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Science (New York, N.Y.) Aug 2023Cells use ubiquitin to mark proteins for proteasomal degradation. Although the proteasome also eliminates proteins that are not ubiquitinated, how this occurs...
Cells use ubiquitin to mark proteins for proteasomal degradation. Although the proteasome also eliminates proteins that are not ubiquitinated, how this occurs mechanistically is unclear. Here, we found that midnolin promoted the destruction of many nuclear proteins, including transcription factors encoded by the immediate-early genes. Diverse stimuli induced midnolin, and its overexpression was sufficient to cause the degradation of its targets by a mechanism that did not require ubiquitination. Instead, midnolin associated with the proteasome via an α helix, used its Catch domain to bind a region within substrates that can form a β strand, and used a ubiquitin-like domain to promote substrate destruction. Thus, midnolin contains three regions that function in concert to target a large set of nuclear proteins to the proteasome for degradation.
Topics: Nuclear Proteins; Proteasome Endopeptidase Complex; Ubiquitin; Ubiquitination; Proteolysis; Genes, Immediate-Early; HEK293 Cells; NIH 3T3 Cells; Transcription, Genetic
PubMed: 37616343
DOI: 10.1126/science.adh5021 -
Trends in Molecular Medicine Aug 2023Ubiquitin-proteasome system (UPS) is a selective proteolytic system that is associated with the expression or function of target proteins and participates in various... (Review)
Review
Ubiquitin-proteasome system (UPS) is a selective proteolytic system that is associated with the expression or function of target proteins and participates in various physiological and pathological processes of breast cancer. Inhibitors targeting the 26S proteasome in combination with other drugs have shown promising therapeutic effects in the clinical treatment of breast cancer. Moreover, several inhibitors/stimulators targeting other UPS components are also effective in preclinical studies, but have not yet been applied in the clinical treatment of breast cancer. Therefore, it is vital to comprehensively understand the functions of ubiquitination in breast cancer and to identify potential tumor promoters or tumor suppressors among UPS family members, with the aim of developing more effective and specific inhibitors/stimulators targeting specific components of this system.
Topics: Humans; Female; Ubiquitin; Proteasome Endopeptidase Complex; Breast Neoplasms; Ubiquitination
PubMed: 37328395
DOI: 10.1016/j.molmed.2023.05.006 -
Trends in Cell Biology Dec 2023Ferroptosis is the type of cell death arising from uncontrolled and excessive lipid peroxidation. NADPH is essential for ferroptosis regulation because it supplies... (Review)
Review
Ferroptosis is the type of cell death arising from uncontrolled and excessive lipid peroxidation. NADPH is essential for ferroptosis regulation because it supplies reducing equivalents for antioxidant defense systems and contributes to the generation of reactive oxygen species. Moreover, NADPH level serves as a biomarker for predicting the sensitivity of cells to ferroptosis. The ubiquitin-proteasome system governs the stability of many ferroptosis effectors. Recent research has revealed MARCHF6, the endoplasmic reticulum ubiquitin ligase, as an unprecedented NADPH sensor in the ubiquitin system and a critical regulator of ferroptosis involved in tumorigenesis and fetal development. This review summarizes the current understanding of NADPH metabolism and the ubiquitin-proteasome system in regulating ferroptosis and highlights the emerging importance of MARCHF6 as a vital connector between NADPH metabolism and ferroptosis.
Topics: Humans; Ferroptosis; Proteasome Endopeptidase Complex; NADP; Ubiquitin; Cell Death; Lipid Peroxidation; Reactive Oxygen Species
PubMed: 37558595
DOI: 10.1016/j.tcb.2023.07.003 -
Nature Mar 2024Targeted protein degradation is a pharmacological modality that is based on the induced proximity of an E3 ubiquitin ligase and a target protein to promote target...
Targeted protein degradation is a pharmacological modality that is based on the induced proximity of an E3 ubiquitin ligase and a target protein to promote target ubiquitination and proteasomal degradation. This has been achieved either via proteolysis-targeting chimeras (PROTACs)-bifunctional compounds composed of two separate moieties that individually bind the target and E3 ligase, or via molecular glues that monovalently bind either the ligase or the target. Here, using orthogonal genetic screening, biophysical characterization and structural reconstitution, we investigate the mechanism of action of bifunctional degraders of BRD2 and BRD4, termed intramolecular bivalent glues (IBGs), and find that instead of connecting target and ligase in trans as PROTACs do, they simultaneously engage and connect two adjacent domains of the target protein in cis. This conformational change 'glues' BRD4 to the E3 ligases DCAF11 or DCAF16, leveraging intrinsic target-ligase affinities that do not translate to BRD4 degradation in the absence of compound. Structural insights into the ternary BRD4-IBG1-DCAF16 complex guided the rational design of improved degraders of low picomolar potency. We thus introduce a new modality in targeted protein degradation, which works by bridging protein domains in cis to enhance surface complementarity with E3 ligases for productive ubiquitination and degradation.
