المؤلفون: Nelson, Samantha, Harris, Timothy, Muli, Christine, Maresh, Marianne, Baker, Braden, Smith, Chloe, Neumann, Chris, Parkinson, Elizabeth, Trader, Darci

المصدر: ChemBioChem. 25(3)

مصطلحات موضوعية: macrocycles, peptides, proteasome, stimulation, Humans, Proteasome Endopeptidase Complex, Peptides, Cyclic, Proteolysis, Proteins, Neurodegenerative Diseases

الوصف: The proteasome degrades proteins, which is essential for cellular homeostasis. Ubiquitin independent proteolysis degrades highly disordered and misfolded proteins. A decline of proteasomal activity has been associated with multiple neurodegenerative diseases due to the accumulation of misfolded proteins. In this work, cyclic peptide proteasome stimulators (CyPPSs) that enhance the clearance of misfolded proteins were discovered. In the initial screen of predicted natural products (pNPs), several cyclic peptides were found to stimulate the 20S core particle (20S CP). Development of a robust structural activity relationship led to the identification of potent, cell permeable CyPPSs. In vitro assays revealed that CyPPSs stimulate degradation of highly disordered and misfolded proteins without affecting ordered proteins. Furthermore, using a novel flow-based assay for proteasome activity, several CyPPSs were found to stimulate the 20S CP in cellulo. Overall, this work describes the development of CyPPSs as chemical tools capable of stimulating the proteasome and provides strong support for proteasome stimulation as a therapeutic strategy for neurodegenerative diseases.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/67h5d70rTest

المؤلفون: Grover, Manish, Gang, Spencer S, Troemel, Emily R, Barkoulas, Michalis

المصدر: PLOS Biology. 22(3)

مصطلحات موضوعية: Medical Microbiology, Biomedical and Clinical Sciences, Biological Sciences, Infectious Diseases, Prevention, Emerging Infectious Diseases, Vaccine Related, Genetics, Aetiology, 2.1 Biological and endogenous factors, Infection, Animals, Caenorhabditis elegans, Transcription Factors, DNA-Binding Proteins, Proteasome Endopeptidase Complex, Caenorhabditis elegans Proteins, Immunity, Innate, Bacterial Infections, Agricultural and Veterinary Sciences, Medical and Health Sciences, Developmental Biology, Agricultural, veterinary and food sciences, Biological sciences, Biomedical and clinical sciences

الوصف: Protein quality control pathways play important roles in resistance against pathogen infection. For example, the conserved transcription factor SKN-1/NRF up-regulates proteostasis capacity after blockade of the proteasome and also promotes resistance against bacterial infection in the nematode Caenorhabditis elegans. SKN-1/NRF has 3 isoforms, and the SKN-1A/NRF1 isoform, in particular, regulates proteasomal gene expression upon proteasome dysfunction as part of a conserved bounce-back response. We report here that, in contrast to the previously reported role of SKN-1 in promoting resistance against bacterial infection, loss-of-function mutants in skn-1a and its activating enzymes ddi-1 and png-1 show constitutive expression of immune response programs against natural eukaryotic pathogens of C. elegans. These programs are the oomycete recognition response (ORR), which promotes resistance against oomycetes that infect through the epidermis, and the intracellular pathogen response (IPR), which promotes resistance against intestine-infecting microsporidia. Consequently, skn-1a mutants show increased resistance to both oomycete and microsporidia infections. We also report that almost all ORR/IPR genes induced in common between these programs are regulated by the proteasome and interestingly, specific ORR/IPR genes can be induced in distinct tissues depending on the exact trigger. Furthermore, we show that increasing proteasome function significantly reduces oomycete-mediated induction of multiple ORR markers. Altogether, our findings demonstrate that proteasome regulation keeps innate immune responses in check in a tissue-specific manner against natural eukaryotic pathogens of the C. elegans epidermis and intestine.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/0nw7p2xxTest

المؤلفون: Lam, Breanna, Lam, Darren, Le, Tiffany, Kao, Andy, Slaiwa, Yousif, Hampton, Randolph, Flagg, Matthew

المصدر: Molecular Biology of the Cell. 34(13)

مصطلحات موضوعية: Humans, Cystic Fibrosis Transmembrane Conductance Regulator, Cystic Fibrosis, Ubiquitin, Proteasome Endopeptidase Complex, Protein Processing, Post-Translational, Mutation

