Co-Chaperones in Targeting and Delivery of Misfolded Proteins to the 26S Proteasome

Protein homeostasis (proteostasis) is essential for the cell and is maintained by a highly conserved protein quality control (PQC) system, which triages newly synthesized, mislocalized and misfolded proteins. The ubiquitin-proteasome system (UPS), molecular chaperones, and co-chaperones are vital PQC elements that work together to facilitate degradation of misfolded and toxic protein species through the 26S proteasome. However, the underlying mechanisms are complex and remain partly unclear. Here, we provide an overview of the current knowledge on the co-chaperones that directly take part in targeting and delivery of PQC substrates for degradation. While J-domain proteins (JDPs) target substrates for the heat shock protein 70 (HSP70) chaperones, nucleotide-exchange factors (NEFs) deliver HSP70-bound substrates to the proteasome. So far, three NEFs have been established in proteasomal delivery: HSP110 and the ubiquitin-like (UBL) domain proteins BAG-1 and BAG-6, the latter acting as a chaperone itself and carrying its substrates directly to the proteasome. A better understanding of the individual delivery pathways will improve our ability to regulate the triage, and thus regulate the fate of aberrant proteins involved in cell stress and disease, examples of which are given throughout the review.

Download Full-text

Cytosolic splice isoform of Hsp70 nucleotide exchange factor Fes1 is required for the degradation of misfolded proteins in yeast

Molecular Biology of the Cell ◽

10.1091/mbc.e15-10-0697 ◽

2016 ◽

Vol 27 (8) ◽

pp. 1210-1219 ◽

Cited By ~ 18

Author(s):

Naveen Kumar Chandappa Gowda ◽

Jayasankar Mohanakrishnan Kaimal ◽

Anna E. Masser ◽

Wenjing Kang ◽

Marc R. Friedländer ◽

...

Keyword(s):

Elevated Temperatures ◽

Protein Quality ◽

Ectopic Expression ◽

Ubiquitin Proteasome System ◽

Misfolded Proteins ◽

Control Mechanisms ◽

Nucleotide Exchange ◽

Exchange Factor ◽

Nucleotide Exchange Factor ◽

Ubiquitin Proteasome

Cells maintain proteostasis by selectively recognizing and targeting misfolded proteins for degradation. In Saccharomyces cerevisiae, the Hsp70 nucleotide exchange factor Fes1 is essential for the degradation of chaperone-associated misfolded proteins by the ubiquitin-proteasome system. Here we show that the FES1 transcript undergoes unique 3′ alternative splicing that results in two equally active isoforms with alternative C-termini, Fes1L and Fes1S. Fes1L is actively targeted to the nucleus and represents the first identified nuclear Hsp70 nucleotide exchange factor. In contrast, Fes1S localizes to the cytosol and is essential to maintain proteostasis. In the absence of Fes1S, the heat-shock response is constitutively induced at normally nonstressful conditions. Moreover, cells display severe growth defects when elevated temperatures, amino acid analogues, or the ectopic expression of misfolded proteins, induce protein misfolding. Importantly, misfolded proteins are not targeted for degradation by the ubiquitin-proteasome system. These observations support the notion that cytosolic Fes1S maintains proteostasis by supporting the removal of toxic misfolded proteins by proteasomal degradation. This study provides key findings for the understanding of the organization of protein quality control mechanisms in the cytosol and nucleus.

Download Full-text

Protein Quality Control Degradation in the Nucleus

Annual Review of Biochemistry ◽

10.1146/annurev-biochem-062917-012730 ◽

2018 ◽

Vol 87 (1) ◽

pp. 725-749 ◽

Cited By ~ 20

Author(s):

Charisma Enam ◽

Yifat Geffen ◽

Tommer Ravid ◽

Richard G. Gardner

Keyword(s):

