06-0542

Cited by 64 publications

(59 citation statements)

References 36 publications

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“…In addition to its function as a DUB, Rpn11 was essential for 26S proteasome structure; thus, the severely impaired function of 26S proteasome in response to RNAi of Rpn11 was at least a consequence of reduced 26S proteasome content. Although several previous studies in yeast, Drosophila, and human cells have shown active-site mutants of Rpn11 to be nonviable, thereby suggesting that Rpn11 DUB activity is essential (Verma et al, 2002;Yao and Cohen, 2002;Lundgren et al, 2004;Gallery et al, 2007), at least one other study has shown an active-site yeast mutant to be viable . Our attempts to distinguish between functional and structural effects of RNAi of Rpn11 by complementation with active site mutants of Rpn11 were unsuccessful due to incomplete incorporation of the transiently expressed subunit (Kulich and DeMartino, unpublished observations).…”

Section: Discussionmentioning

confidence: 94%

Relative Structural and Functional Roles of Multiple Deubiquitylating Proteins Associated with Mammalian 26S Proteasome

Koulich

DeMartino³

2008

MBoC

192

230

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We determined composition and relative roles of deubiquitylating proteins associated with the 26S proteasome in mammalian cells. Three deubiquitylating activities were associated with the 26S proteasome: two from constituent subunits, Rpn11/S13 and Uch37, and one from a reversibly associated protein, Usp14. RNA interference (RNAi) of Rpn11/S13 inhibited cell growth, decreased cellular proteasome activity via disrupted 26S proteasome assembly, and inhibited cellular protein degradation. In contrast, RNAi of Uch37 or Usp14 had no detectable effect on cell growth, proteasome structure or proteolytic capacity, but accelerated cellular protein degradation. RNAi of both Uch37 and Usp14 also had no effect on proteasome structure or proteolytic capacity, but inhibited cellular protein degradation. Thus, proper proteasomal processing of ubiquitylated substrates requires Rpn11 plus either Uch37 or Usp14. Although the latter proteins feature redundant deubiquitylation functions, they also appear to exert noncatalyic effects on proteasome activity that are similar to but independent of one another. These results reveal unexpected functional relationships among multiple deubiquitylating proteins and suggest a model for mammalian 26S proteasome function whereby their concerted action governs proteasome function by linking deubiquitylation to substrate hydrolysis.

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Section: Discussionmentioning

confidence: 94%

Relative Structural and Functional Roles of Multiple Deubiquitylating Proteins Associated with Mammalian 26S Proteasome

Koulich

DeMartino³

2008

MBoC

192

230

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“…We next fitted the Rpn8-Rpn11 core complex crystal structure into the EM densities of the S. cerevisiae 26S proteasome in the substrate-accepting (13) and the substrate-engaged states (27) (Fig. 5 A and B).…”

Section: Resultsmentioning

confidence: 99%

“…The β5 subunits have been clinically validated by the approval of bortezomib and carilfzomib for the treatment of hematologic malignancies. siRNA and mutagenesis studies show that expression of the zinc catalytic domain of hRpn11 is essential for cell survival (27). Inhibition of hRpn11 in combination with EGFR inhibition has been suggested to be beneficial in the treatment of nonsmall cell lung cancer (28).…”

