Search Results (845)

Neurotensin (NTS) binds to the G protein-coupled receptors (GPCRs) NTSR1 and NTSR2. NTSR1 regulates transactivation of the EGFR, HER2, and HER3, but its effects on HER4 are unknown. By Western blot, NTSR1 and HER4 were present in six lung cancer cell lines examined. In NCI-H522 or NCI-H661 cells, adding NTS increased phosphorylation (P) of tyrosine (Y) 1284 on HER4. Because SR48692 antagonized NTS’s ability to increase P-HER4 or P-ERK, NTSR1 may play an important role in NSCLC. SR48692, HER4 siRNA, reactive oxygen species inhibitors, and the tyrosine kinase inhibitor ibrutinib inhibited NTS-induced P-HER4. Adding NTS to NCI-H661 cells increased the formation of HER4/HER4, HER4/ EGFR, and HER4/HER2 dimers. Adding NTS to NSCLC cells increased both P-ERK and P-AKT, which were inhibited by PD98059 and LY294002, respectively. The growth of NCI-H522 or NCI-H661 cells was stimulated by NTS or neuregulin 1 (NRG1), a HER4 ligand, but inhibited by SR48692 or ibrutinib. The results indicate that NTSR1 regulates HER4 transactivation, thereby increasing the proliferation of lung cancer cells. Full article

Cytochrome c (Cytc) tyrosine 48 (Y48) has been previously shown to be phosphorylated in bovine liver, and phosphomimetic substitution (Y48E) inhibits key functions of Cytc in vitro, including respiration and apoptosis. In this study, we investigated the effect of Y48 modification in a double-knockout cell culture model that stably expressed either unphosphorylated wild-type (WT) Cytc, control Y48F Cytc, or phosphomimetic Y48E Cytc. Our findings revealed that Y48E Cytc caused partial inhibition of mitochondrial respiration in intact cells, which corresponded with lower mitochondrial membrane potentials (ΔΨ_m) and reduced reactive oxygen species (ROS) production. When subjected to an oxygen–glucose deprivation/reoxygenation (OGD/R) model, which simulates ischemia/reperfusion injury, the Y48E phosphomimetic cell line showed lower ROS production compared to the unphosphorylated WT and Y48F Cytc cell lines, the latter of which generated higher levels of ROS upon reoxygenation. As a result, the Y48E Cytc cell line had significantly lower cell death rates when exposed to OGD/R, confirming the cytoprotective role of Y48 phosphorylation of Cytc. In summary, our research indicates that the loss of Y48 phosphorylation in Cytc during ischemia leads to reperfusion injury by driving maximum electron transport chain flow, hyperpolarization of ΔΨ_m, bursts of ROS, and death of cells through apoptosis. Full article

Background/Objectives: The clinical challenges of PARP inhibitors in ovarian cancer include the lack of effective maintenance regimens for homologous recombination proficiency (HRP) patients and the emergence of acquired resistance in initially responsive homologous recombination deficiency (HRD) patients. This study aims to explore the synergistic effect and molecular mechanism of the bispecific tyrosine phosphorylation-regulated kinase 1B (DYRK1B) inhibitor AZ191 combined with the PARP inhibitor Niraparib on high-grade serous ovarian cancer (HGSOC). Methods: This study first explored the expression and prognostic significance of DYRK1B in ovarian cancer through bioinformatics analysis. Subsequently, the therapeutic effect of the DYRK1B inhibitor AZ191 combined with Niraparib on HGSOC cells and organoids was evaluated by MTT examination. Flow cytometry and Western blot were used to investigate the synergistic mechanism between the two agents. Results: Bioinformatics analysis shows that the high expression of DYRK1B in serous ovarian cancer is associated with poor prognosis of the patients. The experiments in vitro have shown that the DYRK1B inhibitor AZ191 can enhance the therapeutic effect of Niraparib on HGSOC cells and organoids, whether HRD-positive or not. Mechanistic studies have shown that the combination of AZ191 and Niraparib can synergistically increase the accumulation of DNA damage, thereby intensifying the apoptosis of HGSOC cells. In addition, the combination therapy induces ferroptosis by inhibiting the Nrf2/SLC7A11/GPX4 axis, thereby exerting cytotoxic effects. Conclusions: Our results uncover a novel mechanism by which inhibiting DYRK1B enhances the anti-HGSOC efficacy of Niraparib and may offer a promising treatment strategy to improve the maintenance therapy in both HRD and HRP ovarian cancer patients. Full article

