Changes in cellular physiology and proteomic homeostasis accompanied the initiation and progression of colorectal cancer. Thus, ubiquitination represents a central regulatory mechanism in proteome dynamics. However, the complexity of the ubiquitinating network involved in carcinogenesis remains unclear. This study revealed the tumor-suppressive role of the ubiquitin ligase Cullin4A (CUL4A) in the intestine. We showed that simultaneous loss of CUL4A and hyperactivation of the Wnt pathway promotes tumor development in the distal colon. This tumor development is caused by an accumulation of the inactive SMAD3, a TGF-β pathway mediator. Depletion of CUL4A resulted in stabilization of HUWE1, which attenuated SMAD3 function. We showed a correlation between the intracellular localization of CUL4A and colorectal cancer progression, where nuclear CUL4A localization correlates with advanced colorectal cancer progression. In summary, we identified CUL4A as an important regulator of SMAD3 signal transduction competence in a HUWE1-dependent manner and demonstrated a critical role for the crosstalk between ubiquitination and the Wnt/TGF-β signaling pathways in gastrointestinal homeostasis.

Breast cancer (BC) is the most common malignant tumor and the leading cause of cancer-related deaths among women worldwide. Great progress has been recently achieved in controlling breast cancer; however, mortality from breast cancer remains a substantial challenge, and new treatment mechanisms are being actively sought. Programmed cell death (PCD) is associated with the progression and treatment of many types of human cancers. PCD can be divided into multiple pathways including autophagy, apoptosis, mitotic catastrophe, necroptosis, ferroptosis, pyroptosis, and anoikis. Ubiquitination is a post-translational modification process in which ubiquitin, a 76-amino acid protein, is coupled to the lysine residues of other proteins. Ubiquitination is involved in many physiological events and promotes cancer development and progression. This review elaborates the role of ubiquitin-specific protease (USP) in programmed cell death, which is common in breast cancer cells, and lays the foundation for tumor diagnosis and targeted therapy.

Posttranslational modifications can critically affect conformational changes of amyloid-forming proteins. Ubiquitination of the microtubule-associated tau protein, an intrinsically disordered biomolecule, has been proposed to influence the formation of filamentous deposits in neurodegenerative conditions. Given the reported link between aggregation propensity and intrinsic structural preferences (e.g., transient extended structural motifs or tertiary contacts) in disordered proteins, we sought to explore the conformational landscape of ubiquitinated tau. Exploiting selective conjugation reactions, we produced single- and double-monoubiquitinated protein samples. Next, we examined the ubiquitinated species from different standpoints using NMR spectroscopy, small-angle X-ray scattering experiments, and native ion mobility-mass spectrometry (IM-MS). Moreover, we obtained atomistic representations of the conformational ensembles via scaled MD calculations, consistent with the experimental data. Modifying the repeat domain of tau with ubiquitin had a limited effect on secondary structure propensities and local mobility of distal regions. Instead, ubiquitination enhanced the compaction of the conformational ensemble, with the effect modulated by the site and the number of modifications. Native IM-MS patterns pinpointed similarities and differences between distinct tau proteoforms. It emerges that ubiquitination exerts a position-specific influence on the conformational distribution of tau molecules. This study reveals the unique conformational features of ubiquitinated forms of tau and points to their potential impact on aggregation and phase separation propensities, offering clues for a better understanding of disease-related structural alterations.

NMDA receptors for the neurotransmitter glutamate are widely distributed in the central nervous system, playing important roles in brain development, function and plasticity. Alterations in their activity are also important mediators in neuropsychiatric and neurodegenerative disorders. The different NMDA receptor subunits (GluN1, GluN2A-D and GluN3A, B) share a similar structure and membrane topology, with an intracellular C-terminus tail responsible for the interaction with proteins important for the trafficking of the receptors, and to control their surface distribution and signalling activity. The latter sequence varies among subunits but consistently contains the majority of post-translational modification sites on NMDA receptors. These modifications, including phosphorylation, ubiquitination, and palmitoylation, regulate interactions with intracellular proteins. Differences in the amino acid sequence between NMDA receptor subunits lead to a differential regulation by post-translational modifications. Since NMDA receptors are formed by oligomerization of different subunits, and each subunit is regulated in a specific manner, this creates multiple possibilities for regulation of these receptors, with impact in synaptic function and plasticity. This review addresses the diversity of mechanisms involved in the post-translational modification of NMDA receptor subunits, and their impact on the activity and distribution of the receptors, as well as their function in nerve cells.

Under stress conditions, cells reprogram their molecular machineries to mitigate damage and promote survival. Ubiquitin signaling is globally increased during oxidative stress, controlling protein fate and supporting stress defenses at several subcellular compartments. However, the rules driving subcellular ubiquitin localization to promote concerted response mechanisms remain understudied. Here, we show that K63-linked polyubiquitin chains, known to promote proteasome-independent pathways, accumulate primarily in noncytosolic compartments during oxidative stress induced by sodium arsenite in mammalian cells. Our subcellular ubiquitin proteomic analyses of noncytosolic compartments expanded 2.5-fold the pool of proteins (2,494) and provided a comprehensive number of sites (10,157) known to be ubiquitinated during arsenite stress, suggesting their involvement in a myriad of cellular pathways. Moreover, subcellular proteome analyses revealed proteins that are recruited to noncytosolic compartments under stress, including a significant enrichment of helper ubiquitin-binding adaptors of the ATPase valosin-containing protein (VCP) that processes ubiquitinated substrates for downstream signaling. We further show that VCP recruitment to noncytosolic compartments under arsenite stress occurs in a ubiquitin-dependent manner mediated by its adaptor NPLOC4. Additionally, we show that VCP and NPLOC4 activities are critical to sustain low levels of noncytosolic K63-linked ubiquitin chains, supporting a cyclical model of ubiquitin conjugation and removal that is disrupted by reactive oxygen species. This work deepens our understanding of the role of localized ubiquitin and VCP signaling in the basic mechanisms of stress response and highlights new pathways and molecular players that are essential to reshape the composition and function of the human subcellular proteome under dynamic environments.