Section: Significancementioning

confidence: 99%

Crystal structure of the proteasomal deubiquitylation module Rpn8-Rpn11

Pathare

Nagy

Śledź

et al. 2014

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

119

128

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The ATP-dependent degradation of polyubiquitylated proteins by the 26S proteasome is essential for the maintenance of proteome stability and the regulation of a plethora of cellular processes. Degradation of substrates is preceded by the removal of polyubiquitin moieties through the isopeptidase activity of the subunit Rpn11. Here we describe three crystal structures of the heterodimer of the Mpr1-Pad1-N-terminal domains of Rpn8 and Rpn11, crystallized as a fusion protein in complex with a nanobody. This fusion protein exhibits modest deubiquitylation activity toward a model substrate. Full activation requires incorporation of Rpn11 into the 26S proteasome and is dependent on ATP hydrolysis, suggesting that substrate processing and polyubiquitin removal are coupled. Based on our structures, we propose that premature activation is prevented by the combined effects of low intrinsic ubiquitin affinity, an insertion segment acting as a physical barrier across the substrate access channel, and a conformationally unstable catalytic loop in Rpn11. The docking of the structure into the proteasome EM density revealed contacts of Rpn11 with ATPase subunits, which likely stabilize the active conformation and boost the affinity for the proximal ubiquitin moiety. The narrow space around the Rpn11 active site at the entrance to the ATPase ring pore is likely to prevent erroneous deubiquitylation of folded proteins.n eukaryotes, the ubiquitin (Ub) proteasome system (UPS) is responsible for the regulated degradation of proteins (1-5). The UPS plays a key role in the maintenance of protein homeostasis by removing misfolded or damaged proteins, which could impair cellular functions, and by removing proteins whose functions are no longer needed. Consequently, the UPS is critically involved in numerous cellular processes, including cell cycle progression, apoptosis, and DNA damage repair, and malfunctions of the system often result in disease.The 26S proteasome executes the degradation of substrates that are marked for destruction by the covalent attachment of polyubiquitin chains. It is a molecular machine of 2.5 MDa comprising two subcomplexes, the 20S core particle (CP) and one or two 19S regulatory particles (RPs), which associate with the ends of the cylinder-shaped CP (6-8). The recognition and recruitment of polyubiquitylated substrates, their deubiquitylation, ATP-dependent unfolding, and translocation into the core particle take place in the RP. The structurally and mechanistically well-characterized CP houses the proteolytic activities and sequesters them from the environment, thereby avoiding collateral damage (9).The RPs attach to the outer α-rings of the CP, which control access to the proteolytic chamber formed by the inner β-subunit rings (10). Recently, the molecular architecture of the 26S holocomplex was established using cryo-EM-based approaches (11,12), and a pseudoatomic model of the holocomplex was put forward (13). The RP is formed by two subcomplexes, known as the base and the lid, which assemble independent...

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“…POH1 is a metalloprotease belonging to the JAMM domain family and is an integral part of the 19S RP lid (Verma et al, 2002;Yao and Cohen, 2002). POH1 is essential for the viability of both yeast and metazoan cells, with siRNA depletion resulting in the destabilization of the 19S RP lid complex (Gallery et al, 2007;Rinaldi et al, 1998). POH1 contains a JAMM/MPN + motif sequence where an aspartic acid and two histidine residues coordinate a zinc ion that is essential for DUB activity (Ambroggio et al, 2004;MaytalKivity et al, 2002).…”

Section: The Role Of Proteasome Associated Deubiquitinases (Dubs)mentioning

confidence: 99%

“…In particular targeting proteasome-associated deubiquitination has the potential for increasing the levels of ubiquitin conjugated proteins triggering proteotoxic stress and apoptosis similar to that observed with inhibitors of proteolytic activity. The metalloprotease POH1 is an obvious target since it is absolutely required for cell viability and is overexpressed in a variety of tumours (Gallery et al, 2007). Although neither USP14 nor UCHL5 appears to be essential for cell survival alone, dual knockdown using RNA interference has been shown to lead to the accumulation of polyubiquitinated substrates and loss of cell viability (Koulich et al, 2008;Tian et al, 2013;Wang et al, 2014).…”

Section: Proteasomal Dubs As Drug Targets For Cancer Therapymentioning

confidence: 99%

Inhibition of proteasome deubiquitinase activity: a strategy to overcome resistance to conventional proteasome inhibitors?

Selvaraju

Mazurkiewicz

Wang

et al. 2015

Drug Resistance Updates

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The ubiquitin-proteasome system (UPS) is the primary mechanism controlling the degradation of damaged, unwanted or short-lived proteins in eukaryotic cells. In addition to protein homeostasis, the UPS has also emerged as a critical node in the regulation of signalling pathways implicated in the growth and survival of cancer cells. The absolute dependency of cancer cells on a functioning UPS has been exploited in the development of anti-cancer therapies as exemplified by development of proteasome inhibitors for the treatment of certain leukemic malignancies.Deubiquitinases (DUBs) are enzyme components of the UPS that catalyse re-editing of poly ubiquitin chains and/or removal of ubiquitin en bloc from target substrates leading to alterations in protein stability and/or downstream signalling. There is a growing recognition that targeting DUB activity may be a feasible option for the development of novel anti-cancer therapies. In particular inhibition of proteasomal cysteine DUBs (i.e. USP14 and UCHL5) has been shown to be particularly cytotoxic to cancer cells leading to the accumulation of ubiquitinated proteins and proteotoxic stress. In this review we focus on the mechanisms of action of proteasome DUB inhibitors as well as the potential of such compounds to circumvent acquired drug resistance in cancer patients.

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The JAMM motif of human deubiquitinase Poh1 is essential for cell viability

Cited by 64 publications

References 36 publications

Relative Structural and Functional Roles of Multiple Deubiquitylating Proteins Associated with Mammalian 26S Proteasome

Relative Structural and Functional Roles of Multiple Deubiquitylating Proteins Associated with Mammalian 26S Proteasome

Crystal structure of the proteasomal deubiquitylation module Rpn8-Rpn11

Inhibition of proteasome deubiquitinase activity: a strategy to overcome resistance to conventional proteasome inhibitors?

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