Quality Control ◽

Protein Misfolding ◽

Protein Quality ◽

Current Knowledge ◽

Protein Quality Control ◽

Ubiquitin Proteasome System ◽

Misfolded Proteins ◽

Cellular Processes ◽

Human Disorders ◽

Protein Misfolding And Aggregation

Nuclear proteins participate in diverse cellular processes, many of which are essential for cell survival and viability. To maintain optimal nuclear physiology, the cell employs the ubiquitin-proteasome system to eliminate damaged and misfolded proteins in the nucleus that could otherwise harm the cell. In this review, we highlight the current knowledge about the major ubiquitin-protein ligases involved in protein quality control degradation (PQCD) in the nucleus and how they orchestrate their functions to eliminate misfolded proteins in different nuclear subcompartments. Many human disorders are causally linked to protein misfolding in the nucleus, hence we discuss major concepts that still need to be clarified to better understand the basis of the nuclear misfolded proteins’ toxic effects. Additionally, we touch upon potential strategies for manipulating nuclear PQCD pathways to ameliorate diseases associated with protein misfolding and aggregation in the nucleus.

Download Full-text

The Roles of Ubiquitin-Binding Protein Shuttles in the Degradative Fate of Ubiquitinated Proteins in the Ubiquitin-Proteasome System and Autophagy

Cells ◽

10.3390/cells8010040 ◽

2019 ◽

Vol 8 (1) ◽

pp. 40 ◽

Cited By ~ 27

Author(s):

Katarzyna Zientara-Rytter ◽

Suresh Subramani

Keyword(s):

Protein Quality ◽

Current Knowledge ◽

Binding Protein ◽

Ubiquitin Proteasome System ◽

Oligomeric State ◽

Receptor Associated Protein ◽

Ubiquitin Proteasome ◽

Specific Proteins ◽

And Control ◽

Recognition Code

The ubiquitin-proteasome system (UPS) and autophagy are the two major intracellular protein quality control (PQC) pathways that are responsible for cellular proteostasis (homeostasis of the proteome) by ensuring the timely degradation of misfolded, damaged, and unwanted proteins. Ubiquitination serves as the degradation signal in both these systems, but substrates are precisely targeted to one or the other pathway. Determining how and when cells target specific proteins to these two alternative PQC pathways and control the crosstalk between them are topics of considerable interest. The ubiquitin (Ub) recognition code based on the type of Ub-linked chains on substrate proteins was believed to play a pivotal role in this process, but an increasing body of evidence indicates that the PQC pathway choice is also made based on other criteria. These include the oligomeric state of the Ub-binding protein shuttles, their conformation, protein modifications, and the presence of motifs that interact with ATG8/LC3/GABARAP (autophagy-related protein 8/microtubule-associated protein 1A/1B-light chain 3/GABA type A receptor-associated protein) protein family members. In this review, we summarize the current knowledge regarding the Ub recognition code that is bound by Ub-binding proteasomal and autophagic receptors. We also discuss how cells can modify substrate fate by modulating the structure, conformation, and physical properties of these receptors to affect their shuttling between both degradation pathways.

Download Full-text

Asaronic Acid Inhibits ER Stress and Ameliorates Impaired ERAD in 7Β-Hydroxycholesterol-Loaded Macrophages

Current Developments in Nutrition ◽

10.1093/cdn/nzab037_062 ◽

2021 ◽

Vol 5 (Supplement_2) ◽

pp. 352-352

Author(s):

Hyeongjoo Oh ◽

Young-Hee Kang

Keyword(s):