High grade serous ovarian cancer (HGSOC) is the most lethal of all gynecologic malignancies in which the majority of patients eventually develop chemoresistant recurrent disease. Ubiquitin C-terminal hydrolase L1 (UCHL1) is a deubiquitinating enzyme canonically known for its involvement in neurodegeneration, but recently has been shown to play a key role in tumorigenesis. Furthermore, UCHL1 has garnered attention across a multitude of cancer subtypes as it has the ability to be targeted through small molecule inhibition. Therefore, the goal of this present study was to elucidate mechanistic consequences of small molecule UCHL1 inhibition in HGSOC. Comparative label-free proteomic analysis of HGSOC cell line, OVCAR8 revealed prominent changes in cell metabolism proteins upon treatment with UCHL1 small molecule inhibitor, LDN-57444. Further validation via Western blot analysis revealed that changes in cell metabolism proteins differed in matched chemosensitive versus chemoresistant HGSOC cells. Finally, cell viability analysis demonstrated that a combinatorial carboplatin and LDN-57444 blockade produced a promotion or conversely, inhibition of cell death, in chemoresistant, and chemosensitve HGSOC cells, respectively. This phenomenon was further corroborated by respective differences in activation levels of common tumor cell growth pathways STAT3, MAPK/ERK, and AKT in chemoresistant versus chemosensitive HGSOC cells. Overall, this investigation established that pharmacologic targeting of UCHL1 produces differential effects according to HGSOC chemosensitivity status.

Diabetes has a substantial impact on public health, highlighting the need for novel treatments. Ubiquitination, an intracellular protein modification process, is emerging as a promising strategy for regulating pathological mechanisms. We hypothesize that ubiquitination plays a critical role in the development and progression of diabetes and its complications, and that understanding these mechanisms can lead to new therapeutic approaches.

To uncover the research trends and advances in diabetes ubiquitination and its complications, we conducted a bibliometric analysis.

Studies on ubiquitination in diabetes mellitus and its complications were retrieved from the Web of Science Core Collection. Visual mapping analysis was conducted using the CiteSpace software.

We gathered 791 articles published over the past 23 years, focusing on ubiquitination in diabetes and its associated complications. These articles originated from 54 countries and 386 institutions, with China as the leading contributor. Shanghai Jiao Tong University has the highest number of publications in this field. The most prominent authors contributing to this research area include Wei-Hua Zhang, with Zhang Y being the most frequently cited author. Additionally, The Journal of Biological Chemistry is noted as the most cited in this field. The predominant keywords included expression, activation, oxidative stress, phosphorylation, ubiquitination, degradation, and insulin resistance.

The role of ubiquitination in diabetes and its complications, such as diabetic nephropathy and cardiomyopathy, is a key research focus. However, these areas require further investigations.

The current antiviral agents for the treatment of chronic infection with hepatitis B virus (HBV) do not completely remove covalently closed circular DNA (cccDNA) and integrated viral DNA fragments from patients. Berberine is an isoquinoline alkaloid extracted from various plants and has been reported to inhibit the replication of various types of DNA. In this study, we tested the effects of berberine and its derivatives on HBV infection. Berberine inhibited viral core promoter activity at the highest level among the compounds tested and suppressed HBV production and cccDNA synthesis in primary human hepatocytes and HBV-infected HepG2-NTCP cells at an EC50 value of 3.6 μM and a CC50 value of over 240.0 μM. Compared with other viral promoter activities, berberine treatment potently downregulated core promoter activity and reduced protein levels, but not RNA levels, of hepatic nuclear factor 4α (HNF4α), which primarily enhances enhancer II/core promoter activity. Furthermore, berberine treatment enhanced K48-linked, but not K63-linked, polyubiquitination and subsequent proteasome-dependent degradation of HNF4α. These results suggest that berberine enhances the polyubiquitination- and proteasome-dependent degradation of HNF4α and then inhibits HBV replication via the suppression of core promoter activity. The development of antiviral agents based on berberine may contribute to the amelioration of HBV-related disorders, regardless of the presence of residual cccDNA or integrated viral DNA fragments.

The oncogene xCT plays an indispensable role in tumor growth by protecting cancer cells from oxidative stress and ferroptosis. Emerging evidence indicated xCT function is tightly controlled by posttranslational modifications, especially ubiquitination. However, it still remains unclear what specific regulatory mechanism of xCT by ubiquitin ligases in human cancers. Here, we reported that NEDD4L, an E3 ubiquitin ligases, inhibited esophageal squamous cell carcinoma (ESCC) tumor growth and facilitated ferroptosis by ubiquitination of xCT. NEDD4L expression was declined in ESCC and was associated with tumor invasion, lymph node metastasis and distant metastasis. Silencing NEDD4L triggered ESCC tumor growth. Meanwhile, knock down of NEDD4L prevented the accumulation of ROS, elevated the level of GSH, reduced the content of MDA in ESCC cells, thereby inhibiting ferroptosis. Mechanistically, NEDD4L directly bound to the ∆CT domain of xCT through its WW and HECT domain. More importantly, NEDD4L promoted xCT degradation by facilitating its polyubiquitination in ESCC cells. Collectively, these findings suggest that NEDD4L is crucial in governing the stability of xCT and mediating ferroptosis in ESCC.

