High resolution profiling of cell cycle-dependent protein and phosphorylation abundance changes in non-transformed cells

Nature Communications, Год журнала: 2025, Номер 16(1)

Опубликована: Март 16, 2025

The cell cycle governs a precise series of molecular events, regulated by coordinated changes in protein and phosphorylation abundance, that culminates the generation two daughter cells. Here, we present proteomic phosphoproteomic analysis human hTERT-RPE-1 cells using deep quantitative mass spectrometry isobaric labelling. By analysing non-transformed improving temporal resolution coverage key regulators, dataset cycle-dependent site oscillation offers foundational reference for investigating regulation. These data reveal regulatory intricacies including proteins sites exhibiting oscillation, targeted degradation during mitotic exit. Integrated with complementary resources, our link abundance dynamics to functional are accessible through Cell Cycle database (CCdb), an interactive web-based resource community. authors nontransformed

Язык: Английский

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Defining E3 ligase–substrate relationships through multiplex CRISPR screening

Nature Cell Biology, Год журнала: 2023, Номер 25(10), С. 1535 - 1545

Опубликована: Сен. 21, 2023

Specificity within the ubiquitin-proteasome system is primarily achieved through E3 ubiquitin ligases, but for many E3s their substrates-and in particular molecular features (degrons) that they recognize-remain largely unknown. Current approaches assigning to cognate substrates are tedious and low throughput. Here we developed a multiplex CRISPR screening platform assign ligases at scale. A proof-of-principle screen successfully performed ~100 screens single experiment, refining known C-degron pathways identifying an additional pathway which Cul2

Язык: Английский

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DEGRONOPEDIA: a web server for proteome-wide inspection of degrons

Nucleic Acids Research, Год журнала: 2024, Номер 52(W1), С. W221 - W232

Опубликована: Апрель 3, 2024

Abstract E3 ubiquitin ligases recognize substrates through their short linear motifs termed degrons. While degron-signaling has been a subject of extensive study, resources for its systematic screening are limited. To bridge this gap, we developed DEGRONOPEDIA, web server that searches degrons and maps them to nearby residues can undergo ubiquitination disordered regions, which may act as protein unfolding seeds. Along with an evolutionary assessment degron conservation, the also reports on post-translational modifications mutations modulate availability. Acknowledging prevalence at termini, DEGRONOPEDIA incorporates machine learning assess N-/C-terminal stability, supplemented by simulations proteolysis identify in newly formed termini. An experimental validation predicted C-terminal destabilizing motif, coupled confirmation post-proteolytic another case, exemplifies practical application. be freely accessed degronopedia.com.

Язык: Английский

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PRKN-linked familial Parkinson’s disease: cellular and molecular mechanisms of disease-linked variants

Cellular and Molecular Life Sciences, Год журнала: 2024, Номер 81(1)

Опубликована: Май 20, 2024

Abstract Parkinson’s disease (PD) is a common and incurable neurodegenerative disorder that arises from the loss of dopaminergic neurons in substantia nigra mainly characterized by progressive motor function. Monogenic familial PD associated with highly penetrant variants specific genes, notably PRKN gene, where homozygous or compound heterozygous loss-of-function predominate. encodes Parkin, an E3 ubiquitin-protein ligase important for protein ubiquitination mitophagy damaged mitochondria. Accordingly, Parkin plays central role mitochondrial quality control but itself also subject to strict system rapidly eliminates certain disease-linked variants. Here, we summarize cellular molecular functions highlighting various mechanisms which gene result loss-of-function. We emphasize importance high-throughput assays computational tools clinical classification how detailed insights into pathogenic may impact development personalized therapeutics.

Язык: Английский

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Targeted protein degradation in CNS disorders: a promising route to novel therapeutics?

Frontiers in Molecular Neuroscience, Год журнала: 2024, Номер 17

Опубликована: Апрель 15, 2024

Targeted protein degradation (TPD) is a rapidly expanding field, with various PROTACs (proteolysis-targeting chimeras) in clinical trials and molecular glues such as immunomodulatory imide drugs (IMiDs) already well established the treatment of certain blood cancers. Many current approaches are focused on oncology targets, leaving numerous potential applications underexplored. Targeting proteins for offers novel therapeutic route targets whose inhibition remains challenging, aggregates neurodegenerative diseases. This mini review focuses prospect utilizing TPD disease particularly PROTAC glue formats opportunities CNS E3 ligases. Some key challenges modalities including design degrader molecules, drug delivery brain barrier penetrance will be discussed.

