• Journal of Cancer Prevention
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• 제23권4호
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• pp.153-161
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• 2018

Imbalance of protein homeostasis (proteostasis) is known to cause cellular malfunction, cell death, and diseases. Elaborate regulation of protein synthesis and degradation is one of the important processes in maintaining normal cellular functions. Protein degradation pathways in eukaryotes are largely divided into proteasome-mediated degradation and lysosome-mediated degradation. Proteasome is a multisubunit complex that selectively degrades 80% to 90% of cellular proteins. Proteasome-mediated degradation can be divided into 26S proteasome (20S proteasome + 19S regulatory particle) and free 20S proteasome degradation. In 1980, it was discovered that during ubiquitination process, wherein ubiquitin binds to a substrate protein in an ATP-dependent manner, ubiquitin acts as a degrading signal to degrade the substrate protein via proteasome. Conversely, 20S proteasome degrades the substrate protein without using ATP or ubiquitin because it recognizes the oxidized and structurally modified hydrophobic patch of the substrate protein. To date, most studies have focused on protein degradation via 26S proteasome. This review describes the 26S/20S proteasomal pathway of protein degradation and discusses the potential of proteasome as therapeutic targets for cancer treatment as well as against diseases caused by abnormalities in the proteolytic system.

• 한국동물학회지
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• 제37권3호
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• pp.324-330
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• 1994

The multicatalvtic 205 proteasome consisting of 12-15 subunits of 22-35 kDa is the catalytic core of the ATP/ubiquitin-dependent 26S protease complex that also is comprised of multiple subunits of 22-110 KDa. In order to determine whether the proteolvtic activities change during muscle development, the enzyme preparations were obtained from 11-, 14- and 17-day old chick embryonic muscle using a BioGel A-1.5m column. The 26S complex preparation from 14- or 17-day old muscle hvdr olvz e d both N -s uccinvl- Le u- Le u -Val-Tvr-7- amido -4- methvlco umarin ( Suc- LLVY- AMC) and ubiquitin-Ivsozvme conjugates about 50% as well as that from 11-day old muscle. In addition, the activity of 20S proteBsome against Suc-LLVY-AMC also decreased by about 20-30%. However, the protein level of 265 complex remained constant during the entire development period, while that of 205 proteasome increased 5- to 6-fold, as analyzed by nondenaturins polyacrvlamide gel elenrophoresis followed by immunoblot analysis using the antibodies raised against the purified enzymes. Thus, the specific activity of 20S proteasome against the peptide must decrease rather dramatically during the muscle development. These results suggest that the development-dependent changes in the proteolytic activities of both 20S proteasome and 26S protease complect from embryonic muscle are due to alterations in the expression of certain subunits in the enzvmes that are responsible for their specific cataIVtic functions but not to overall changes in the enzyme amounts.

• BMB Reports
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• 제49권9호
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• pp.459-473
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• 2016

Neurodegenerative diseases (NDs) often involve the formation of abnormal and toxic protein aggregates, which are thought to be the primary factor in ND occurrence and progression. Aged neurons exhibit marked increases in aggregated protein levels, which can lead to increased cell death in specific brain regions. As no specific drugs/therapies for treating the symptoms or/and progression of NDs are available, obtaining a complete understanding of the mechanism underlying the formation of protein aggregates is needed for designing a novel and efficient removal strategy. Intracellular proteolysis generally involves either the lysosomal or ubiquitin-proteasome system. In this review, we focus on the structure and assembly of the proteasome, proteasome-mediated protein degradation, and the multiple dynamic regulatory mechanisms governing proteasome activity. We also discuss the plausibility of the correlation between changes in proteasome activity and the occurrence of NDs.

• 약학회지
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• 제55권6호
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• pp.466-472
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• 2011

NF-E2-related factor 2 (Nrf2), a master regulator of antioxidant genes in animals, has been associated with the resistance of cancer cells to several cytotoxic chemotherapeutics. Bortezomib, a reversible inhibitor of the 26S proteasome, is a novel class anti-cancer therapeutics approved for the treatment of refractory multiple myeloma. However, the molecular mechanism of drug-resistance remains elusive. In the present study, bortezomib sensitivity has been investigated in Nrf2 knockdown ovarian cancer cells. When Nrf2 expression is stably repressed using interfering RNA expression, bortezomib-induced apoptosis and cell death were significantly enhanced compared to nonspecific RNA control cells. Knockdown cells showed elevated expression in the catalytic subunit PSMB5, PSMB6, and PSMB7 compared to the control, and failed to induce heme oxygenase-1 expression following bortezomib treatment. These indicate that differential proteasome levels and altered expression of stress-response genes could be underlying mechanisms of bortezomib sensitization in Nrf2-inhibited ovarian cancer cells.

• Molecules and Cells
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• 제28권4호
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• pp.375-382
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• 2009

The 26S proteasome is a 2-MDa complex with a central role in protein turn over. The 26S proteasome is comprised of one 20S core particle and two 19S regulatory particles (RPs). The RPN12a protein, a non-ATPase subunit of the 19S RP, was previously shown to be involved in cytokinin signaling in Arabidopsis. To further investigate cellular roles of RPN12a, RNAi transgenic plants of RPN12a were constructed. As expected, the 35S:RNAi-RPN12a plants showed cytokinin signaling defective phenotypes, including abnormal formation of leaves and inflorescences. Furthermore, RNAi knock-down transgenic plants exhibited additional unique phenotypes, including concave and heart-shape cotyledons, triple cotyledons, irregular and clustered guard cells, and defects in phyllotaxy, all of which are typical for defective cytokinin signaling. We next examined the mRNA level of cytokinin signaling components, including type-A ARRs, type-B ARRs, and CRFs. The expression of type-A ARRs, encoding negative regulators of cytokinin signaling, was markedly reduced in 35S:RNAi-RPN12a transgenic plants relative to that in wild type plants, while type-B ARRs and CRFs were unaffected. Our results also indicate that in vivo stability of the ARR5 protein, a negative regulator of cytokinin signaling, is mediated by the 26S proteasome complex. These results suggest that RPN12a participates in feedback inhibitory mechanism of cytokinin signaling through modulation of the abundance of ARR5 protein in Arabidopsis.

