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International Journal of Molecular... Oct 2023This study investigated modifications to the ubiquitin proteasome system (UPS) in a mouse model of type 2 diabetes mellitus (T2DM) and their relationship to heart...
This study investigated modifications to the ubiquitin proteasome system (UPS) in a mouse model of type 2 diabetes mellitus (T2DM) and their relationship to heart complications. mice heart tissues were compared with mice tissues using RNA sequencing, qRT-PCR, and protein analysis to identify cardiac UPS modifications associated with diabetes. The findings unveiled a distinctive gene profile in the hearts of mice with decreased levels of mRNA and increased levels of , indicating potential cardiac dysfunction. The mRNA levels of (deubiquitinating enzyme), , and (proteasome β-subunits) were down-regulated in mice, while the mRNA levels of RNF167 (E3 ligase) were increased. Corresponding LMP2 and LMP7 proteins were down-regulated in mice, and RNF167 was elevated in diabetic mice. The reduced expression of LMP2 and LMP7, along with increased RNF167 expression, may contribute to the future cardiac deterioration commonly observed in diabetes. This study enhances our understanding of UPS imbalances in the hearts of diabetic mice and raises questions about the interplay between the UPS and other cellular processes, such as autophagy. Further exploration in this area could provide valuable insights into the mechanisms underlying diabetic heart complications and potential therapeutic targets.
Topics: Mice; Animals; Proteasome Endopeptidase Complex; Ubiquitin; Diabetes Mellitus, Type 2; Diabetes Mellitus, Experimental; Diabetic Cardiomyopathies; Diabetes Complications; RNA, Messenger
PubMed: 37895057
DOI: 10.3390/ijms242015376 -
Biochimica Et Biophysica Acta.... Jan 2024The ubiquitin-proteasome system (UPS) is a fundamental regulatory mechanism in cells, vital for maintaining cellular homeostasis, compiling signaling transduction, and... (Review)
Review
The ubiquitin-proteasome system (UPS) is a fundamental regulatory mechanism in cells, vital for maintaining cellular homeostasis, compiling signaling transduction, and determining cell fates. These biological processes require the coordinated signal cascades of UPS members, including ubiquitin ligases, ubiquitin-conjugating enzymes, deubiquitinases, and proteasomes, to ubiquitination and de-ubiquitination on substrates. Recent studies indicate that ubiquitination code rewriting is particularly prominent in pancreatic cancer. High frequency mutation or aberrant hyperexpression of UPS members dysregulates ferroptosis, tumor microenvironment, and metabolic rewiring processes and contribute to tumor growth, metastasis, immune evasion, and acquired drug resistance. We conduct an in-depth overview of ubiquitination process in pancreatic cancer, highlighting the role of ubiquitin code in tumor-promoting and tumor-suppressor pathways. Furthermore, we review current UPS modulators and analyze the potential of UPS modulators as cancer therapy.
Topics: Humans; Ubiquitin; Ubiquitin-Protein Ligases; Ubiquitination; Ubiquitin-Conjugating Enzymes; Pancreatic Neoplasms; Proteasome Endopeptidase Complex; Tumor Microenvironment
PubMed: 37704111
DOI: 10.1016/j.bbadis.2023.166884 -
Cell Reports Aug 2023The proteasome plays a central role in intracellular protein degradation. Age-dependent decline in proteasome activity is associated with cellular senescence and...
The proteasome plays a central role in intracellular protein degradation. Age-dependent decline in proteasome activity is associated with cellular senescence and organismal aging; however, the mechanism by which the proteasome plays a role in senescent cells remains elusive. Here, we show that nuclear foci that contain the proteasome and exhibit liquid-like properties are formed in senescent cells. The formation of senescence-associated nuclear proteasome foci (SANPs) is dependent on ubiquitination and RAD23B, similar to previously known nuclear proteasome foci, but also requires proteasome activity. RAD23B knockdown suppresses SANP formation and increases mitochondrial activity, leading to reactive oxygen species production without affecting other senescence traits such as cell-cycle arrest and cell morphology. These findings suggest that SANPs are an important feature of senescent cells and uncover a mechanism by which the proteasome plays a role in senescent cells.