α-Synuclein (α-Syn) aggregation in Lewy bodies and Lewy neurites has emerged as a key pathogenetic feature in Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy. Various factors, including posttranslational modifications (PTMs), can influence the propensity of α-Syn to misfold and aggregate. PTMs are biochemical modifications of a protein that occur during or after translation and are typically mediated by enzymes. PTMs modulate several characteristics of proteins including their structure, activity, localization, and stability. α-Syn undergoes various posttranslational modifications, including phosphorylation, ubiquitination, SUMOylation, acetylation, glycation, O-GlcNAcylation, nitration, oxidation, polyamination, arginylation, and truncation. Different PTMs of a protein can physically interact with one another or work together to influence a particular physiological or pathological feature in a process known as PTMs crosstalk. The development of detection techniques for the cooccurrence of PTMs in recent years has uncovered previously unappreciated mechanisms of their crosstalk. This has led to the emergence of evidence supporting an association between α-Syn PTMs crosstalk and synucleinopathies. In this review, we provide a comprehensive evaluation of α-Syn PTMs, their impact on misfolding and pathogenicity, the pharmacological means of targeting them, and their potential as biomarkers of disease. We also highlight the importance of the crosstalk between these PTMs in α-Syn function and aggregation. Insight into these PTMS and the complexities of their crosstalk can improve our understanding of the pathogenesis of synucleinopathies and identify novel targets of therapeutic potential. SIGNIFICANCE STATEMENT: α-Synuclein is a key pathogenic protein in Parkinson's disease and other synucleinopathies, making it a leading therapeutic target for disease modification. Multiple posttranslational modifications occur at various sites in α-Synuclein and alter its biophysical and pathological properties, some interacting with one another to add to the complexity of the pathogenicity of this protein. This review details these modifications, their implications in disease, and potential therapeutic opportunities.

Cell-cell adhesion in endothelial monolayers is tightly controlled and crucial for vascular integrity. Recently, we reported on the importance of fast protein turnover for maintenance of endothelial barrier function. Specifically, continuous ubiquitination and degradation of the Rho GTPase RhoB is crucial to preserve quiescent endothelial integrity. Here, we sought to identify other barrier regulators, which are characterized by a short half-life, using a proteomics approach. Following short-term inhibition of ubiquitination with E1 ligase inhibitor MLN7243 or Cullin E3 ligase inhibitor MLN4924 in primary human endothelial cells, we identified sixty significantly differentially expressed proteins. Intriguingly, our data showed that AAMP and MTSS1 are novel negative regulators of endothelial barrier function and that their turnover is tightly controlled by ubiquitination. Mechanistically, AAMP regulates the stability and activity of RhoA and RhoB, and colocalizes with F-actin and cortactin at membrane ruffles, possibly regulating F-actin dynamics. Taken together, these findings demonstrate the critical role of protein turnover of specific proteins in the regulation of endothelial barrier function, contributing to our options to target dysregulation of vascular permeability.

The ubiquitination is crucial for controlling cellular homeostasis and protein modification, in which ubiquitin-conjugating enzyme (E2) acts as the central player in the ubiquitination system. Ubiquitin-conjugating enzymes, which have special domains that catalyse substrates, have sequence discrepancies and modulate various pathophysiological processes in different cells of multiple organisms. E2s take part in the mitosis of primordial germ cells, meiosis of spermatocytes and the formation of mature haploid spermatids to maintain normal male fertility. In this review, we summarize the various types of E2s and their functions during distinct stages of spermatogenesis.

The extensive cellular signalling events controlled by posttranslational ubiquitination are tightly regulated through the action of specialized proteases termed deubiquitinases. Among them, the OTU family of deubiquitinases can play very specialized roles in the regulation of discrete subtypes of ubiquitin signals that control specific cellular functions. To exert control over host cellular functions, some pathogenic bacteria have usurped the OTU deubiquitinase fold as a secreted virulence factor that interferes with ubiquitination inside infected cells. Herein, we provide a review of the function of bacterial OTU deubiquitinases during infection, the structural basis for their deubiquitinase activities and the bioinformatic approaches leading to their identification. Understanding bacterial OTU deubiquitinases holds the potential for discoveries not only in bacterial pathogenesis but in eukaryotic biology as well.

Under stress conditions, cells reprogram their molecular machineries to mitigate damage and promote survival. Ubiquitin signaling is globally increased during oxidative stress, controlling protein fate and supporting stress defenses at several subcellular compartments. However, the rules driving subcellular ubiquitin localization to promote these concerted response mechanisms remain understudied. Here, we show that K63-linked ubiquitin chains, known to promote proteasome-independent pathways, accumulate primarily in non-cytosolic compartments during oxidative stress induced by sodium arsenite in mammalian cells. Our subcellular ubiquitin proteomic analyses of non-cytosolic compartments expanded 10-fold the pool of proteins known to be ubiquitinated during arsenite stress (2,046) and revealed their involvement in pathways related to immune signaling and translation control. Moreover, subcellular proteome analyses revealed proteins that are recruited to non-cytosolic compartments under stress, including a significant enrichment of helper ubiquitin-binding adaptors of the ATPase VCP that processes ubiquitinated substrates for downstream signaling. We further show that VCP recruitment to non-cytosolic compartments under arsenite stress occurs in a ubiquitin-dependent manner mediated by its adaptor NPLOC4. Additionally, we show that VCP and NPLOC4 activities are critical to sustain low levels of non-cytosolic K63-linked ubiquitin chains, supporting a cyclical model of ubiquitin conjugation and removal that is disrupted by cellular exposure to reactive oxygen species. This work deepens our understanding of the role of localized ubiquitin and VCP signaling in the basic mechanisms of stress response and highlights new pathways and molecular players that are essential to reshape the composition and function of the human subcellular proteome under dynamic environments.