Background: YES1 is an Src family non-receptor tyrosine-protein kinase that regulates cell growth, migration, survival, and oncogenic signaling. Although YES1 activation mechanisms and substrates have been extensively studied, its phosphosite-specific regulation across diverse biological contexts remains poorly understood. Methods: We performed a large-scale integrative analysis of 3825 publicly available human mass spectrometry-based phosphoproteomic datasets to map YES1 phosphorylation events. Co-modulation, co-occurrence, evolutionary conservation, and disease-association analyses were conducted to characterize the functional and clinical relevance of site-specific YES1 phosphorylation. Results: Y426 emerged as the predominant YES1 phosphosite across diverse biological conditions, localized within the activation loop of the kinase domain and conserved across Src family kinases. Co-modulation analysis identified 421 positively and 102 negatively associated phosphosites enriched in biological processes related to cell cycle regulation, transcription, cytoskeletal remodeling, apoptosis, and carcinogenesis. Among these high-confidence protein phosphosites, we identified 24 binary interactors, 5 upstream regulators, and 8 candidate downstream substrates. Comparison with DisGeNet cancer biomarkers showed overlap between YES1-associated phosphoproteomic signatures and site-specific oncogenic markers across multiple cancers, such as breast cancer, colorectal cancer, leukemia, and lung adenocarcinoma. Conclusions: This study provides a systems-level, phosphosite-focused view of YES1 signaling and supports a central regulatory role for Y426 within global phosphoregulatory and cancer-associated networks. Full article

Cognitive impairment in schizophrenia remains insufficiently addressed by existing treatments. Current FDA-approved therapies primarily modulate neurotransmitter systems, resulting in incomplete symptom control and substantial adverse effects. There is therefore a critical need for therapeutic strategies that more directly address the intracellular signaling mechanisms underlying synaptic dysfunction and cognitive deficits in schizophrenia. Protein phosphatases represent an essential but historically underexplored class of signaling enzymes that regulate phosphorylation-dependent control of synaptic receptor trafficking, plasticity, and neuronal circuit function. Although multiple phosphatases have been implicated in schizophrenia through genetic, post-mortem, and functional studies, their therapeutic targeting has been limited by challenges related to selectivity, cellular permeability, and pleiotropy. Here, we review the etiology of schizophrenia and limitations of current pharmacological approaches, synthesize evidence linking specific protein phosphatases to schizophrenia pathophysiology, and discuss emerging strategies, including allosteric modulation and targeted protein degradation, that may enable selective intervention in phosphatase-driven signaling pathways. We highlight the striatal-enriched tyrosine phosphatase STEP (PTPN5) as a case study illustrating how selective phosphatase modulation can restore synaptic signaling in schizophrenia-relevant models. Full article