Язык: Английский

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Mechanism of Ψ-Pro/C-degron recognition by the CRL2FEM1B ubiquitin ligase

Nature Communications, Год журнала: 2024, Номер 15(1)

Опубликована: Апрель 26, 2024

Abstract The E3 ligase-degron interaction determines the specificity of ubiquitin‒proteasome system. We recently discovered that FEM1B, a substrate receptor Cullin 2-RING ligase (CRL2), recognizes C-degrons containing C-terminal proline. By solving several cryo-EM structures CRL2 FEM1B bound to different C-degrons, we elucidate dimeric assembly complex. Furthermore, reveal distinct dimerization states unmodified and neddylated uncover NEDD8-mediated activation mechanism . Our research also indicates that, utilizes bipartite recognize both proline an upstream aromatic residue within substrate. These structural findings, complemented by in vitro ubiquitination vivo cell-based assays, demonstrate -mediated polyubiquitination subsequent protein turnover depend on FEM1B-degron interactions state Overall, this study deepens our molecular understanding how Cullin-RING selection mediates turnover.

Язык: Английский

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Dipeptidyl peptidases and E3 ligases of N-degron pathways cooperate to regulate protein stability

The Journal of Cell Biology, Год журнала: 2024, Номер 223(8)

Опубликована: Июнь 14, 2024

N-degrons are short sequences located at protein N-terminus that mediate the interaction of E3 ligases (E3s) with substrates to promote their proteolysis. It is well established can be exposed following protease cleavage allow recognition by E3s. However, our knowledge regarding how proteases and E3s cooperate in quality control mechanisms remains minimal. Using a systematic approach monitor stability an N-terminome library, we found proline residue third N-terminal position (hereafter “P+3”) promotes instability. Genetic perturbations identified dipeptidyl peptidases DPP8 DPP9 primary N-degron pathways, UBR proteins, as regulators P+3 bearing substrate turnover. Interestingly, significantly enriched for secretory proteins. We proteins relying on signal peptide (SP) targeting contain “built-in” within SP. This degron becomes DPP8/9 upon translocation failure designated compartments, thus enabling clearance mislocalized UBRs maintain proteostasis.

Язык: Английский

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Recognition of BACH1 quaternary structure degrons by two F-box proteins under oxidative stress

Cell, Год журнала: 2024, Номер unknown

Опубликована: Ноя. 1, 2024

Highlights•FBXO22 and FBXL17 play non-redundant roles in regulating the stability of BACH1•FBXO22 binds a BACH1 quaternary structure degron presented by its dimeric BTB fold•S-nitrosylation destabilizes domain dimer prevents FBXO22 binding•Destabilized is recognized remodeled pair FBXL17SummaryUbiquitin-dependent proteolysis regulates diverse cellular functions with high substrate specificity, which hinges on ability ubiquitin E3 ligases to decode targets' degradation signals, i.e., degrons. Here, we show that BACH1, transcription repressor antioxidant response genes, features two distinct unconventional degrons encrypted homodimeric domain. These are both functionalized oxidative stress deciphered complementary E3s. recognizes constructed interface, unmasked from transcriptional co-repressors after releases chromatin. When this impaired oxidation, second manifested destabilized probed proteins remodels into E3-bound monomers for ubiquitination. Our findings highlight multidimensionality protein signals functional complementarity different targeting same substrate.Graphical abstract

Язык: Английский

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A quantitative intracellular peptide binding assay reveals recognition determinants and context dependence of short linear motifs

Journal of Biological Chemistry, Год журнала: 2025, Номер unknown, С. 108225 - 108225

Опубликована: Янв. 1, 2025

Язык: Английский

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Phosphodegrons in Health and Disease: From Cellular Homeostasis to Therapeutic Potential

Kinases and Phosphatases, Год журнала: 2025, Номер 3(1), С. 3 - 3

Опубликована: Фев. 6, 2025

Phosphodegrons are critical motifs that play a pivotal role in the regulation of protein stability and function via phosphorylation-dependent signaling pathways. These serve as recognition elements for ubiquitin ligases, facilitating targeted degradation proteins. By modulating key cellular processes such cell cycle progression, DNA repair, apoptosis, phosphodegrons essential maintaining homeostasis. Dysregulation has been implicated wide range diseases, including cancer neurodegenerative disorders, highlighting their potential therapeutic targets. This review provides an overview phosphodegron functions along with biological significance health disease. Additionally, we discuss current methodologies studying explore emerging trends identification targeting. synthesizing recent advances field, this article aims to offer insights into future directions challenges research, ultimately underscoring importance disease pathology.

Язык: Английский

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