Proteins are the keystone for the execution of various life activities, and the maintenance of protein normalization is crucial for organisms. Ubiquitination, as a post-transcriptional modification, is widely present in organisms, and it relies on the sophisticated ubiquitin-proteasome (UPS) system that controls protein quality and modulates protein lifespan. Deubiquitinases (DUBs) counteract ubiquitination and are essential for the maintenance of homeostasis. Ubiquitin specific peptidase 3 (USP3) is a member of the DUBs that has received increasing attention in recent years. USP3 is a novel chromatin modifier that tightly regulates the DNA damage response (DDR) and maintains genome integrity. Meanwhile, USP3 acts as a key regulator of inflammatory vesicles and sustains the normal operation of the innate immune system. In addition, USP3 is aberrantly expressed in a wide range of cancers, such as gastric cancer, glioblastoma and neuroblastoma, implicating that USP3 could be an effective target for targeted therapies. In this review, we retrace all the current researches of USP3, describe the structure of USP3, elucidate its functions in DNA damage, immune and inflammatory responses and the cell cycle, and summarize the important role of USP3 in multiple cancers and diseases.

Granulation of juice sacs is a physiological disorder, which affects pomelo fruit quality. Here, the transcriptome and ubiquitinome of the granulated juice sacs were analyzed in Guanxi pomelo. We found that lignin accumulation in the granulated juice sacs was regulated at transcription and protein modification levels. In transcriptome data, we found that the genes in lignin biosynthesis pathway and antioxidant enzyme system of the granulated juice sacs were significantly upregulated. However, in ubiquitinome data, we found that ubiquitinated antioxidant enzymes increased in abundance but the enzyme activities decreased after the modification, which gave rise to reactive oxygen species (ROS) contents in granulated juice sacs. This finding suggests that ubiquitination level of the antioxidant enzymes is negatively correlated with the enzyme activities. Increased H2O2 is considered to be a signaling molecule to activate the key gene expressions in lignin biosynthesis pathway, which leads to the lignification in granulated juice sacs of pomelo. This regulatory mechanism in juice sac granulation of pomelo was further confirmed through the verification experiment using tissue culture by adding H2O2 or dimethylthiourea (DMTU). Our findings suggest that scavenging H2O2 and other ROS are important for reducing lignin accumulation, alleviating juice sac granulation and improving pomelo fruit quality.

Ubiquitination of the proteins is crucial for governing protein degradation and regulating fundamental cellular processes. Deubiquitinases (DUBs) have emerged as significant regulators of multiple pathways associated with cancer and other diseases, owing to their capacity to remove ubiquitin from target substrates and modulate signaling. Consequently, they represent potential therapeutic targets for cancer and other life-threatening conditions. USP43 belongs to the DUBs family involved in cancer development and progression. This review aims to provide a comprehensive overview of the existing scientific evidence implicating USP43 in cancer development. Additionally, it will investigate potential small-molecule inhibitors that target DUBs that may have the capability to function as anti-cancer medicines.

Niemann-Pick Type C (NPC) represents an autosomal recessive disorder with an incidence rate of 1 in 150,000 live births, classified within lysosomal storage diseases (LSDs). The abnormal accumulation of unesterified cholesterol characterizes the pathophysiology of NPC. This phenomenon is not unique to NPC, as analogous accumulations have also been observed in Alzheimer's disease, Parkinson's disease, and other neurodegenerative disorders. Interestingly, disturbances in the folding of the mutant protein NPC1 I1061T are accompanied by the aggregation of proteins such as hyperphosphorylated tau, α-synuclein, TDP-43, and β-amyloid peptide. These accumulations suggest potential disruptions in proteostasis, a regulatory process encompassing four principal mechanisms: synthesis, folding, maintenance of folding, and protein degradation. The dysregulation of these processes leads to excessive accumulation of abnormal proteins that impair cell function and trigger cytotoxicity. This comprehensive review delineates reported alterations across proteostasis mechanisms in NPC, encompassing changes in processes from synthesis to degradation. Additionally, it discusses therapeutic interventions targeting pharmacological facets of proteostasis in NPC. Noteworthy among these interventions is valproic acid, a histone deacetylase inhibitor (HDACi) that modulates acetylation during NPC1 synthesis. In addition, various therapeutic options addressing protein folding modulation, such as abiraterone acetate, DHBP, calnexin, and arimoclomol, are examined. Additionally, treatments impeding NPC1 degradation, exemplified by bortezomib and MG132, are explored as potential strategies. This review consolidates current knowledge on proteostasis dysregulation in NPC and underscores the therapeutic landscape targeting diverse facets of this intricate process.