Background/Objectives: Protein kinases play crucial roles in signal transduction pathways that regulate growth and differentiation in Trypanosoma cruzi. These protein kinases are attractive targets to develop new drugs to treat Chagas disease. Methods: We used several protein kinase inhibitors targeting the p38 MAPK, MEK, and ERK pathways to evaluate their effects on the in vivo phosphorylation status of T. cruzi proteins, particularly DNA polymerase beta (TcPolβ). We also used Genistein, a protein tyrosine kinase inhibitor, to assess its effects on global protein phosphorylation and TcPolβ phosphorylation. Also, we investigated the effect of oxidative stress on global tyrosine phosphorylation. Finally, we determined the phosphorylation sites on TcPolβ by the protein kinases TcPKC2 and TcWee570 in vitro. Results: p38 MAPK and MEK protein kinase inhibitors inhibited approximately 50% of the Ser/Thr phosphorylation of TcPolβ. Genistein inhibited both Ser/Thr and Tyr phosphorylation of several polypeptides in epimastigotes. Oxidative stress increases global Tyr phosphorylation by about twofold and also TcPolβ phosphorylation. TcPKC2 and TcWee570 were able to phosphorylate TcPolβ at both Ser/Thr and Tyr residues. Conclusions: Small-molecule protein kinase inhibitors can affect the phosphorylation status of TcPolβ in vivo. Since Genistein can inhibit both Ser/Thr and Tyr protein phosphorylation, and TcPKC2 and TcWee570 can phosphorylate both Ser/Thr and Tyr residues, it suggests the existence of dual protein kinases in T. cruzi. However, this possibility must be further studied. Full article

Prolonged occupational exposure to oil mist particulate matter (OMPM) poses health risks, yet its neurotoxic effects and underlying mechanisms remain poorly understood. Here, OMPM generated from turbine oil commonly used in occupational labor environments was used to expose rats. The rats were divided into the control and OMPM groups. Following 42 days of exposure, a multidimensional assessment was performed using untargeted metabolomics, phosphoproteomics, behavioral testing, hematoxylin–eosin (HE) staining, transmission electron microscopy (TEM), colorimetric assays, enzyme-linked immunosorbent assay, and Western blotting (WB) to evaluate metabolic alterations, protein phosphorylation, and tissue integrity in the striatum. Integrated omics analyses revealed that differentially phosphorylated proteins and metabolites were remarkably enriched in dopaminergic synapse, Parkinson’s disease, and amphetamine addiction pathways (FDR < 0.05), with a regulatory axis involving L-tyrosine, tyrosine hydroxylase (TH), and dopamine (DA) identified. OMPM-exposed rats exhibited depression- and anxiety-like behaviors, alongside striatal pathological and ultrastructural damage. Biochemical analyses showed elevated malondialdehyde and reactive oxygen species levels; reduced superoxide dismutase, glutathione, and glutathione peroxidase activities and total antioxidant capacity; increased glutathione disulfide and inducible nitric oxide synthase expression; and decreased DA and L-tyrosine levels. Additionally, proinflammatory mediators (IL-1β, IL-6, TNF-α, MCP-1, and PGD₂) were significantly upregulated in the striatum. WB analysis further confirmed significant reductions in the relative phosphorylation levels of key regulators in dopaminergic and calcium signaling pathways, including CALM3, CaMK2b, GSK-3β, PRKCG, and TH. Collectively, these findings reveal critical molecular and biochemical alterations in the rat striatum following OMPM exposure and provide a mechanistic basis for understanding depression-like behaviors associated with prolonged OMPM exposure in occupational workers. Full article

Background: Decreased dopaminergic cells and tyrosine hydroxylase (TH) in the substantia nigra (SN) lead to Parkinson’s disease (PD); but its cause remains unknown. PD is characterized by α-synuclein (α-syn) accumulation in Lewy bodies; most of which is phosphorylated at Ser129 (pSer129 α-syn). Serping1 is an important gene for controlling blood vessel maintenance; including the process of inflammation. Methods: Increased expression of Serping1 affects dopaminergic cell death in the SN of a chronic PD mouse model induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP); and Serping1 siRNA treatment has a therapeutic effect in this model. Results: We demonstrated that this treatment shows a normal status in the motor ability test and TH level in the SN and striatum. Serping1 siRNA was found to react to decreased Serping1 levels in the SN. In the pSer129-α-syn level of the SN region; Serping1 siRNA had a greater positive effect on PD than N-acetylcysteine by inhibiting pSer129-α-syn formation. Cyclooxygenase-2 and inducible nitric oxide synthase levels were decreased by Serping1 siRNA treatment; thereby indicating its effect on inflammation. Conclusions: Our findings suggest that Serping1 siRNA may represent a potential therapeutic approach for PD; warranting further investigation. Full article