الوصف: In both health and disease, the ubiquitin-proteasome system (UPS) degrades point mutants that retain partial function but have decreased stability compared with their wild-type counterparts. This class of UPS substrate includes routine translational errors and numerous human disease alleles, such as the most common cause of cystic fibrosis, ΔF508-CFTR. Yet, there is no systematic way to discover novel examples of these minimally misfolded substrates. To address that shortcoming, we designed a genetic screen to isolate functional-but-degraded point mutants, and we used the screen to study soluble, monomeric proteins with known structures. These simple parent proteins yielded diverse substrates, allowing us to investigate the structural features, cytotoxicity, and small-molecule regulation of minimal misfolding. Our screen can support numerous lines of inquiry, and it provides broad access to a class of poorly understood but biomedically critical quality-control substrates.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/27c1z789Test

المؤلفون: Zhang, Ruizhu, Pan, Shuxian, Zheng, Suya, Liao, Qingqing, Jiang, Zhaodi, Wang, Dixian, Li, Xuemei, Hu, Ao, Li, Xinran, Zhu, Yezhang, Shen, Xiaoqi, Lei, Jing, Zhong, Siming, Zhang, Xiaomei, Huang, Lingyun, Wang, Xiaorong, Shen, Li, Song, Bao-Liang, Zhao, Jing-Wei, Wang, Zhiping, Yang, Bing, Guo, Xing, Huang, Lan

المصدر: Science Advances. 9(48)

مصطلحات موضوعية: Humans, Animals, Mice, Proteasome Endopeptidase Complex, Membrane Proteins, Proteostasis, Endoplasmic Reticulum-Associated Degradation, Mice, Nude, Lipids

الوصف: Protein degradation in eukaryotic cells is mainly carried out by the 26S proteasome, a macromolecular complex not only present in the cytosol and nucleus but also associated with various membranes. How proteasomes are anchored to the membrane and the biological meaning thereof have been largely unknown in higher organisms. Here, we show that N-myristoylation of the Rpt2 subunit is a general mechanism for proteasome-membrane interaction. Loss of this modification in the Rpt2-G2A mutant cells leads to profound changes in the membrane-associated proteome, perturbs the endomembrane system, and undermines critical cellular processes such as cell adhesion, endoplasmic reticulum-associated degradation and membrane protein trafficking. Rpt2G2A/G2A homozygous mutation is embryonic lethal in mice and is sufficient to abolish tumor growth in a nude mice xenograft model. These findings have defined an evolutionarily conserved mechanism for maintaining membrane protein homeostasis and underscored the significance of compartmentalized protein degradation by myristoyl-anchored proteasomes in health and disease.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/9h60k1j7Test

المؤلفون: Ribeiro, Felipe, Cozachenco, Danielle, Heimfarth, Luana, Fortuna, Juliana, de Freitas, Guilherme, de Sousa, Jorge, Alves-Leon, Soniza, Leite, Renata, Suemoto, Claudia, De Felice, Fernanda, Lourenco, Mychael, Ferreira, Sergio, Grinberg, Lea

المصدر: Communications Biology. 6(1)

مصطلحات موضوعية: Mice, Animals, Alzheimer Disease, Amyloid beta-Peptides, Proteasome Endopeptidase Complex, Neuronal Plasticity, Memory Disorders

الوصف: The proteasome plays key roles in synaptic plasticity and memory by regulating protein turnover, quality control, and elimination of oxidized/misfolded proteins. Here, we investigate proteasome function and localization at synapses in Alzheimers disease (AD) post-mortem brain tissue and in experimental models. We found a marked increase in ubiquitinylated proteins in post-mortem AD hippocampi compared to controls. Using several experimental models, we show that amyloid-β oligomers (AβOs) inhibit synaptic proteasome activity and trigger a reduction in synaptic proteasome content. We further show proteasome inhibition specifically in hippocampal synaptic fractions derived from APPswePS1ΔE9 mice. Reduced synaptic proteasome activity instigated by AβOs is corrected by treatment with rolipram, a phosphodiesterase-4 inhibitor, in mice. Results further show that dynein inhibition blocks AβO-induced reduction in dendritic proteasome content in hippocampal neurons. Finally, proteasome inhibition induces AD-like pathological features, including reactive oxygen species and dendritic spine loss in hippocampal neurons, inhibition of hippocampal mRNA translation, and memory impairment in mice. Results suggest that proteasome inhibition may contribute to synaptic and memory deficits in AD.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/3208p37pTest