Primary liver cancer is known for its high incidence and fatality rate. Over the years, therapeutic strategies for primary liver cancer have advanced significantly. Nonetheless, a substantial number of patients have not benefited from these methods, underscoring the pressing need for new and effective treatments for primary liver cancer. Ubiquitination is a critical post-translational modification that enables proteins to fulfill their normal biological functions and maintain their expression stability within cells. Importantly, increasing evidence suggests that the progression of liver cancer cells is often accompanied by disruptions in protein ubiquitination and deubiquitination processes. In this comprehensive review, we have compiled pertinent research about dysregulated ubiquitination in hepatocellular carcinoma (HCC) to broaden our understanding in this field. We elucidate the connections between the ubiquitination proteasome system, deubiquitination, and HCC. Furthermore, we shed light on the role of ubiquitination in cells situated within the tumor microenvironment of HCC including its involvement in mediating the activation of oncogenic pathways, reprogramming metabolic processes, and perturbing normal cellular functions. In conclusion, targeting the dysregulation of ubiquitination in HCC holds promise as a prospective and complementary therapeutic approach to existing treatments.

Linc-ROR plays an important role in gastric cancer (GC) development and progression. This study sought to determine how the aberrant expression of Linc-ROR impacts GC progression and immune evasion, and to identify new targets for GC therapy. GC cells overexpressing Linc-ROR and GSAGS cells were cocultured with NK-92 cells, respectively, and Linc-ROR expression was determined using reverse transcription polymerase chain reaction. Linc-ROR overexpression experiments were used to measure the expression of MICB, a tumor protein that is recognized by natural killer (NK) cells. Bioinformatics analysis identified retinoid X receptor α (RXRA) and YY1 as MICB-specific transcription factors. Cotransfection and ubiquitinated drug experiments found that Linc-ROR promoted the ubiquitination and degradation of RXRA. Linc-ROR was upregulated in GC tissue and high expression was associated with tumor escape from NK-92 cell-mediated immunity. Linc-ROR overexpression inhibited the expression of MICB on the cell surface by degrading RXRA. These findings indicate that Linc-ROR promotes the binding of RXRA and E3 ligase UBE4B, reducing RXRA and MICB expression, and limiting NK cell-killing activity. Linc-ROR is a critical long noncoding RNA with a tumor-promoting function in GC and thus may serve as a potential therapeutic target.

Smurf1 and Smurf2 are two closely related member of the HECT (homologous to E6AP carboxy terminus) E3 ubiquitin ligase family and play important roles in the regulation of various cellular processes. Both were initially identified to regulate transforming growth factor-β and bone morphogenetic protein signaling pathways through regulating Smad protein stability and are now implicated in various pathological processes. Generally, E3 ligases, of which over 800 exist in humans, are ideal targets for inhibition as they determine substrate specificity; however, there are few inhibitors with the ability to precisely target a particular E3 ligase of interest. In this work, we explored a panel of ubiquitin variants (UbVs) that were previously identified to bind Smurf1 or Smurf2. In vitro binding and ubiquitination assays identified a highly specific Smurf2 inhibitor, UbV S2.4, which was able to inhibit ligase activity with high potency in the low nanomolar range. Orthologous cellular assays further demonstrated high specificity of UbV S2.4 toward Smurf2 and no cross-reactivity toward Smurf1. Structural analysis of UbV S2.4 in complex with Smurf2 revealed its mechanism of inhibition was through targeting the E2 binding site. In summary, we investigated several protein-based inhibitors of Smurf1 and Smurf2 and identified a highly specific Smurf2 inhibitor that disrupts the E2-E3 protein interaction interface.

Zika virus (ZIKV) is a re-emerging RNA virus and causes major public health events due to its link to severe neurological complications in foetuses and neonates. The cGAS-STING signalling pathway regulates innate immunity and plays an important role in the invasion of DNA and RNA viruses. This study reveals a distinct mechanism by which ZIKV restricts the cGAS-STING signalling to repress IFN-β expression. ZIKV attenuates IFN-β expression induced by DNA viruses (herpes simplex virus type 1, HSV-1) or two double-stranded DNAs (dsDNA90 and HSV120) in mouse embryonic fibroblasts (MEFs). Notably, ZIKV NS5, the viral RNA-dependent RNA polymerase, was responsible for the repression of IFN-β. NS5 interacts with STING in the cytoplasm, suppresses IRF3 phosphorylation and nucleus localization and promotes the cleavage of STING K48-linked polyubiquitination. Furthermore, the NS5 methyltransferase (MTase) domain interacts with STING to restrict STING-induced IFN-β expression. Interestingly, point mutation analyses of conserved methyltransferase active site residue D146 indicate that it is critical for repressing IFN-β expression induced by STING stimulation in cGAS-STING signalling.

Ubiquitylation refers to the attachment of mono- or poly-ubiquitin molecules to a substrate protein. To shield ubiquitin chains against potential hydrolysis, a facile, click-chemistry based approach was recently established for the generation of site-specifically conjugated ubiquitin dimers relying on triazole-linkage. Here, the preparation of such ubiquitin chains was advanced by the generation of homotypic Lys11-linked ubiquitin trimers considering an isotopic labeling scheme in a moiety-wise manner. The structural and dynamical impact on the ubiquitin unit at proximal, central, or distal position that is potentially invoked by the respective other two moieties was systematically probed by heteronuclear high-resolution NMR spectroscopic approaches. As a result, conjugating a third ubiquitin moiety to the proximal or distal site of a ubiquitin dimer does not alter structural and dynamical characteristics as it has been seen for ubiquitin dimers. This observation suggests that recognition of a homotypically assembled ubiquitin chain by a potential substrate is primarily done by screening the length of a ubiquitin chain rather than relying on subtle changes in structure or dynamic properties of single ubiquitin moieties composing the chain.

