Search Results (56)

Background: Vitiligo, a chronic depigmentation disorder driven by oxidative stress and immune dysregulation, remains poorly understood mechanistically. The Keap1/NRF2/ARE pathway is critical for melanocyte protection against oxidative damage; however, the role of Cullin-3 (CUL3), a scaffold for E3 ubiquitin ligases that regulate NRF2 degradation, and its interplay with inflammatory mediators in vitiligo pathogenesis are underexplored. This study investigates CUL3, NRF2, and the associated regulatory networks in vitiligo, integrating clinical profiling and computational docking to identify therapeutic targets. Methods: A case-control study compared non-segmental vitiligo patients with age-/sex-matched controls. Lesional skin biopsies were analyzed by qRT-PCR for the expression of CUL3, NRF2, miRNA-146a, FOXP3, NF-κB, IL-6, TNF-α, and P53. Molecular docking was used to evaluate vitexin’s binding affinity to Keap1, validated by root mean square deviation (RMSD) calculations. Results: Patients with vitiligo exhibited significant downregulation of CUL3 (0.27 ± 0.03 vs. 1 ± 0.58; p = 0.013), NRF2 (0.37 ± 0.26 vs. 1 ± 0.8; p = 0.001), and FOXP3 (0.09 ± 0.2 vs. 1 ± 0.3; p = 0.001), alongside the upregulation of miRNA-146a (4.7 ± 1.9 vs. 1 ± 0.8; p = 0.001), NF-κB (4.7 ± 1.9 vs. 1 ± 0.5; p = 0.001), IL-6 (2.8 ± 1.5 vs. 1 ± 0.4; p = 0.001), and TNF-α (2.2 ± 1.1 vs. 1 ± 0.3; p = 0.001). P53 showed no differential expression (p > 0.05). Docking revealed a strong binding of vitexin to Keap1 (RMSD: 0.23 Å), mirroring the binding of the control ligand CDDO-Im. Conclusions: Dysregulation of the CUL3/Keap1/NRF2 axis and elevated miRNA-146a levels correlate with vitiligo progression, suggesting a role for oxidative stress and immune imbalance. Vitexin’s high-affinity docking to Keap1 positions it as a potential modulator of the NRF2 pathway, offering novel therapeutic avenues. This study highlights the translational potential of targeting the ubiquitin–proteasome and antioxidant pathways in the management of vitiligo. Full article

Chagas disease, caused by the protozoan parasite Trypanosoma cruzi (T. cruzi), continues to pose significant public health challenges due to the toxicity, poor tolerability, and limited efficacy of current treatments. Targeted protein degradation (TPD) using proteolysis-targeting chimeras (PROTACs) represents a novel therapeutic avenue by leveraging the ubiquitin–proteasome system to selectively degrade essential parasite proteins. This review introduces the conceptual framework of “TrypPROTACs” as a prospective strategy for T. cruzi, integrating a comprehensive analysis of druggable targets across critical biological pathways, including ergosterol biosynthesis, redox metabolism, glycolysis, nucleotide synthesis, protein kinases, molecular chaperones such as heat shock protein 90 (Hsp90), and epigenetic regulators such as T. cruzi bromodomain factor 3 (TcBDF3). It is important to note that no TrypPROTAC compound has yet been synthesized or experimentally validated in T. cruzi; the approach discussed herein remains theoretical and forward-looking. Representative inhibitors for each target class are compiled, highlighting potency, selectivity, and structural features relevant to ligand design. We also examine the parasite’s ubiquitination machinery and compare it to the human system to identify putative E3 ubiquitin ligases. Key aspects of linker engineering and ternary complex stabilization are discussed, alongside potential validation techniques such as the cellular thermal shift assay (CETSA) and bioluminescence resonance energy transfer (NanoBRET). Collectively, these insights outline a roadmap for the rational design of TrypPROTACs and support the feasibility of expanding targeted protein degradation strategies to neglected tropical diseases. Full article