Er Stress ◽

Protein Quality ◽

Metabolic Stress ◽

Ubiquitin Proteasome System ◽

Misfolded Proteins ◽

Immunocytochemical Staining ◽

Funding Sources ◽

Subsequent Degradation ◽

Ubiquitin Proteasome ◽

Stress Damage

Abstract Objectives Misfolded proteins were formed in the endoplasmic reticulum (ER) due to diverse stresses including metabolic stress and oxidative stress. Accumulation of unfolded proteins in the ER stimulates chaperone expression and ER-associated degradation (ERAD) process. This process involves the recognition of misfolded proteins to maintain the protein quality control, which in turn eliminates in association with the ER membrane. Upregulation of ubiquitination enzymes is an essential mechanism by which ER stress enhances ERAD. Asaronic acid (2,4,5-trimethoxybenzoic acid), identified as one of purple perilla constituents, has anti-diabetic and anti-inflammatory effects. This study attempted to examine whether asaronic acid attenuated the 7Β-hydroxycholesterol-elicited ER stress of macrophages. Methods J774A.1 murine macrophage was incubated with 28 μM 7Β-hydroxycholesterol in absence and presence of 1–20 μΜ asaronic acid up to 24 h. Cytotoxicity was assessed by MTT assay. Expression levels of ER stress-responsive chaperones and ERAD biomarkers were measured by Western blot analysis and immunocytochemical staining with a specific antibody. Results Asaronic acid at 1–20 μM had a cytoprotective effect on macrophages against 7Β-hydroxycholesterol-induced toxicity. Asaronic acid diminished the induction and activation of ER stress sensors such as Grp/BiP, IRE1, and PERK in macrophages exposed to 7Β-hydroxycholesterol. Also, asaronic acid positively influenced the induction of ERAD process-linked components of EDEM1, OS9, SEl1L, HRD1, and VCP1/p97. Furthermore, asaronic acid promoted subsequent degradation reduced by 7Β-hydroxycholesterol via the cytosolar ubiquitin-proteasome system of macrophages. Conclusions These results demonstrate that asaronic acid attenuated 7Β-hydroxycholesterol-induced ER stress and improved impaired ER stress-mediated degradation systems. Therefore, asaronic acid may be a potent agent protecting macrophages against pathological ER stress damage. Funding Sources This work was supported by the BK21 FOUR(Fostering Outstanding Universities for Research, 4220200913807) funded by the National Research Foundation of Korea (NRF).

Download Full-text

Neurodegenerative Diseases as Protein Misfolding Disorders

10.1093/med/9780190233563.003.0002 ◽

2016 ◽

Author(s):

Tomohiro Nakamura ◽

Stuart A. Lipton

Keyword(s):

Quality Control ◽

Neurodegenerative Diseases ◽

Protein Misfolding ◽

Protein Quality ◽

Protein S ◽

Ubiquitin Proteasome System ◽

Misfolded Proteins ◽

Redox Stress ◽

Chemical Mechanisms ◽

Ubiquitin Proteasome

Neurodegenerative diseases (NDDs) often represent disorders of protein folding. Rather than large aggregates, recent evidence suggests that soluble oligomers of misfolded proteins are the most neurotoxic species. Emerging evidence points to small, soluble oligomers of misfolded proteins as the cause of synaptic dysfunction and loss, the major pathological correlate to disease progression in many NDDs including Alzheimer’s disease. The protein quality control machinery of the cell, which includes molecular chaperones as found in the endoplasmic reticulum (ER), the ubiquitin-proteasome system (UPS), and various forms of autophagy, can counterbalance the accumulation of misfolded proteins to some extent. Their ability to eliminate the neurotoxic effects of misfolded proteins, however, declines with age. A plausible explanation for the age-dependent deterioration of the quality control machinery involves compromise of these systems by excessive generation of reactive oxygen species (ROS), such as superoxide anion (O2-), and reactive nitrogen species (RNS), such as nitric oxide (NO). The resulting redox stress contributes to the accumulation of misfolded proteins. Here, we focus on aberrantly increased generation of NO-related species since this process appears to accelerate the manifestation of key neuropathological features, including protein misfolding. We review the chemical mechanisms of posttranslational modification by RNS such as protein S-nitrosylation of critical cysteine thiol groups and nitration of tyrosine residues, showing how they contribute to the pathogenesis of NDDs.

Download Full-text

Releasing the Lockdown: An Emerging Role for the Ubiquitin-Proteasome System in the Breakdown of Transient Protein Inclusions

Biomolecules ◽

10.3390/biom10081168 ◽

2020 ◽

Vol 10 (8) ◽

pp. 1168 ◽

Cited By ~ 1

Author(s):

Yuval Reiss ◽

Elisheva Gur ◽

Tommer Ravid

Keyword(s):