المؤلفون: Gressler, A, Zinecker, Heidi, Simon, Anna, Leng, Houfu

مصطلحات موضوعية: Aging, Degradation, Inflamm-aging, Proteostasis, T cell, Translation, Humans, Aged, Proteostasis, T-Cell Senescence, Aging, Proteasome Endopeptidase Complex, Autophagy

الوصف: Aging leads to a decline in immune cell function, which leaves the organism vulnerable to infections and age-related multimorbidities. One major player of the adaptive immune response are T cells, and recent studies argue for a major role of disturbed proteostasis contributing to reduced function of these cells upon aging. Proteostasis refers to the state of a healthy, balanced proteome in the cell and is influenced by synthesis (translation), maintenance and quality control of proteins, as well as degradation of damaged or unwanted proteins by the proteasome, autophagy, lysosome and cytoplasmic enzymes. This review focuses on molecular processes impacting on proteostasis in T cells, and specifically functional or quantitative changes of each of these upon aging. Importantly, we describe the biological consequences of compromised proteostasis in T cells, which range from impaired T cell activation and function to enhancement of inflamm-aging by aged T cells. Finally, approaches to improve proteostasis and thus rejuvenate aged T cells through pharmacological or physical interventions are discussed.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/8vg2414cTest

المؤلفون: Balmanno, Kathryn, Kidger, Andrew M, Byrne, Dominic P, Sale, Matthew J, Nassman, Nejma, Eyers, Patrick A, Cook, Simon J

المصدر: Biochemical Journal. 480(9)

مصطلحات موضوعية: Biochemistry and Cell Biology, Biological Sciences, Prevention, 5.1 Pharmaceuticals, Development of treatments and therapeutic interventions, Phosphorylation, MAP Kinase Signaling System, Proteasome Endopeptidase Complex, Protein Processing, Post-Translational, Ubiquitination, ERK inhibitors, MEK, RAF, RAS, extracellular signal-regulated kinases, ubiquitin proteasome system, Chemical Sciences, Medical and Health Sciences, Biochemistry & Molecular Biology, Biochemistry and cell biology

الوصف: Innate or acquired resistance to small molecule BRAF or MEK1/2 inhibitors (BRAFi or MEKi) typically arises through mechanisms that sustain or reinstate ERK1/2 activation. This has led to the development of a range of ERK1/2 inhibitors (ERKi) that either inhibit kinase catalytic activity (catERKi) or additionally prevent the activating pT-E-pY dual phosphorylation of ERK1/2 by MEK1/2 (dual-mechanism or dmERKi). Here, we show that eight different ERKi (both catERKi or dmERKi) drive the turnover of ERK2, the most abundant ERK isoform, with little or no effect on ERK1. Thermal stability assays show that ERKi do not destabilise ERK2 (or ERK1) in vitro, suggesting that ERK2 turnover is a cellular consequence of ERKi binding. ERK2 turnover is not observed upon treatment with MEKi alone, suggesting it is ERKi binding to ERK2 that drives ERK2 turnover. However, MEKi pre-treatment, which blocks ERK2 pT-E-pY phosphorylation and dissociation from MEK1/2, prevents ERK2 turnover. ERKi treatment of cells drives the poly-ubiquitylation and proteasome-dependent turnover of ERK2 and pharmacological or genetic inhibition of Cullin-RING E3 ligases prevents this. Our results suggest that ERKi, including current clinical candidates, act as 'kinase degraders', driving the proteasome-dependent turnover of their major target, ERK2. This may be relevant to the suggestion of kinase-independent effects of ERK1/2 and the therapeutic use of ERKi.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/5s95c7vqTest

المؤلفون: King, Elizabeth, Cho, Yoojin, Hsu, Nathan, Dovala, Dustin, McKenna, Jeffrey, Tallarico, John, Schirle, Markus, Nomura, Daniel

المصدر: Cell chemical biology. 30(4)

مصطلحات موضوعية: E2 ligase, NFKB1, UBE2D, activity-based protein profiling, molecular glue, targeted protein degradation, transcription factor, NF-kappa B, Ligands, Proteomics, Proteolysis, Proteasome Endopeptidase Complex, Ubiquitin-Protein Ligases