Signal transducer and activator of transcription 3 (STAT3) is a central oncogenic hub in several tumors including the Triple-Negative Breast Cancer (TNBC) subtype, where its constitutive activity supports proliferation, metabolic flexibility, tumor progression, immune evasion, and therapeutic resistance. Therapeutic development has largely focused on canonical STAT3 activation driven by tyrosine 705 phosphorylation (p-Y705), which enables dimerization and transcriptional programs. However, accumulating evidence indicates that phosphorylation at serine 727 (p-S727) defines a functionally distinct STAT3 axis, underpinning non-canonical activities across extranuclear compartments that include mitochondria and endoplasmic reticulum/mitochondria-associated membranes. In TNBC, p-S727 STAT3 is frequently prevalent and may sustain oncogenic signaling when p-Y705 is low or pharmacologically suppressed, contributing to metabolic rewiring, redox control, apoptosis resistance, and metastatic fitness. Here, we review the mechanistic basis and clinical correlations of STAT3 p-S727 across cancers with emphasis on TNBC, and discuss how compartmentalized STAT3 pools may integrate kinase signaling, nutrient sensing, and stress responses. We also summarize emerging therapeutic strategies that modulate p-S727—often in conjunction with p-Y705—highlighting proof-of-concept for dual targeting or specific p-S727 to overcome limitations of Y705-centric approaches. Finally, we propose that integrating p-S727/p-Y705 distribution and activity into patient stratification could improve the efficacy–toxicity balance of STAT3-directed therapies in TNBC. Full article

Multiple myeloma (MM) is characterised by the clonal expansion of plasma cells in the bone marrow followed by end-organ damage. Despite a significant increase in the five-year survival rate in recent years, MM is still considered an incurable disease as patients will repeatedly relapse and develop resistance to standard-of-care therapies. A central theme for the personalization of MM therapy is understanding the biological mechanisms of drug resistance and identifying clinically relevant biomarkers of therapeutic response. Highly effective protocols for the enrichment of phosphorylated peptides followed by high-resolution mass spectrometry makes possible the quantitation of thousands of site-specific phosphorylation events, principally on serine, threonine or tyrosine residues. In this study, phosphoproteomic analysis of 20 MM patient cell lysates was performed, stratified based on their ex vivo drug response profiles to Bortezomib and Lenalidomide, two of the most foundational therapeutic agents in the management of MM. In this study, patients who are highly sensitive to these drugs show increased phosphorylation of proteins concerned with translation and RNA processing including the spliceosome, RNA transport and RNA binding pathways, while highly resistant patients demonstrated an increased phosphorylation of proteins involved with tight junctions, the Rap1 signalling pathway and the phosphatidylinositol signalling system. This study has established a phosphoproteomic dataset displaying unique phosphorylation signatures associated with drug sensitivity in MM patient plasma cells. The identification of phosphorylation signatures associated with drug resistance provides the foundation for further exploration of these mechanisms and associated signalling pathways to further characterise drug resistance mechanisms in MM and identify promising biomarkers of therapeutic response and targets for drug re-sensitization in MM. Full article