Protein Quality ◽

Ubiquitin Proteasome System ◽

Biological Properties ◽

26S Proteasome ◽

Turnover Rates ◽

Functional Link ◽

Protein Deposits ◽

Dead End ◽

Ubiquitin Proteasome ◽

Quality Control Mechanism

Intracellular protein inclusions are diverse cellular entities with distinct biological properties. They vary in their protein content, sequestration sites, physiological function, conditions for their generation, and turnover rates. Major distinctions have been recognized between stationary amyloids and dynamic, misfolded protein deposits. The former being a dead end for irreversibly misfolded proteins, hence, cleared predominantly by autophagy, while the latter consists of a protein-quality control mechanism, important for cell endurance, where proteins are sequestered during proteotoxic stress and resolved upon its relief. Accordingly, the disaggregation of transient inclusions is a regulated process consisting of protein solubilization, followed by a triage step to either refolding or to ubiquitin-mediated degradation. Recent studies have demonstrated an indispensable role in disaggregation for components of the chaperone and the ubiquitin–proteasome systems. These include heat-shock chaperones of the 40/70/100 kDa families, the proteasome, proteasome substrate shuttling factors, and deubiquitylating enzymes. Thus, a functional link has been established between the chaperone machinery that extracts proteins from transient deposits and 26S proteasome-dependent disaggregation, indicative of a coordinated process. In this review, we discuss data emanating from these important studies and subsequently consolidate the information in the form of a working model for the disaggregation mechanism.

Download Full-text

The Cytoplasmic Hsp70 Chaperone Machinery Subjects Misfolded and Endoplasmic Reticulum Import-incompetent Proteins to Degradation via the Ubiquitin–Proteasome System

Molecular Biology of the Cell ◽

10.1091/mbc.e06-04-0338 ◽

2007 ◽

Vol 18 (1) ◽

pp. 153-165 ◽

Cited By ~ 113

Author(s):

Sae-Hun Park ◽

Natalia Bolender ◽

Frederik Eisele ◽

Zlatka Kostova ◽

Junko Takeuchi ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Protein Quality ◽

Signal Sequence ◽

Degradation Process ◽

Ubiquitin Proteasome System ◽

Protein Domain ◽

Misfolded Proteins ◽

Misfolded Protein ◽

Ubiquitin Proteasome ◽

Shock Proteins

The mechanism of protein quality control and elimination of misfolded proteins in the cytoplasm is poorly understood. We studied the involvement of cytoplasmic factors required for degradation of two endoplasmic reticulum (ER)-import–defective mutated derivatives of carboxypeptidase yscY (ΔssCPY* and ΔssCPY*-GFP) and also examined the requirements for degradation of the corresponding wild-type enzyme made ER-import incompetent by removal of its signal sequence (ΔssCPY). All these protein species are rapidly degraded via the ubiquitin–proteasome system. Degradation requires the ubiquitin-conjugating enzymes Ubc4p and Ubc5p, the cytoplasmic Hsp70 Ssa chaperone machinery, and the Hsp70 cochaperone Ydj1p. Neither the Hsp90 chaperones nor Hsp104 or the small heat-shock proteins Hsp26 and Hsp42 are involved in the degradation process. Elimination of a GFP fusion (GFP-cODC), containing the C-terminal 37 amino acids of ornithine decarboxylase (cODC) directing this enzyme to the proteasome, is independent of Ssa1p function. Fusion of ΔssCPY* to GFP-cODC to form ΔssCPY*-GFP-cODC reimposes a dependency on the Ssa1p chaperone for degradation. Evidently, the misfolded protein domain dictates the route of protein elimination. These data and our further results give evidence that the Ssa1p-Ydj1p machinery recognizes misfolded protein domains, keeps misfolded proteins soluble, solubilizes precipitated protein material, and escorts and delivers misfolded proteins in the ubiquitinated state to the proteasome for degradation.

Download Full-text

p62-containing, proteolytically active nuclear condensates, increase the efficiency of the ubiquitin–proteasome system

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2107321118 ◽

2021 ◽

Vol 118 (33) ◽

pp. e2107321118

Author(s):

Afu Fu ◽

Victoria Cohen-Kaplan ◽

Noa Avni ◽

Ido Livneh ◽

Aaron Ciechanover

Keyword(s):