Transcription factors (TFs) play central roles in gene regulation and disease progression but have long been considered undruggable due to the absence of well-defined binding pockets and their reliance on protein–protein or protein–DNA interactions. Proteolysis-targeting chimeras (PROTACs) offer a novel strategy to overcome these limitations by inducing selective degradation of TFs via the ubiquitin–proteasome system. This review highlights recent advances in TF-targeting PROTACs, focusing on key oncogenic TFs such as androgen receptor (AR), estrogen receptor alpha (ERα), BRD4, c-Myc, and STAT family members. Strategies for ligand design—including small molecules, peptides, and nucleic acid-based elements—are discussed alongside the use of various E3 ligases such as VHL, CRBN, and IAP. Several clinically advanced PROTACs, including ARV-110 and ARV-471, demonstrate the therapeutic potential of this technology. Despite challenges in pharmacokinetics and E3 ligase selection, emerging data suggest that PROTACs can successfully target TFs, paving the way for new treatment strategies across oncology and other disease areas. Full article

B30.2 domains, sometimes referred to as PRY/SPRY domains, were originally identified by sequence profiling methods at the gene level. The B30.2 domain comprises a concanavalin A-like fold consisting of two twisted seven-stranded anti-parallel β-sheets. B30.2 domains are present in about 150 human and 700 eukaryotic proteins, usually fused to other domains. The B30.2 domain represents a scaffold, which, through six variable loops, binds different unrelated peptides or endogenous low-molecular-weight compounds. At the cellular level, B30.2 proteins engage in supramolecular assemblies with important signaling functions. In humans, B30.2 domains are often found in E3-ligases, such as tripartite motif (Trim) proteins, SPRY domain-containing SOCS box proteins, Ran binding protein 9 and −10, Ret-finger protein-like, and Ring-finger proteins. The B30.2 protein recognizes the target and recruits the E2-conjugase by means of the fused domains, often involving specific adaptor proteins. Further well-studied B30.2 proteins are the methyltransferase adaptor protein Ash2L, some butyrophilins, and Ryanodine Receptors. Although the affinity of an isolated B30.2 domain to its ligand might be weak, it can increase strongly due to avidity effects upon recognition of oligomeric targets or in the context of macromolecular machines. Full article

This study focuses on the synthesis of Biguanide-PROTACs, formed by conjugating the biguanide motif with a spacer and a ligand for recognition subunits of two E3 ubiquitin ligases. Evaluation of their activity on pancreatic cancer cell (KP4) proliferation established a correlation between membrane permeability and median effective concentration. Mechanistic insights revealed that only two compounds exhibited biguanide-like AMPK activation, while only one hydrophobic compound uniquely altered mitochondrial protein levels. The prospect of developing and expanding the Biguanide-PROTAC library holds several promises, offering potential insights into biguanide mechanisms and the creation of more potent anticancer agents. This study contributes to understanding the intricate interplay between compound structure, permeability, and anticancer activity, paving the way for targeted drug development in pancreatic cancer treatment. Full article

Bivalve mollusks, comprising animals enclosed in two shell valves, are well-adapted to benthic life in many intertidal zones. Clams have evolved the buried lifestyle, which depends on their unique soft tissue structure and their wedge-shaped muscular foot and long extendible siphons. However, molecular mechanisms of adaptative phenotype evolution remain largely unknown. In the present study, we obtain the high-quality chromosome-level genome of Manila clam R. philippinarum, an economically important marine bivalve in many coastal areas. The genome is constructed by the Hi-C assisted assembly, which yields 19 chromosomes with a total of 1.17 Gb and BUSCO integrity of 92.23%. The de novo assembled genome has a contig N50 length of 307.7 kb and scaffold N50 of 59.5 Mb. Gene family expansion analysis reveals that a total of 24 single-copy gene families have undergone the significant expansion or contraction, including E3 ubiquitin ligase and dynein heavy chain. The significant expansion of transposable elements has been also identified, including long terminal repeats (LTR) and non-LTR retrotransposons. The comparative transcriptomics among different clam tissues reveals that extracellular matrix (ECM) receptors and neuroactive ligand receptors may play the important roles in tissue structural support and neurotransmission during their infaunal life. These findings of gene family expansion and tissue-specific expression may reflect the unique soft tissue structure of clams, suggesting the evolution of lineage-specific morphological novelties. The high-quality genome and transcriptome data of R. philippinarum will not only facilitate the genetic studies on clams but will also provide valuable information on morphological novelties in mollusks. Full article