Background/Objectives: Alzheimer’s disease (AD) is characterized by progressive neurodegeneration driven by interconnected mechanisms, including oxidative stress, mitochondrial dysfunction, neuroinflammation, synaptic impairment, and abnormal protein aggregation. MicroRNAs (miRNAs) have emerged as post-transcriptional regulators of these complex pathways; however, efficient delivery remains a major limitation. Small extracellular vesicles (sEVs) have been proposed as biologically compatible carriers for miRNA delivery. Methods: In this study, milk-derived sEVs were isolated, characterized, and loaded with microRNA-137-5p (miR-137-5p). Their effects were evaluated in an amyloid-β (Aβ)-induced in vitro AD model using SH-SY5Y human neuroblastoma cells. Oxidative stress markers, including reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), lactate dehydrogenase (LDH), and glutathione peroxidase 1 (GPX1), were assessed. Inflammation- and neuroprotection-related gene expression analyses included intercellular adhesion molecule 1 (ICAM1), tumor necrosis factor alpha (TNF-α), and brain-derived neurotrophic factor (BDNF). Cytoskeletal injury was evaluated using neurofilament light chain (NfL). Mitochondrial stress markers included cytochrome c (Cyt-c), 8-hydroxy-2′-deoxyguanosine (8-OHdG), PTEN-induced kinase 1 (PINK1), dynamin-1-like protein (DNM1L), and mitochondrial transcription factor A (TFAM). Synaptic and extracellular matrix-associated proteins, including complexin-2 (CPLX2), SPARC-related modular calcium-binding protein 1 (SMOC1), and receptor tyrosine kinase-like orphan receptor 1 (ROR1), as well as AD-related biomarkers, including total tau, phosphorylated tau at threonine 181 (pTau-181), phosphorylated tau at threonine 217 (pTau-217), and amyloid-β 1–40 (Aβ1–40), were evaluated using molecular and biochemical approaches. Results: Aβ exposure was associated with increased oxidative stress, inflammatory activation, mitochondrial and cytoskeletal alterations, synaptic-related disturbances, and elevations in tau- and amyloid-associated proteins. Treatment with unloaded sEVs was associated with partial modulation of several parameters, whereas miR-137-5p-loaded sEVs were consistently associated with normalization of multiple pathological markers toward control levels. Conclusions: These findings indicate that miR-137-5p-enriched sEVs may represent a useful experimental platform for multi-target modulation of AD-related cellular alterations. Further mechanistic and in vivo studies are required to clarify translational relevance. Full article

Objective: Exposure to rotenone, a naturally occurring pesticide, has been linked to an increased risk of developing Parkinson’s disease (PD). Rotenone strongly inhibits complex I of the mitochondrial respiratory chain, inducing oxidative stress both in vitro and in vivo, ultimately leading to cell death. The objective of this study was to evaluate the cytoprotective effects of the multifunctional agonist D-512 against rotenone-induced toxicity in neuronal PC12 and dopaminergic MN9D cell lines. Methods: Various cell-based assays, including cell viability, antioxidant activity, caspase-mediated apoptosis, and other related assays, were performed. Results: Rotenone was found to be toxic to both dopaminergic MN9D cells and neuronal PC12 cells. However, treatment with D-512 protected both cell types from rotenone-induced toxicity in a dose-dependent manner. Rotenone-induced impairment of mitochondrial membrane potential and increased production of reactive oxygen species were reversed by D-512 treatment. Furthermore, rotenone-induced caspase-mediated apoptotic signaling in MN9D cells was inhibited by D-512. In addition, D-512 restored levels of phosphorylated tyrosine hydroxylase in rotenone-exposed cells across various doses, indicating protection of the dopaminergic system. Finally, rotenone-induced activation of phosphorylated ERK was reversed by D-512 treatment, further supporting its neuroprotective potential. Conclusions: This study demonstrates the ability of D-512 to reverse the toxic effects of rotenone across multiple experimental models. The data presented here are consistent with previously reported neuroprotective properties of D-512. The multifunctional nature of D-512, which combines potent dopamine agonist activity with neuroprotective and other beneficial properties, may address therapeutic needs in PD beyond symptomatic relief and could have potential application across PD subgroups as part of a personalized therapeutic approach. Full article