الوصف: Targeted protein degradation has arisen as a powerful therapeutic modality for degrading disease targets. While proteolysis-targeting chimera (PROTAC) design is more modular, the discovery of molecular glue degraders has been more challenging. Here, we have coupled the phenotypic screening of a covalent ligand library with chemoproteomic approaches to rapidly discover a covalent molecular glue degrader and associated mechanisms. We have identified a cysteine-reactive covalent ligand EN450 that impairs leukemia cell viability in a NEDDylation and proteasome-dependent manner. Chemoproteomic profiling revealed covalent interaction of EN450 with an allosteric C111 in the E2 ubiquitin-conjugating enzyme UBE2D. Quantitative proteomic profiling revealed the degradation of the oncogenic transcription factor NFKB1 as a putative degradation target. Our study thus puts forth the discovery of a covalent molecular glue degrader that uniquely induced the proximity of an E2 with a transcription factor to induce its degradation in cancer cells.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/513376ddTest

المؤلفون: Almaliti, Jehad, Fajtová, Pavla, Calla, Jaeson, LaMonte, Gregory M, Feng, Mudong, Rocamora, Frances, Ottilie, Sabine, Glukhov, Evgenia, Boura, Evzen, Suhandynata, Raymond T, Momper, Jeremiah D, Gilson, Michael K, Winzeler, Elizabeth A, Gerwick, William H, O'Donoghue, Anthony J

المصدر: Chemistry - A European Journal. 29(20)

مصطلحات موضوعية: Chemical Sciences, Orphan Drug, Rare Diseases, Vector-Borne Diseases, Infectious Diseases, Malaria, 5.1 Pharmaceuticals, Development of treatments and therapeutic interventions, Infection, Good Health and Well Being, Mice, Animals, Humans, Proteasome Inhibitors, Proteasome Endopeptidase Complex, Plasmodium, Plasmodium falciparum, Antimalarials, epoxyketone, inhibition, malaria, plasmodium, proteasome, General Chemistry, Chemical sciences

الوصف: Here, we present remarkable epoxyketone-based proteasome inhibitors with low nanomolar in vitro potency for blood-stage Plasmodium falciparum and low cytotoxicity for human cells. Our best compound has more than 2,000-fold greater selectivity for erythrocytic-stage P. falciparum over HepG2 and H460 cells, which is largely driven by the accommodation of the parasite proteasome for a D-amino acid in the P3 position and the preference for a difluorobenzyl group in the P1 position. We isolated the proteasome from P. falciparum cell extracts and determined that the best compound is 171-fold more potent at inhibiting the β5 subunit of P. falciparum proteasome when compared to the same subunit of the human constitutive proteasome. These compounds also significantly reduce parasitemia in a P. berghei mouse infection model and prolong survival of animals by an average of 6 days. The current epoxyketone inhibitors are ideal starting compounds for orally bioavailable anti-malarial drugs.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/6x99c667Test

المؤلفون: Chua, Bernadette A, Lennan, Connor J, Sunshine, Mary Jean, Dreifke, Daniela, Chawla, Ashu, Bennett, Eric J, Signer, Robert AJ

المصدر: Cell Stem Cell. 30(4)

مصطلحات موضوعية: Biochemistry and Cell Biology, Biological Sciences, Stem Cell Research, Aging, Regenerative Medicine, Aetiology, 2.1 Biological and endogenous factors, Underpinning research, 1.1 Normal biological development and functioning, Proteostasis, Macroautophagy, Proteasome Endopeptidase Complex, Autophagy, Transcription Factors, Hematopoietic Stem Cells, Bag3, aggrephagy, aggresome, aging, autophagy, hematopoietic stem cell, proteasome, protein degradation, proteostasis, stem cell, Medical and Health Sciences, Developmental Biology, Biological sciences, Biomedical and clinical sciences

الوصف: Hematopoietic stem cells (HSCs) regenerate blood cells throughout life. To preserve their fitness, HSCs are particularly dependent on maintaining protein homeostasis (proteostasis). However, how HSCs purge misfolded proteins is unknown. Here, we show that in contrast to most cells that primarily utilize the proteasome to degrade misfolded proteins, HSCs preferentially traffic misfolded proteins to aggresomes in a Bag3-dependent manner and depend on aggrephagy, a selective form of autophagy, to maintain proteostasis in vivo. When autophagy is disabled, HSCs compensate by increasing proteasome activity, but proteostasis is ultimately disrupted as protein aggregates accumulate and HSC function is impaired. Bag3-deficiency blunts aggresome formation in HSCs, resulting in protein aggregate accumulation, myeloid-biased differentiation, and diminished self-renewal activity. Furthermore, HSC aging is associated with a severe loss of aggresomes and reduced autophagic flux. Protein degradation pathways are thus specifically configured in young adult HSCs to preserve proteostasis and fitness but become dysregulated during aging.

وصف الملف: application/pdf

الوصول الحر: https://escholarship.org/uc/item/6jz57666Test