Notch signaling is a conserved pathway crucial for nervous system development. Disruptions in this pathway are linked to neurodevelopmental disorders, neurodegenerative diseases, and brain tumors. Hairy/E(spl) (HES) genes, major downstream targets of Notch, are commonly used as markers for Notch activation. However, these genes can be activated, inhibited, or function independently of Notch signaling, and their response to Notch disruption varies across tissues and developmental stages. MIB1/Mib1 is an E3 ubiquitin ligase that enables Notch receptor activation by processing ligands like Delta and Serrate. We investigated Notch signaling disruption using the zebrafish Mib1 mutant line, mib1^ta52b, focusing on changes in the expression of Hairy/E(spl) (her) genes. Our findings reveal significant variability in her gene expression across different neural cell types, regions, and developmental stages following Notch disruption. This variability questions the reliability of Hairy/E(spl) genes as universal markers for Notch activation, as their response is highly context-dependent. This study highlights the complex and context-specific nature of Notch signaling regulation. It underscores the need for a nuanced approach when using Hairy/E(spl) genes as markers for Notch activity. Additionally, it provides new insights into Mib1’s role in Notch signaling, contributing to a better understanding of its involvement in Notch signaling-related disorders. Full article

Cyclophilin A (CypA), the cellular receptor of the immunosuppressant cyclosporin A (CsA), is an abundant cytosolic protein and is involved in a variety of diseases. For example, CypA supports cancer proliferation and mediates viral infections, such as the human immunodeficiency virus 1 (HIV-1). Here, we present the design of PROTAC (proteolysis targeting chimera) compounds against CypA to induce its intracellular proteolysis and to investigate their effect on immune cells. Interestingly, upon connecting to E3 ligase ligands, both peptide-based low-affinity binders and CsA-based high-affinity binders can degrade CypA at nM concentration in HeLa cells and fibroblast cells. As the immunosuppressive effect of CsA is not directly associated with the binding of CsA to CypA but the inhibition of phosphatase calcineurin by the CypA:CsA complex, we investigated whether a CsA-based PROTAC compound could induce CypA degradation without affecting the activation of immune cells. P3, the most efficient PROTAC compound discovered from this study, could deplete CypA in lymphocytes without affecting cell proliferation and cytokine production. This work demonstrates the feasibility of the PROTAC approach in depleting the abundant cellular protein CypA at low drug dosage without affecting immune cells, allowing us to investigate the potential therapeutic effects associated with the endogenous protein in the future. Full article

TRAF6 is an E3 ubiquitin ligase that plays a crucial role in cell signaling. It is known that MMP is involved in tumor metastasis, and TRAF6 induces MMP-9 expression by binding to BSG. However, inhibiting TRAF6’s ubiquitinase activity without disrupting the RING domain is a challenge that requires further research. To address this, we conducted computer-based drug screening to identify potential TRAF6 inhibitors. Using a ligand–receptor complex pharmacophore based on the inhibitor EGCG, known for its anti-tumor properties, we screened 52,765 marine compounds. After the molecular docking of 405 molecules with TRAF6, six compounds were selected for further analysis. By replacing fragments of non-binding compounds and conducting second docking, we identified two promising molecules, CMNPD9212-16 and CMNPD12791-8, with strong binding activity and favorable pharmacological properties. ADME and toxicity predictions confirmed their potential as TRAF6 inhibitors. Molecular dynamics simulations showed that CMNPD12791-8 maintained a stable structure with the target protein, comparable to EGCG. Therefore, CMNPD12791-8 holds promise as a potential inhibitor of TRAF6 for inhibiting tumor growth and metastasis. Full article