Mitochondrial dysfunction contributes to diabetic cardiomyopathy, yet how genetic predisposition and diet interact to reshape cardiac metabolism in diabetic and prediabetic states remains unclear. The Cohen diabetic rat model, comprising diabetes-resistant (CDr) and diabetes-sensitive (CDs) strains, provides a unique platform to dissect this interplay. Here, we present an integrative global proteomic and bioenergetic characterization of cardiac tissue from CDr and CDs rats fed either a regular or a diabetogenic diet. Proteomic pathway mapping revealed downregulation of cytochrome c oxidase (CcO) subunits, strain-dependent rewiring of fatty-acid oxidation pathways, and CcO subunits switch from “heart-type” to “liver-type” isoforms in the sensitive strain. These changes were accompanied by impaired mitochondrial respiration, ATP depletion, and disruption of mitochondrial quality-control mechanisms, together with increased accumulation of tyrosine 304 phosphorylation of cytochrome c oxidase subunit I, indicative of inflammation-driven regulatory inhibition in a diet-specific manner. These findings establish an understanding of how genetic susceptibility and diet contribute to cardiac mitochondrial dysfunction in the Cohen diabetic rat model. Full article

Background: Fibroblast growth factor receptors (FGFRs) play a crucial role in tissue homeostasis and organ development by regulating cellular processes, including proliferation, differentiation, and survival. Dysregulation of FGFRs contributes to developmental disorders and carcinogenesis. As membrane-bound receptors, they represent promising targets for therapeutic intervention and drug development. Methods: This study employed a systematic in silico analysis of publicly available phosphoproteomics datasets to provide a comprehensive overview of the phosphorylation regulatory network of the FGFR family. Results: We identified predominant phosphosites in FGFR1-4 that exhibited differential abundance across diverse experimental conditions, specifically, Y653 in FGFR1; S453, Y586, Y656, and Y657 in FGFR2; S444 and S445 in FGFR3; and S573 in FGFR4. Our analysis identified 32 and 89 significantly co-modulated phosphosites on other proteins with FGFR3 and FGFR4, respectively. Beyond the upstream kinases from the FGFR family, we also identified MAPK1 as a potential upstream kinase of FGFR4. Furthermore, disease enrichment analysis revealed that proteins co-modulated with FGFR3 were primarily involved in skeletal developmental disorders, such as brachydactyly, short toe, and syndactyly of fingers, whereas those associated with FGFR4 were linked to various cancers. Conclusions: Our findings highlight key disease-associated phosphosites within the FGFRs and offer a foundation for advancing phosphosite-focused therapeutic research. Full article

Background: Microplastics and nanoplastics, as pervasive and persistent environmental pollutants, are raising growing concerns regarding their potential risks to reproductive health, particularly pregnancy outcomes. Although the reproductive toxicity of polystyrene nanoplastics (PS-NPs) has been reported, the specific mechanisms underlying their effects on placental development and offspring health following gestational exposure remain unclear. Method: This study aimed to investigate the effects of gestational exposure to PS-NPs of different sizes (50 and 200 nm) and concentrations (1, 3, and 10 mg/mL) on placental function and embryonic development in ICR mice. An exposure model was established via tail vein injection, and samples were collected on embryonic Day 14.5 (E14.5). Results: the exposed groups tended towards increased embryo weight, embryo length, and embryo head circumference. Transcriptomic analysis revealed that PS-NP exposure significantly downregulated the expression of Ndufa5 (a subunit of mitochondrial respiratory chain complex I) and mt-CO1 (a core subunit of complex IV), but upregulated the expression of the genes Cldn1 (tight junction protein) and Erbb3 (receptor tyrosine kinase) in the placenta. Differentially expressed genes were enriched primarily in pathways related to oxidative phosphorylation, the tricarboxylic acid (TCA) cycle, and ErbB signalling. Conclusions: These changes collectively led to decreased mitochondrial ATP production, increased oxidative stress in the placenta, and potentially altered placental barrier function and trophoblast cell proliferation signalling. This study reveals a novel mechanism by which PS-NPs disrupt placental development and embryonic growth through impairment of placental energy metabolic homeostasis and key signalling pathways, thus providing crucial experimental evidence for assessing the reproductive and developmental toxicity of nanoplastics. Full article