Topics: Bromodomain Containing Proteins; Cell Cycle Proteins; Drug Design; Proteasome Endopeptidase Complex; Proteolysis; Proteolysis Targeting Chimera; Transcription Factors; Ubiquitin-Protein Ligases; Ubiquitination; Protein Binding; Substrate Specificity; Protein Domains
PubMed: 38383787
DOI: 10.1038/s41586-024-07089-6 -
Cell Feb 2024Oocytes are among the longest-lived cells in the body and need to preserve their cytoplasm to support proper embryonic development. Protein aggregation is a major threat...
Oocytes are among the longest-lived cells in the body and need to preserve their cytoplasm to support proper embryonic development. Protein aggregation is a major threat for intracellular homeostasis in long-lived cells. How oocytes cope with protein aggregation during their extended life is unknown. Here, we find that mouse oocytes accumulate protein aggregates in specialized compartments that we named endolysosomal vesicular assemblies (ELVAs). Combining live-cell imaging, electron microscopy, and proteomics, we found that ELVAs are non-membrane-bound compartments composed of endolysosomes, autophagosomes, and proteasomes held together by a protein matrix formed by RUFY1. Functional assays revealed that in immature oocytes, ELVAs sequester aggregated proteins, including TDP-43, and degrade them upon oocyte maturation. Inhibiting degradative activity in ELVAs leads to the accumulation of protein aggregates in the embryo and is detrimental for embryo survival. Thus, ELVAs represent a strategy to safeguard protein homeostasis in long-lived cells.
Topics: Animals; Female; Mice; Autophagosomes; Cytoplasmic Vesicles; Lysosomes; Oocytes; Proteasome Endopeptidase Complex; Protein Aggregates; Proteolysis
PubMed: 38382525
DOI: 10.1016/j.cell.2024.01.031 -
Annual Review of Pharmacology and... Jan 2024Thalidomide and its derivatives are powerful cancer therapeutics that are among the best-understood molecular glue degraders (MGDs). These drugs selectively reprogram... (Review)
Review
Thalidomide and its derivatives are powerful cancer therapeutics that are among the best-understood molecular glue degraders (MGDs). These drugs selectively reprogram the E3 ubiquitin ligase cereblon (CRBN) to commit target proteins for degradation by the ubiquitin-proteasome system. MGDs create novel recognition interfaces on the surface of the E3 ligase that engage in induced protein-protein interactions with neosubstrates. Molecular insight into their mechanism of action opens exciting opportunities to engage a plethora of targets through a specific recognition motif, the G-loop. Our analysis shows that current CRBN-based MGDs can in principle recognize over 2,500 proteins in the human proteome that contain a G-loop. We review recent advances in tuning the specificity between CRBN and its MGD-induced neosubstrates and deduce a set of simple rules that govern these interactions. We conclude that rational MGD design efforts will enable selective degradation of many more proteins, expanding this therapeutic modality to more disease areas.
Topics: Humans; Thalidomide; Proteolysis; Ubiquitin-Protein Ligases; Proteasome Endopeptidase Complex
PubMed: 37585660
DOI: 10.1146/annurev-pharmtox-022123-104147 -
Nature Communications Oct 2023Proteolysis-targeting chimera (PROTAC) and other targeted protein degradation (TPD) molecules that induce degradation by the ubiquitin-proteasome system (UPS) offer new...
Proteolysis-targeting chimera (PROTAC) and other targeted protein degradation (TPD) molecules that induce degradation by the ubiquitin-proteasome system (UPS) offer new opportunities to engage targets that remain challenging to be inhibited by conventional small molecules. One fundamental element in the degradation process is the E3 ligase. However, less than 2% amongst hundreds of E3 ligases in the human genome have been engaged in current studies in the TPD field, calling for the recruiting of additional ones to further enhance the therapeutic potential of TPD. To accelerate the development of PROTACs utilizing under-explored E3 ligases, we systematically characterize E3 ligases from seven different aspects, including chemical ligandability, expression patterns, protein-protein interactions (PPI), structure availability, functional essentiality, cellular location, and PPI interface by analyzing 30 large-scale data sets. Our analysis uncovers several E3 ligases as promising extant PROTACs. In total, combining confidence score, ligandability, expression pattern, and PPI, we identified 76 E3 ligases as PROTAC-interacting candidates. We develop a user-friendly and flexible web portal ( https://hanlaboratory.com/E3Atlas/ ) aimed at assisting researchers to rapidly identify E3 ligases with promising TPD activities against specifically desired targets, facilitating the development of these therapies in cancer and beyond.
Topics: Humans; Ubiquitin-Protein Ligases; Proteasome Endopeptidase Complex; Proteolysis; Ubiquitination; Neoplasms
PubMed: 37845222
DOI: 10.1038/s41467-023-42233-2 -
Nature Cell Biology Oct 2023Specificity within the ubiquitin-proteasome system is primarily achieved through E3 ubiquitin ligases, but for many E3s their substrates-and in particular the molecular...