Ubiquitin Ligase ◽

Protein Quality ◽

Ubiquitin Proteasome System ◽

Cellular Protein ◽

26S Proteasome ◽

Ubiquitin Chain ◽

Recognition Element ◽

Oxidative Stresses ◽

Ubiquitin Ligase E3 ◽

Ubiquitin Proteasome

Degradation of a protein by the ubiquitin–proteasome system (UPS) is a multistep process catalyzed by sequential reactions. Initially, ubiquitin is conjugated to the substrate in a process mediated by concerted activity of three enzymes; the last of them—a ubiquitin ligase (E3)—belongs to a family of several hundred members, each recognizing a few specific substrates. This is followed by repeated addition of ubiquitin moieties to the previously conjugated one to generate a ubiquitin chain that serves as a recognition element for the proteasome, which then degrades the substrate. Ubiquitin is recycled via the activity of deubiquitinating enzymes (DUBs). It stands to reason that efficiency of such a complex process would depend on colocalization of the different components in an assembly that allows the reactions to be carried out sequentially and processively. Here we describe nuclear condensates that are dynamic in their composition. They contain p62 as an essential component. These assemblies are generated by liquid–liquid phase separation (LLPS) and also contain ubiquitinated targets, 26S proteasome, the three conjugating enzymes, and DUBs. Under basal conditions, they serve as efficient centers for proteolysis of nuclear proteins (e.g., c-Myc) and unassembled subunits of the proteasome, suggesting they are involved in cellular protein quality control. Supporting this notion is the finding that such foci are also involved in degradation of misfolded proteins induced by heat and oxidative stresses, following recruitment of heat shock proteins and their associated ubiquitin ligase CHIP.

Download Full-text

The Degradation-promoting Roles of Deubiquitinases Ubp6 and Ubp3 in Cytosolic and ER Protein Quality Control

10.1101/2020.01.22.915793 ◽

2020 ◽

Author(s):

Hongyi Wu ◽

Davis T.W. Ng ◽

Ian Cheong ◽

Paul Matsudaira

Keyword(s):

Saccharomyces Cerevisiae ◽

Quality Control ◽

Molecular Chaperones ◽

Protein Quality ◽

Protein Quality Control ◽

Ubiquitin Proteasome System ◽

Misfolded Proteins ◽

Intracellular Proteins ◽

Ubiquitin Proteasome ◽

Free Ubiquitin

AbstractThe quality control of intracellular proteins is achieved by degrading misfolded proteins which cannot be refolded by molecular chaperones. In eukaryotes, such degradation is handled primarily by the ubiquitin-proteasome system. However, it remains unclear whether and how protein quality control deploys various deubiquitinases. To address this question, we screened deletions or mutation of the 20 deubiquitinase genes in Saccharomyces cerevisiae and discovered that almost half of the mutations slowed the removal of misfolded proteins whereas none of the remaining mutations accelerated this process significantly. Further characterization revealed that Ubp6 maintains the level of free ubiquitin to promote the elimination of misfolded cytosolic proteins, while Ubp3 supports the degradation of misfolded cytosolic and ER luminal proteins by different mechanisms.

Download Full-text

Protein Quality Control of the Endoplasmic Reticulum and Ubiquitin–Proteasome-Triggered Degradation of Aberrant Proteins: Yeast Pioneers the Path

Annual Review of Biochemistry ◽

10.1146/annurev-biochem-062917-012749 ◽

2018 ◽

Vol 87 (1) ◽

pp. 751-782 ◽

Cited By ~ 51

Author(s):

Nicole Berner ◽

Karl-Richard Reutter ◽

Dieter H. Wolf

Keyword(s):

Quality Control ◽

Endoplasmic Reticulum ◽

Protein Quality ◽

Protein Structures ◽

Protein Quality Control ◽

Ubiquitin Proteasome System ◽

Misfolded Proteins ◽

Secretory Proteins ◽

Structural Alterations ◽

Ubiquitin Proteasome

Cells must constantly monitor the integrity of their macromolecular constituents. Proteins are the most versatile class of macromolecules but are sensitive to structural alterations. Misfolded or otherwise aberrant protein structures lead to dysfunction and finally aggregation. Their presence is linked to aging and a plethora of severe human diseases. Thus, misfolded proteins have to be rapidly eliminated. Secretory proteins constitute more than one-third of the eukaryotic proteome. They are imported into the endoplasmic reticulum (ER), where they are folded and modified. A highly elaborated machinery controls their folding, recognizes aberrant folding states, and retrotranslocates permanently misfolded proteins from the ER back to the cytosol. In the cytosol, they are degraded by the highly selective ubiquitin–proteasome system. This process of protein quality control followed by proteasomal elimination of the misfolded protein is termed ER-associated degradation (ERAD), and it depends on an intricate interplay between the ER and the cytosol.

Download Full-text