Suppressor of deltex (Su(dx)) is a Drosophila melanogaster member of the NEDD4 family of the HECT domain E3 ubiquitin ligases. Su(dx) acts as a regulator of Notch endocytic trafficking, promoting Notch lysosomal degradation and the down-regulation of both ligand-dependent and ligand-independent signalling, the latter involving trafficking through the endocytic pathway and activation of the endo/lysosomal membrane. Mutations of Su(dx) result in developmental phenotypes in the Drosophila wing that reflect increased Notch signalling, leading to gaps in the specification of the wing veins, and Su(dx) functions to provide the developmental robustness of Notch activity to environmental temperature shifts. The full developmental functions of Su(dx) are unclear; however, this is due to a lack of a clearly defined null allele. Here we report the first defined null mutation of Su(dx), generated by P-element excision, which removes the complete open reading frame. We show that the mutation is recessive-viable, with the Notch gain of function phenotypes affecting wing vein and leg development. We further uncover new roles for Su(dx) in Drosophila oogenesis, where it regulates interfollicular stalk formation, egg chamber separation and germline cyst enwrapment by the follicle stem cells. Interestingly, while the null allele exhibited a gain in Notch activity during oogenesis, the previously described Su(dx)^SP allele, which carries a seven amino acid in-frame deletion, displayed a Notch loss of function phenotypes and an increase in follicle stem cell turnover. This is despite both alleles displaying similar Notch gain of function in wing development. We attribute this unexpected context-dependent outcome of Su(dx)^sp being due to the partial retention of function by the intact C2 and WW domain regions of the protein. Our results extend our understanding of the developmental role of Su(dx) in the tissue renewal and homeostasis of the Drosophila ovary and illustrate the importance of examining an allelic series of mutations to fully understand developmental functions. Full article

Programmed death ligand 1 (PD-L1) plays a pivotal role in cancer immune evasion and is a critical target for cancer immunotherapy. This review focuses on the regulation of PD-L1 through the dynamic processes of ubiquitination and deubiquitination, which are crucial for its stability and function. Here, we explored the intricate mechanisms involving various E3 ubiquitin ligases and deubiquitinating enzymes (DUBs) that modulate PD-L1 expression in cancer cells. Specific ligases are discussed in detail, highlighting their roles in tagging PD-L1 for degradation. Furthermore, we discuss the actions of DUBs that stabilize PD-L1 by removing ubiquitin chains. The interplay of these enzymes not only dictates PD-L1 levels but also influences cancer progression and patient response to immunotherapies. Furthermore, we discuss the therapeutic implications of targeting these regulatory pathways and propose novel strategies to enhance the efficacy of PD-L1/PD-1-based therapies. Our review underscores the complexity of PD-L1 regulation and its significant impact on the tumor microenvironment and immunotherapy outcomes. Full article

SWI/SNF enzymes are heterogeneous multi-subunit complexes that utilize the energy from ATP hydrolysis to remodel chromatin structure, facilitating transcription, DNA replication, and repair. In mammalian cells, distinct sub-complexes, including cBAF, ncBAF, and PBAF exhibit varying subunit compositions and have different genomic functions. Alterations in the SWI/SNF complex and sub-complex functions are a prominent feature in cancer, making them attractive targets for therapeutic intervention. Current strategies in cancer therapeutics involve the use of pharmacological agents designed to bind and disrupt the activity of SWI/SNF complexes or specific sub-complexes. Inhibitors targeting the catalytic subunits, SMARCA4/2, and small molecules binding SWI/SNF bromodomains are the primary approaches for suppressing SWI/SNF function. Proteolysis-targeting chimeras (PROTACs) were generated by the covalent linkage of the bromodomain or ATPase-binding ligand to an E3 ligase-binding moiety. This engineered connection promotes the degradation of specific SWI/SNF subunits, enhancing and extending the impact of this pharmacological intervention in some cases. Extensive preclinical studies have underscored the therapeutic potential of these drugs across diverse cancer types. Encouragingly, some of these agents have progressed from preclinical research to clinical trials, indicating a promising stride toward the development of effective cancer therapeutics targeting SWI/SNF complex and sub-complex functions. Full article