Specificity within the ubiquitin-proteasome system is primarily achieved through E3 ubiquitin ligases, but for many E3s their substrates-and in particular the molecular features (degrons) that they recognize-remain largely unknown. Current approaches for assigning E3s to their cognate substrates are tedious and low throughput. Here we developed a multiplex CRISPR screening platform to assign E3 ligases to their cognate substrates at scale. A proof-of-principle multiplex screen successfully performed ~100 CRISPR screens in a single experiment, refining known C-degron pathways and identifying an additional pathway through which Cul2 targets C-terminal proline. Further, by identifying substrates for Cul1, Cul2, Cul3, Cul3, Cul3 and Cul3, we demonstrate that the approach is compatible with pools of full-length protein substrates of varying stabilities and, when combined with site-saturation mutagenesis, can assign E3 ligases to their cognate degron motifs. Thus, multiplex CRISPR screening will accelerate our understanding of how specificity is achieved within the ubiquitin-proteasome system.
Topics: Ubiquitin-Protein Ligases; Proteasome Endopeptidase Complex; Clustered Regularly Interspaced Short Palindromic Repeats; Ubiquitin
PubMed: 37735597
DOI: 10.1038/s41556-023-01229-2 -
Nature Reviews. Drug Discovery Feb 2024Targeted protein degradation (TPD) has emerged in the past decade as a major new drug modality to remove intracellular proteins with bispecific small molecules that... (Review)
Review
Targeted protein degradation (TPD) has emerged in the past decade as a major new drug modality to remove intracellular proteins with bispecific small molecules that recruit the protein of interest (POI) to an E3 ligase for degradation in the proteasome. Unlike classic occupancy-based drugs, intracellular TPD (iTPD) eliminates the target and works catalytically, and so can be more effective and sustained, with lower dose requirements. Recently, this approach has been expanded to the extracellular proteome, including both secreted and membrane proteins. Extracellular targeted protein degradation (eTPD) uses bispecific antibodies, conjugates or small molecules to degrade extracellular POIs by trafficking them to the lysosome for degradation. Here, we focus on recent advances in eTPD, covering degrader systems, targets, molecular designs and parameters to advance them. Now almost any protein, intracellular or extracellular, is addressable in principle with TPD.
Topics: Humans; Proteolysis; Proteasome Endopeptidase Complex; Membrane Proteins; Drug Discovery; Ubiquitin-Protein Ligases
PubMed: 38062152
DOI: 10.1038/s41573-023-00833-z -
Circulation Research Jan 2024Increasing evidence suggests that long noncoding RNAs play significant roles in vascular biology and disease development. One such long noncoding RNA, , has been...
BACKGROUND
Increasing evidence suggests that long noncoding RNAs play significant roles in vascular biology and disease development. One such long noncoding RNA, , has been implicated in the development of tumors. Nevertheless, the precise role of in cardiovascular diseases, particularly atherosclerosis, has not been thoroughly elucidated. Thus, the primary aim of this investigation is to assess the influence of on vascular inflammation and the initiation of atherosclerosis.
METHODS
We generated knockin and (Apolipoprotein E) knockout mice () and global and proteasome subunit-β type-9 () double knockout mice (). To explore the roles of and in atherosclerosis, we fed the mice with a Western diet for 12 weeks.
RESULTS
Long noncoding RNA is significantly elevated in human atherosclerotic plaques. Strikingly, mice exhibited increased atherosclerosis development, plaque vulnerability, and vascular inflammation compared with mice. Moreover, the levels of VCAM1 (vascular adhesion molecule 1) and ICAM1 (intracellular adhesion molecule 1) were significantly upregulated in atherosclerotic lesions and serum of mice. Consistently, in vitro gain- and loss-of-function studies demonstrated that induced monocyte/macrophage adhesion to endothelial cells and increased VCAM1 and ICAM1 levels in a PSMB9-dependent manner. Mechanistic studies revealed that induced transcription by recruiting the transcription factor NONO (non-POU domain-containing octamer-binding protein) and binding to the promoter. PSMB9 (proteasome subunit-β type-9) elevated VCAM1 and ICAM1 expression via the upregulation of ZEB1 (zinc finger E-box-binding homeobox 1). deficiency decreased atherosclerotic lesion size, plaque vulnerability, and vascular inflammation in mice in vivo. Importantly, endothelial overexpression of -increased atherosclerosis and vascular inflammation were attenuated by knockout.
CONCLUSIONS
promotes vascular inflammation and atherosclerosis via the NONOPSMB9/ZEB1 axis. Our findings support the development of new long noncoding RNA-based strategies to counteract atherosclerotic cardiovascular disease.
Topics: Animals; Humans; Mice; Apolipoproteins E; Atherosclerosis; Endothelial Cells; Inflammation; Mice, Inbred C57BL; Mice, Knockout; Plaque, Atherosclerotic; Proteasome Endopeptidase Complex; RNA, Long Noncoding
PubMed: 38084631
DOI: 10.1161/CIRCRESAHA.122.322360