Ubiquitination, a crucial post-translational modification, plays a role in nearly all physiological processes. Its functional execution depends on a series of catalytic reactions involving numerous proteases. TRIM26, a protein belonging to the TRIM family, exhibits E3 ubiquitin ligase activity because of its RING structural domain, and is present in diverse cell lineages. Over the last few decades, TRIM26 has been documented to engage in numerous physiological and pathological processes as a controller, demonstrating a diverse array of biological roles. Despite the growing research interest in TRIM26, there has been limited attention given to examining the protein's structure and function in existing reviews. This review begins with a concise overview of the composition and positioning of TRIM26 and then proceeds to examine its roles in immune response, viral invasion, and inflammatory processes. Simultaneously, we demonstrate the contribution of TRIM26 to the progression of various diseases, encompassing numerous malignancies and neurologic conditions. Finally, we have investigated the potential areas for future research on TRIM26.

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.

Pellino1 (Peli1) is a highly conserved E3 Ub ligase that exerts its biological functions by mediating target protein ubiquitination. Extensive evidence has demonstrated the crucial role of Peli1 in regulating inflammation by modulating various receptor signaling pathways, including interleukin-1 receptors, Toll-like receptors, nuclear factor-κB, mitogen-activated protein kinase, and phosphoinositide 3-kinase/AKT pathways. Peli1 has been implicated in the development of several diseases by influencing inflammation, apoptosis, necrosis, pyroptosis, autophagy, DNA damage repair, and glycolysis. Peli1 is a risk factor for most cancers, including breast cancer, lung cancer, and lymphoma. Conversely, Peli1 protects against herpes simplex virus infection, systemic lupus erythematosus, esophageal cancer, and toxic epidermolysis bullosa. Therefore, Peli1 is a potential therapeutic target that warrants further investigation. This comprehensive review summarizes the target proteins of Peli1, delineates their involvement in major signaling pathways and biological processes, explores their role in diseases, and discusses the potential clinical applications of Peli1-targeted therapy, highlighting the therapeutic prospects of Peli1 in various diseases.

Radiotherapy, a treatment method employing radiation to eradicate tumor cells and subsequently reduce or eliminate tumor masses, is widely applied in the management of numerous patients with tumors. However, its therapeutic effectiveness is somewhat constrained by various drug-resistant factors. Recent studies have highlighted the ubiquitination/deubiquitination system, a reversible molecular modification pathway, for its dual role in influencing tumor behaviors. It can either promote or inhibit tumor progression, impacting tumor proliferation, migration, invasion, and associated therapeutic resistance. Consequently, delving into the potential mechanisms through which ubiquitination and deubiquitination systems modulate the response to radiotherapy in malignant tumors holds paramount significance in augmenting its efficacy. In this paper, we comprehensively examine the strides made in research and the pertinent mechanisms of ubiquitination and deubiquitination systems in governing radiotherapy resistance in tumors. This underscores the potential for developing diverse radiosensitizers targeting distinct mechanisms, with the aim of enhancing the effectiveness of radiotherapy.

Ubiquitination (or ubiquitylation) and de-ubiquitination, which are both post-translational modifications (PTMs) of proteins, have become a research hotspot in recent years. Some ubiquitinated or de-ubiquitinated signaling proteins have been found to promote or suppress innate immunity through Toll-like receptor (TLR), RIG-like receptor (RIG-I-like receptor, RLR), NOD-like receptor (NLR), and the cyclic guanosine monophosphate (GMP)-adenosine monophosphate (AMP) synthase (cGAS)-STING pathway. This article aimed to provide a review on the role of ubiquitination and de-ubiquitination, especially ubiquitin ligase enzymes and de-ubiquitinating enzymes, in the above four pathways. We hope that our work can contribute to the research and development of treatment strategies for innate immunity-related diseases such as inflammatory bowel disease.

Protein degradation is a general process to maintain cell homeostasis. The intracellular protein quality control system mainly includes the ubiquitin-proteasome system and the lysosome pathway. Inspired by the physiological process, strategies to degrade specific proteins have developed, which emerge as potent and effective tools in biological research and drug discovery. This review focuses on recent advances in targeted protein degradation techniques, summarizing the principles, advantages, and challenges. Moreover, the potential applications and future direction in biological science and clinics are also discussed.

Ubiquitin-specific peptidase 9X (USP9X) is involved in certain human diseases, including malignancies, atherosclerosis and certain diseases of the nervous system. USP9X promotes the deubiquitination and stabilization of diverse substrates, thereby exerting a versatile range of effects on pathological and physiological processes. USP9X serves vital roles in the processes of cell survival, invasion and migration in various types of cancer. The present review aims to highlight the current knowledge of USP9X in terms of its structure and the possible mediatory mechanisms involved in certain types of cancer, providing a thorough introduction to its biological functions in carcinogenesis and further outlining its oncogenic or suppressive properties in a diverse range of cancer types. Finally, several perspectives regarding USP9X-targeted pharmacological therapeutics in cancer development are discussed.

Beclin-1 is the firstly-identified mammalian protein of the autophagy machinery, which functions as a molecular scaffold for the assembly of PI3KC3 (class III phosphatidylinositol 3 kinase) complex, thus controlling autophagy induction and other cellular trafficking events. Notably, there is mounting evidence establishing the implications of Beclin-1 in diverse tumorigenesis processes, including tumor suppression and progression as well as resistance to cancer therapeutics and CSC (cancer stem-like cell) maintenance. More importantly, Beclin-1 has been confirmed as a potential target for the treatment of multiple cancers. In this review, we provide a comprehensive survey of the structure, functions, and regulations of Beclin-1, and we discuss recent advances in understanding the controversial roles of Beclin-1 in oncology. Moreover, we focus on summarizing the targeted Beclin-1-regulating strategies in cancer therapy, providing novel insights into a promising strategy for regulating Beclin-1 to improve cancer therapeutics in the future.