Topics: Proteasome Endopeptidase Complex; Cell Nucleus; Ubiquitination; Cellular Senescence
PubMed: 37541257
DOI: 10.1016/j.celrep.2023.112880 -
Chemical Society Reviews Apr 2024Targeted protein degradation (TPD) has been established as a viable alternative to attenuate the function of a specific protein of interest in both biological and... (Review)
Review
Targeted protein degradation (TPD) has been established as a viable alternative to attenuate the function of a specific protein of interest in both biological and clinical contexts. The unique TPD mode-of-action has allowed previously undruggable proteins to become feasible targets, expanding the landscape of "druggable" properties and "privileged" target proteins. As TPD continues to evolve, a range of innovative strategies, which do not depend on recruiting E3 ubiquitin ligases as in proteolysis-targeting chimeras (PROTACs), have emerged. Here, we present an overview of direct lysosome- and proteasome-engaging modalities and discuss their perspectives, advantages, and limitations. We outline the chemical composition, biochemical activity, and pharmaceutical characteristics of each degrader. These alternative TPD approaches not only complement the first generation of PROTACs for intracellular protein degradation but also offer unique strategies for targeting pathologic proteins located on the cell membrane and in the extracellular space.
Topics: Proteolysis; Proteasome Endopeptidase Complex; Lysosomes; Cell Membrane; Ubiquitin-Protein Ligases
PubMed: 38369971
DOI: 10.1039/d3cs00344b -
Circulation Research Sep 2023A better understanding of the regulation of proteasome activities can facilitate the search for new therapeutic strategies. A cell culture study shows that PKA...
BACKGROUND
A better understanding of the regulation of proteasome activities can facilitate the search for new therapeutic strategies. A cell culture study shows that PKA (cAMP-dependent protein kinase or protein kinase A) activates the 26S proteasome by pS14-Rpn6 (serine14-phosphorylated Rpn6), but this discovery and its physiological significance remain to be established in vivo.
METHODS
Male and female mice with Ser14 of Rpn6 (regulatory particle non-ATPase 6) mutated to Ala (S14A [Rpn6/Psmd11]) or Asp (S14D) to respectively block or mimic pS14-Rpn6 were created and used along with cells derived from them. cAMP/PKA were manipulated pharmacologically. Ubiquitin-proteasome system functioning was evaluated with the GFPdgn (green fluorescence protein with carboxyl fusion of the CL1 degron) reporter mouse and proteasomal activity assays. Impact of S14A and S14D on proteotoxicity was tested in mice and cardiomyocytes overexpressing the misfolded protein R120G-CryAB (R120G [arginine120 to glycine missense mutant alpha B-crystallin]).
RESULTS
PKA activation increased pS14-Rpn6 and 26S proteasome activities in wild-type but not S14A embryonic fibroblasts (mouse embryonic fibroblasts), adult cardiomyocytes, and mouse hearts. Basal 26S proteasome activities were significantly greater in S14D myocardium and adult mouse cardiomyocytes than in wild-type counterparts. S14D::GFPdgn mice displayed significantly lower myocardial GFPdgn protein but not mRNA levels than GFPdgn mice. In R120G mice, a classic model of cardiac proteotoxicity, basal myocardial pS14-Rpn6 was significantly lower compared with nontransgenic littermates, which was not always associated with reduction of other phosphorylated PKA substrates. Cultured S14D neonatal cardiomyocytes displayed significantly faster proteasomal degradation of R120G than wild-type neonatal cardiomyocytes. Compared with R120G mice, S14D/S14D::R120G mice showed significantly greater myocardial proteasome activities, lower levels of total and K48-linked ubiquitin conjugates, and of aberrant CryAB (alpha B-crystallin) protein aggregates, less fetal gene reactivation, and cardiac hypertrophy, and delays in cardiac malfunction.
CONCLUSIONS
This study establishes in animals that pS14-Rpn6 mediates the activation of 26S proteasomes by PKA and that the reduced pS14-Rpn6 is a key pathogenic factor in cardiac proteinopathy, thereby identifying a new therapeutic target to reduce cardiac proteotoxicity.