• Animal cells and systems
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• 제14권1호
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• pp.1-10
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• 2010

Previously we showed that CHIP, a co-chaperone of Hsp70 and E3 ubiquitin ligase, can promote the degradation of mutant SOD1 linked to familial amyotrophic lateral sclerosis (fALS) via a mechanism not involving SOD1 ubiquitylation. Here we present evidence that CHIP functions in the interaction of mutant SOD1 with 26S proteasomes. Bag-1, a coupling factor between molecular chaperones and the proteasomes, formed a complex with SOD1 in an hsp70-dependent manner but had no direct effect on the degradation of mutant SOD1. Instead, Bag-1 stimulated interaction between CHIP and the proteasome-associated protein VCP (p97), which do not associate normally. Over-expressed CHIP interfered with the association between mutant SOD1 and VCP. Conversely, the binding of CHIP to mutant SOD1 was inhibited by VCP, implying that the chaperone complex and proteolytic machinery are competing for the common substrates. Finally we observed that mutant SOD1 strongly associated with the 19S complex of proteasomes and CHIP over-expression specifically reduced the interaction between S6/S6' ATPase subunits and mutant SOD1. These results suggest that CHIP, together with ubiquitin-binding proteins such as Bag-1 and VCP, promotes the degradation of mutant SOD1 by facilitating its translocation from ATPase subunits of 19S complex to the 20S core particle.

• The Plant Pathology Journal
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• 제24권4호
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• pp.367-374
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• 2008

The genomes of plants contain more than 600 genes encoding a diverse set of proteases and the subunits of proteasomes. These proteases and proteasomes consist of plant proteolytic systems, which are involved in various cellular metabolic processes. Plant proteolytic systems have been shown to have diverse roles in defense responses, such as execution of the attack on the invading organisms, participation in signaling cascades, and perception of the invaders. In order to provide a framework for illustrating the importance of proteolytic systems in plant defense, characteristics of non-proteasome proteases and the 26S proteasome are summarized. The involvement of caspase-like proteases, saspases, apoplastic proteases, and the 26S proteasome in pathogen defense suggests that plant proteolytic systems are essential for defense and further clarity on the roles of plant proteases in defense is challenging but fundamentally important to understand plant-microbe interactions.

• BMB Reports
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• 제50권8호
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• pp.393-400
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• 2017

Plants are constantly exposed to a variety of abiotic stresses, such as drought, heat, cold, flood, and salinity. To survive under such unfavorable conditions, plants have evolutionarily developed their own resistant-mechanisms. For several decades, many studies have clarified specific stress response pathways of plants through various molecular and genetic studies. In particular, it was recently discovered that ubiquitin proteasome system (UPS), a regulatory mechanism for protein turn over, is greatly involved in the stress responsive pathways. In the UPS, many E3 ligases play key roles in recognizing and tethering poly-ubiquitins on target proteins for subsequent degradation by the 26S proteasome. Here we discuss the roles of RING ligases that have been defined in related to abiotic stress responses in plants.

• BMB Reports
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• 제47권5호
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• pp.292-297
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• 2014

Age-related macular degeneration (AMD) is the leading cause of blindness in the world. Evidence indicates that the suppression of the ubiquitin-proteasome system (UPS) contributes to the accumulation of toxic proteins and inflammation in retinal pigment epithelium (RPE), the functional abnormalities and/or the degeneration of which are believed to be the initiators and major pathologies of AMD. To identify new protein associations with the altered UPS in AMD, we used LC-ESI-MS/MS to perform a proteomic analysis of the aqueous humor (AH) of AMD patients and matched control subjects. Six UPS-related proteins were present in the AH of the patients and control subjects. Four of the proteins, including 26S proteasome non-ATPase regulatory subunit 1 (Rpn2), were increased in patients, according to semi-quantitative proteomic profiling. An LC-MRM assay revealed a significant increase of Rpn2 in 15 AMD patients compared to the control subjects, suggesting that this protein could be a biomarker for AMD.

• IMMUNE NETWORK
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• 제22권3호
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• pp.28.1-28.11
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• 2022

The 26S proteasome irreversibly hydrolyzes polyubiquitylated substrates to maintain protein homeostasis; it also regulates immune responses by generating antigenic peptides. An alternative form of the 26S proteasome is the immunoproteasome, which contains substituted catalytic subunits (β1i/PSMB9, β2i/PSMB10, and β5i/PSMB8) instead of constitutively expressed counterparts (β1/PSMB6, β2/PSMB7, and β5/PSMB5). The immunoproteasome expands the peptide repertoire presented on MHC class I molecules. However, how its activity changes in this context is largely elusive, possibly due to the lack of a standardized methodology to evaluate its specific activity. Here, we describe an assay protocol that measures the immunoproteasome activity of whole-cell lysates using commercially available fluorogenic peptide substrates. Our results showed that the most accurate assessment of immunoproteasome activity could be achieved by combining β5i-targeting substrate Ac-ANW-AMC and immunoproteasome inhibitor ONX-0914. This simple and reliable protocol may contribute to future studies of immunoproteasomes and their pathophysiological roles during viral infection, inflammation, and tumorigenesis.