Finding effective treatments for cancer requires a thorough understanding of how it develops and progresses. Recent research has revealed the crucial role that Zinc and ring finger 2 (ZNRF2) play in the progression of non-small cell lung cancer (NSCLC) by controlling cell growth and death. However, a comprehensive analysis of ZNRF2's role in cancer as a whole has yet to be conducted. Our study sought to investigate the impact of ZNRF2 on diverse human tumours, as well as the molecular pathways involved, using databases such as TCGA (The Cancer Genome Atlas), GEO (Gene Expression Omnibus) and the Human Protein Atlas (HPA), as well as several bioinformatic tools. Our findings indicate that ZNRF2 is generally expressed at higher levels in tumours than in normal tissues, and in some cancers, its levels correlate positively with disease stage, potentially predicting a poor prognosis for patients. We also discovered genetic changes in ZNRF2 among cancer patients, as well as its relationship with cancer-related fibroblasts, endothelial cells and immune cell infiltration. Additionally, we explored potential molecular mechanisms of ZNRF2 in tumours, finding that it increases in hepatocellular carcinoma (HCC) tissues and that inhibiting its expression through ZNRF2 siRNA can limit HepG2 cell proliferation. Overall, our study provides a comprehensive overview of ZNRF2's oncogenic roles across various cancers.

Dynamic monitoring of intracellular ubiquitin (Ub) conjugates is instrumental to understanding the Ub regulatory machinery. Although many biochemical approaches have been developed to characterize protein ubiquitination, chemical tools capable of temporal resolution probing of ubiquitination events remain to be developed. Here, we report the development of the first cell-permeable and stimuli-responsive Ub probe and its application for the temporal resolution profiling of ubiquitinated substrates in live cells. The probe carrying the photolabile group N-(2-nitrobenzyl)-Gly (Nbg) on the amide bond between Ub Gly75 and Gly76 is readily prepared through chemical synthesis and can be delivered to live cells by conjugation via a disulfide bond with the cyclic cell-penetrating peptide cR10D (i.e., 4-((4-(dimethylamino)phenyl)-azo)-benzoic acid-modified cyclic deca-arginine). Both in vitro and in vivo experiments showed that Ub-modifying enzymes (E1, E2s, and E3s) could not install the Ub probe onto substrate proteins prior to removal of the nitrobenzyl group, which was easily accomplished via photoirradiation. The utility and practicality of this probe were exemplified by the time-resolved biochemical and proteomic investigation of ubiquitination events in live cells during a H2O2-mediated oxidative stress response. This work shows a conceptually new family of chemical Ub tools for the time-resolved studies on dynamic protein ubiquitination in different biological processes and highlights the utility of modern chemical protein synthesis in obtaining custom-designed tools for biological studies.

Tissue-infiltrating neutrophils (TINs) secrete various signaling molecules to establish paracrine communication within the inflammatory milieu. It is imperative to identify molecular mediators that control this secretory phenotype of TINs. The present study uncovers a secretory neutrophil subset that exhibits increased pro-inflammatory cytokine production and enhanced migratory capacity which is highly related with periodontal pathogenesis. Further analysis identifies the OTU domain-containing protein 1 (OTUD1) plays a regulatory role in this secretory neutrophil polarization. In human and mouse periodontitis, the waning of inflammation is correlated with OTUD1 upregulation, whereas severe periodontitis is induced when neutrophil-intrinsic OTUD1 is depleted. Mechanistically, OTUD1 interacts with SEC23B, a component of the coat protein II complex (COPII). By removing the K63-linked polyubiquitin chains on SEC23B Lysine 81, the deubiquitinase OTUD1 negatively regulates the COPII secretory machinery and limits protein ER-to-Golgi trafficking, thus restricting the surface expression of integrin-regulated proteins, CD9 and CD47. Accordingly, blockade of protein transport by Brefeldin A (BFA) curbs recruitment of Otud1-deficient TINs and attenuates inflammation-induced alveolar bone destruction. The results thus identify OTUD1 signaling as a negative feedback loop that limits the polarization of neutrophils with secretory phenotype and highlight the potential application of BFA in the treatment of periodontal inflammation.

Ubiquitination modulates protein turnover or activity depending on the number and location of attached ubiquitin (Ub) moieties. Proteins marked by a lysine 48 (K48)-linked polyubiquitin chain are usually targeted to the 26S proteasome for degradation; however, other polyubiquitin chains, such as those attached to K63, usually regulate other protein properties. Here, we show that 2 PLANT U-BOX E3 ligases, PUB25 and PUB26, facilitate both K48- and K63-linked ubiquitination of the transcriptional regulator INDUCER OF C-REPEAT BINDING FACTOR (CBF) EXPRESSION1 (ICE1) during different periods of cold stress in Arabidopsis (Arabidopsis thaliana), thus dynamically modulating ICE1 stability. Moreover, PUB25 and PUB26 attach both K48- and K63-linked Ub chains to MYB15 in response to cold stress. However, the ubiquitination patterns of ICE1 and MYB15 mediated by PUB25 and PUB26 differ, thus modulating their protein stability and abundance during different stages of cold stress. Furthermore, ICE1 interacts with and inhibits the DNA-binding activity of MYB15, resulting in an upregulation of CBF expression. This study unravels a mechanism by which PUB25 and PUB26 add different polyubiquitin chains to ICE1 and MYB15 to modulate their stability, thereby regulating the timing and degree of cold stress responses in plants.