Protein–protein interactions play crucial roles in a wide range of biological processes, including metabolic pathways, cell cycle progression, signal transduction, and the proteasomal system. For PPIs to fulfill their biological functions, they require the specific recognition of a multitude of interacting partners. In many cases, however, protein–protein interaction domains are capable of binding different partners in the intracellular environment, but they require precise regulation of the binding events in order to exert their function properly and avoid misregulation of important molecular pathways. In this work, we focused on the MATH domain of the E3 Ligase adaptor protein SPOP in order to decipher the molecular features underlying its interaction with two different peptides that mimic its physiological partners: Puc and MacroH2A. By employing stopped-flow kinetic binding experiments, together with extensive site-directed mutagenesis, we addressed the roles of specific residues, some of which, although far from the binding site, govern these transient interactions. Our findings are compatible with a scenario in which the binding of the MATH domain with its substrate is characterized by a fine energetic network that regulates its interactions with different ligands. Results are briefly discussed in the context of previously existing work regarding the MATH domain. Full article

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor and a substrate protein of a Cullin 4B E3 ligase complex responsible for diverse cellular processes. In the lung, this receptor is responsible for the bioactivation of benzo[a]pyrene during tumorigenesis. Realizing that the AHR function is affected by its expression level, we are interested in the degradation mechanism of AHR in the lung. Here, we have investigated the mechanism responsible for AHR degradation using human lung epithelial A549 cells. We have observed that the AHR protein levels increase in the presence of chloroquine (CQ), an autophagy inhibitor, in a dose-dependent manner. Treatment with 6-aminonicotinamide (6-AN), a chaperone-mediated autophagy (CMA) activator, decreases AHR protein levels in a concentration-dependent and time-dependent manner. This decrease suppresses the ligand-dependent activation of the AHR target gene transcription, and can be reversed by CQ but not MG132. Knockdown of lysosome-associated membrane protein 2 (LAMP2), but not autophagy-related 5 (ATG5), suppresses the chloroquine-mediated increase in the AHR protein. AHR is resistant to CMA when its CMA motif is mutated. Suppression of the epithelial-to-mesenchymal transition in A549 cells is observed when the AHR gene is knocked out or the AHR protein level is reduced by 6-AN. Collectively, we have provided evidence supporting that AHR is continuously undergoing CMA and activation of CMA suppresses the AHR function in A549 cells. Full article

Glioblastoma (GBM) is the most lethal brain cancer, causing inevitable deaths of patients owing to frequent relapses of cancer stem cells (CSCs). The significance of the NOTCH signaling pathway in CSCs has been well recognized; however, there is no NOTCH-selective treatment applicable to patients with GBM. We recently reported that Jagged1 (JAG1), a NOTCH ligand, drives a NOTCH receptor-independent signaling pathway via JAG1 intracellular domain (JICD1) as a crucial signal that renders CSC properties. Therefore, mechanisms regulating the JICD1 signaling pathway should be elucidated to further develop a selective therapeutic regimen. Here, we identified annexin A2 (ANXA2) as an essential modulator to stabilize intrinsically disordered JICD1. The binding of ANXA2 to JICD1 prevents the proteasomal degradation of JICD1 by heat shock protein-70/90 and carboxy-terminus of Hsc70 interacting protein E3 ligase. Furthermore, JICD1-driven propagation and tumor aggressiveness were inhibited by ANXA2 knockdown. Taken together, our findings show that ANXA2 maintains the function of the NOTCH receptor-independent JICD1 signaling pathway by stabilizing JICD1, and the targeted suppression of JICD1-driven CSC properties can be achieved by blocking its interaction with ANXA2. Full article