Topics: Female; Male; Animals; Mice; Proteasome Endopeptidase Complex; alpha-Crystallin B Chain; Fibroblasts; Myocytes, Cardiac; Cyclic AMP-Dependent Protein Kinases; Ubiquitins
PubMed: 37641975
DOI: 10.1161/CIRCRESAHA.123.322887 -
Clinical and Translational Medicine Aug 2023CCN6 is a matricellular protein that critically regulates the tumourigenesis and progression of breast cancer. Although the tumour-suppressive function of CCN6 has been...
BACKGROUND
CCN6 is a matricellular protein that critically regulates the tumourigenesis and progression of breast cancer. Although the tumour-suppressive function of CCN6 has been extensively studied, molecular mechanisms regulating protein levels of CCN6 remain largely unclear. This study aims to investigate the regulation of CCN6 by ubiquitination and deubiquitinating enzymes (DUBs) in breast cancer.
METHODS
A screening assay was performed to identify OTUB1 as the DUB for CCN6. Various biochemical methods were applied to elucidate the molecular mechanism of OTUB1 in the regulation of CCN6. The role of OTUB1-CCN6 interaction in breast cancer was studied with cell experiments and the allograft model. The correlation of OTUB1 and CCN6 in human breast cancer was determined by immunohistochemistry and Western blot.
RESULTS
We found that CCN6 protein levels were controlled by the ubiquitin-proteasome system. The K48 ubiquitination and degradation of CCN6 was inhibited by OTUB1, which directly interacted with CCN6 through its linker domain. Furthermore, OTUB1 inhibited the ubiquitination of CCN6 in a non-canonical manner. Deletion of OTUB1, concomitant with reduced CCN6 abundance, increased the migration, proliferation and viability of breast cancer cells. Supplementation of CCN6 abolished the effect of OTUB1 deletion on breast cancer. Importantly, OTUB1 expression was downregulated in human breast cancer and positively correlated with CCN6 levels.
CONCLUSION
This study identified OTUB1 as a novel regulator of CCN6 in breast cancer.
Topics: Humans; Blotting, Western; Carcinogenesis; Cell Transformation, Neoplastic; Cytoplasm; Proteasome Endopeptidase Complex; Deubiquitinating Enzymes; CCN Intercellular Signaling Proteins
PubMed: 37608493
DOI: 10.1002/ctm2.1385 -
Molecular Cell Jan 2024The posttranslational modifier ubiquitin regulates most cellular processes. Its ability to form polymeric chains of distinct linkages is key to its diverse...
The posttranslational modifier ubiquitin regulates most cellular processes. Its ability to form polymeric chains of distinct linkages is key to its diverse functionality. Yet, we still lack the experimental tools to induce linkage-specific polyubiquitylation of a protein of interest in cells. Here, we introduce a set of engineered ubiquitin protein ligases and matching ubiquitin acceptor tags for the rapid, inducible linear (M1-), K48-, or K63-linked polyubiquitylation of proteins in yeast and mammalian cells. By applying the so-called "Ubiquiton" system to proteasomal targeting and the endocytic pathway, we validate this tool for soluble cytoplasmic and nuclear as well as chromatin-associated and integral membrane proteins and demonstrate how it can be used to control the localization and stability of its targets. We expect that the Ubiquiton system will serve as a versatile, broadly applicable research tool to explore the signaling functions of polyubiquitin chains in many biological contexts.