Cell death is critical for the development and homeostasis of almost all multicellular organisms. Moreover, its dysregulation leads to diverse disease states. Historically, apoptosis was thought to be the major regulated cell death pathway, whereas necrosis was considered to be an unregulated form of cell death. However, research in recent decades has uncovered several forms of regulated necrosis that are implicated in degenerative diseases, inflammatory conditions and cancer. The growing insight into these regulated, non-apoptotic cell death pathways has opened new avenues for therapeutic targeting. Here, we describe the regulatory pathways of necroptosis, pyroptosis, parthanatos, ferroptosis, cuproptosis, lysozincrosis and disulfidptosis. We discuss small-molecule inhibitors of the pathways and prospects for future drug discovery. Together, the complex mechanisms governing these pathways offer strategies to develop therapeutics that control non-apoptotic cell death.

The pyruvate kinase enzyme PKM2 catalyzes the final step in glycolysis and converts phosphoenolpyruvate (PEP) to pyruvate. PKM2 is often overexpressed in cancer and plays a role in the Warburg effect. The expression of PKM2 can be regulated at different levels. While it has been proven that PKM2 can be regulated by ubiquitination, little is known about its de-ubiquitination regulation.

Immunoprecipitation was applied to identify the PKM2 interaction protein and to determine the interaction region between PKM2 and USP4. Immunofluorescence was performed to determine the cellular localization of USP4 and PKM2. The regulation of PKM2 by USP4 was examined by western blot and ubiquitination assay. MTT assays, glucose uptake, and lactate production were performed to analyze the biological effects of USP4 in gastric cancer cells.

USP4 interacts with PKM2 and catalyzes the de-ubiquitination of PKM2. Overexpression of USP4 promotes cell proliferation, glucose uptake, and lactate production in gastric cancer cells. Knockdown of USP4 reduces PKM2 levels and results in a reduction in cell proliferation and the glycolysis rate.

USP4 plays a tumor-promoting role in gastric cancer cells by regulating PKM2.

With the rapidly increasing incidence of bladder cancer in China and worldwide, great efforts have been made to understand the detailed mechanism of bladder cancer tumorigenesis. Recently, the introduction of immune checkpoint inhibitor-based immunotherapy has changed the treatment strategy for bladder cancer, especially for advanced bladder cancer, and has improved the survival of patients. The ubiquitin-proteasome system, which affects many biological processes, plays an important role in bladder cancer. Several E3 ubiquitin ligases and deubiquitinases target immune checkpoints, either directly or indirectly. In this review, we summarize the recent progress in E3 ubiquitin ligases and deubiquitinases in bladder cancer tumorigenesis and further highlight the implications for bladder cancer immunotherapies.

In Alzheimer's disease and related disorders called tauopathies, the microtubule-associated protein tau accumulates in the brain in the form of amyloid-like supramolecular filaments. As an intrinsically disordered protein, tau undergoes many post-translational modifications, including ubiquitination. Alterations to the levels of ubiquitination of tau have been observed at various stages of neurodegenerative conditions. We focus on proteoform-specific interrogations to obtain mechanistic insight into the effects of ubiquitination on disease-related conformational transitions of tau. Single and double ubiquitination of tau at residues Lys311 and Lys317 is strongly associated with pathological conditions. In this study, we leveraged disulfide-directed chemistry to install ubiquitin at one or both of those positions in the isolated microtubule-binding repeat domain of tau. We obtained homogeneously modified tau proteins and observed that they retained disordered character in solution. We found that ubiquitination in position 317 (with or without ubiquitination in position 311) impaired the formation of ordered fibrillar structures via oligomeric intermediates. Since the transition to fibrillar species may proceed via an alternative condensation pathway involving liquid droplet intermediates, we further tested the ability of the ubiquitinated proteoforms to phase separate. Single monoubiquitinated tau species were able to coacervate, however no liquid droplets were observed for the double ubiquitinated form. Taken together, the data indicate that double ubiquitination in the third repeat of tau disfavors the formation of amyloid aggregates by distinct mechanisms, suggesting that the presence of ubiquitinated residues 311 and 317 in insoluble tau may result from modifications in advanced stages of aggregation. These findings contribute to our understanding of the influence of site-specific ubiquitination on the pathological conformational transitions of a prototypical intrinsically disordered protein.

Ubiquitin C-terminal hydrolase-L1 (UCH-L1), a member of the lesser-known deubiquitinating enzyme family, has deubiquitinase and ubiquitin (Ub) ligase activity and the role of stabilizing Ub. UCH-L1 was first discovered in the brain and is associated with regulating cell differentiation, proliferation, transcriptional regulation and numerous other biological processes. UCH-L1 is predominantly expressed in the brain and serves a role in tumor promotion or inhibition. There is still controversy about the effect of UCH-L1 dysregulation in cancer and its mechanisms are unknown. Extensive research to investigate the mechanism of UCH-L1 in different types of cancer is key for the future treatment of UCH-L1-associated cancer. The present review details the molecular structure and function of UCH-L1. The role of UCH-L1 in different types of cancer is also summarized and how novel treatment targets provide a theoretical foundation in cancer research is discussed.