Topics: Animals; Ubiquitin; Ubiquitin-Protein Ligases; Polyubiquitin; Signal Transduction; Proteasome Endopeptidase Complex; Ubiquitination; Mammals
PubMed: 38103558
DOI: 10.1016/j.molcel.2023.11.016 -
Biomolecules Aug 2023The 26S proteasome is the largest and most complicated protease known, and changes to proteasome assembly or function contribute to numerous human diseases. Assembly of... (Review)
Review
The 26S proteasome is the largest and most complicated protease known, and changes to proteasome assembly or function contribute to numerous human diseases. Assembly of the 26S proteasome from its ~66 individual polypeptide subunits is a highly orchestrated process requiring the concerted actions of both intrinsic elements of proteasome subunits, as well as assistance by extrinsic, dedicated proteasome assembly chaperones. With the advent of near-atomic resolution cryo-electron microscopy, it has become evident that the proteasome is a highly dynamic machine, undergoing numerous conformational changes in response to ligand binding and during the proteolytic cycle. In contrast, an appreciation of the role of conformational dynamics during the biogenesis of the proteasome has only recently begun to emerge. Herein, we review our current knowledge of proteasome assembly, with a particular focus on how conformational dynamics guide particular proteasome biogenesis events. Furthermore, we highlight key emerging questions in this rapidly expanding area.
Topics: Proteasome Endopeptidase Complex; Protein Conformation; Models, Molecular; Molecular Chaperones; Humans; Cryoelectron Microscopy; Proteolysis; Ubiquitin
PubMed: 37627288
DOI: 10.3390/biom13081223 -
Frontiers in Immunology 2023Mutations in genes coding for proteasome subunits and/or proteasome assembly helpers typically cause recurring autoinflammation referred to as chronic atypical...
Mutations in genes coding for proteasome subunits and/or proteasome assembly helpers typically cause recurring autoinflammation referred to as chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperatures (CANDLE) or proteasome-associated autoinflammatory syndrome (PRAAS). Patients with CANDLE/PRAAS present with mostly chronically elevated type I interferon scores that emerge as a consequence of increased proteotoxic stress by mechanisms that are not fully understood. Here, we report on five unrelated patients with CANDLE/PRAAS carrying novel inherited proteasome missense and/or nonsense variants. Four patients were compound heterozygous for novel pathogenic variants in the known CANDLE/PRAAS associated genes, and , whereas one patient showed additive loss-of-function mutations in . Variants in two previously not associated proteasome genes, and , were found in a patient who also carried the founder mutation, p.T75M. All newly identified mutations substantially impact the steady-state expression of the affected proteasome subunits and/or their incorporation into mature 26S proteasomes. Our observations expand the spectrum of PRAAS-associated genetic variants and improve a molecular diagnosis and genetic counseling of patients with sterile autoinflammation.
Topics: Humans; Proteasome Endopeptidase Complex; Syndrome; Cytoplasm; Dermatitis
PubMed: 37600812
DOI: 10.3389/fimmu.2023.1190104 -
Human Molecular Genetics Dec 2023MeCP2 (Methyl CpG binding protein 2) is an intrinsically disordered protein that binds to methylated genome regions. The protein is a critical transcriptional regulator... (Review)
Review
MeCP2 (Methyl CpG binding protein 2) is an intrinsically disordered protein that binds to methylated genome regions. The protein is a critical transcriptional regulator of the brain, and its mutations account for 95% of Rett syndrome (RTT) cases. Early studies of this neurodevelopmental disorder revealed a close connection with dysregulations of the ubiquitin system (UbS), notably as related to UBE3A, a ubiquitin ligase involved in the proteasome-mediated degradation of proteins. MeCP2 undergoes numerous post-translational modifications (PTMs), including ubiquitination and sumoylation, which, in addition to the potential functional outcomes of their monomeric forms in gene regulation and synaptic plasticity, in their polymeric organization, these modifications play a critical role in proteasomal degradation. UbS-mediated proteasomal degradation is crucial in maintaining MeCP2 homeostasis for proper function and is involved in decreasing MeCP2 in some RTT-causing mutations. However, regardless of all these connections to UbS, the molecular details involved in the signaling of MeCP2 for its targeting by the ubiquitin-proteasome system (UPS) and the functional roles of monomeric MeCP2 ubiquitination and sumoylation remain largely unexplored and are the focus of this review.
Topics: Humans; Methyl-CpG-Binding Protein 2; Sumoylation; Proteasome Endopeptidase Complex; Rett Syndrome; Ubiquitination; Ubiquitin
PubMed: 37694858
DOI: 10.1093/hmg/ddad150