Search Results (70)

Pancreatic β-cells maintain glucose homeostasis through tightly regulated insulin biosynthesis, storage, and secretion. To prevent pathological accumulation of excess or aging secretory granules (SGs), β-cells use crinophagy, a selective lysosomal degradation pathway in which mature insulin-containing granules fuse directly with lysosomes to form hybrid organelles termed crinosomes. Crinophagy was historically considered a simple mechanism for discarding obsolete, aged SGs. The acidic, protease-rich environment of crinosomes is proposed to generate unconventional insulin-derived epitopes through cathepsin-mediated proteolysis and transpeptidation reactions. These cryptic epitopes, which include hybrid insulin peptides (HIPs) resulting from the covalent fusion of insulin fragments with peptides from co-resident granule proteins, are largely absent from the thymic epitope repertoire. This creates a “peripheral–thymic mismatch” that allows autoreactive CD4+ T cells to escape central tolerance, ultimately driving β-cell destruction in type 1 diabetes (T1D). Recent studies demonstrate that pharmacological or genetic inhibition of crinophagy reduces crinosome abundance, narrows the pathogenic epitope repertoire, and delays the onset of diabetes in preclinical models. In type 2 diabetes (T2D), a related pathway termed stress-induced nascent granule degradation (SINGD) diverts newly synthesized insulin granules to lysosomes under glucolipotoxic conditions, contributing to insulin depletion and progressive β-cell failure. This review summarizes the current understanding of the molecular mechanisms behind crinophagy. It discusses its two main functions: maintaining physiological quality control and generating pathological antigens. Additionally, the review explores how crinophagy interacts with other cellular stress pathways and highlights new therapeutic strategies aimed at targeting this process to protect pancreatic β-cell function and potentially prevent or delay diabetes. Full article

The picornavirus 3CD protein is a precursor to the 3C main protease and the 3D RNA-dependent RNA polymerase. In addition to its functions in proteolytic processing of the virus polyprotein and cleavage of key host factors, the 3C domain interacts with cis-acting replication elements (CREs) within the viral genome to regulate replication and translation events. We investigated the molecular determinants of RNA binding to 3C using a wide range of biophysical and computational methods. These studies showed that 3C binds to a broad spectrum of RNA oligonucleotides, displaying minimal sequence and structure dependence, at least for these shorter RNAs. However, they also uncovered a novel aspect of these interactions, that is, 3C-RNA binding can induce liquid–liquid phase separation (LLPS), with 3CD–RNA interactions likewise leading to LLPS. This may be a general phenomenon for other 3C and 3C-like proteases and polyproteins incorporating 3C domains. These findings have potential implications in understanding virally induced apoptosis and the control of stress granules, which involve LLPS and include other proteins with known interactions with 3C/3CD. Full article

Rice sheath rot has progressively developed into a growing threat to global rice production, particularly in intensively managed systems conducive to disease development. Therefore, accurate identification of the causal pathogen and the development of sustainable management strategies represent urgent scientific requirements. In this study, we isolated the causal organism of rice sheath rot from infected rice tissues and identified it as Fusarium verticillioides based on multi-locus sequence analysis. Eight endophytic bacterial strains were recovered from healthy rice root systems. Among the isolates, Bacillus velezensis isolate 7-A exhibited the strongest antifungal activity against F. verticillioides. This isolate demonstrated broad-spectrum antifungal activity, with inhibition rates ranging from 54.8% to 71.8%. Phylogenetic analysis based on 16S rRNA and gyrB gene sequences identified it as B. velezensis. Further characterization revealed that B. velezensis 7-A is capable of secreting proteases and synthesizing siderophores. The filtered liquid from sterile fermentation markedly inhibited the growth of mycelium in F. verticillioides and induced marked morphological abnormalities. Liquid LC-MS analysis identified multiple antifungal active substances, including camphor, ginkgolides B, salicin, cinnamic acid, hydroxygenkwanin, stearamide, β-carotene, and others. A pot experiment demonstrated that the fermentation broth of B. velezensis 7-A effectively suppressed the occurrence of rice sheath rot, achieving a relative control efficacy of 61.3%, which is comparable to that of a 10% carbendazim water-dispersible granule (WDG). Additionally, isolate 7-A enhances plant disease resistance by activating the activities of key defense enzymes. These findings provide preliminary insights into its potential application in integrated and sustainable disease management programs. Full article

The heparin-binding protein (HBP) is an enzymatically inactive protein of the serine protease family that plays an important role in host response to stress, especially infection and sepsis. It is produced by activated neutrophils due to a variety of stimuli and is part of the immune response that leads to macrophage, lymphocyte, and neutrophil activation and monocyte adhesion. Its most common repository is the azurophilic granules of the neutrophils. HBP has been studied as a biomarker for several infections, including central nervous system infection, respiratory tract infection, and urinary tract infection, and in several settings, including the Emergency Department and Intensive Care Unit, with promising results. As a biomarker for infection and sepsis, HBP has been compared to other commonly used biomarkers such as Neutrophil to Lymphocyte Ratio, White Blood Count, C-reactive protein, and Procalcitonin, with at least comparable performance. Its sharp increase is promising for the early detection of sepsis. The ability to differentiate inflammatory conditions from infections and bacterial from non-bacterial causes of infection has also been demonstrated. The sepsis-related organ damage, as it is represented by the Sequential Organ Failure Assessment score, can also be reflected by the proportional increase in HBP. Consequently, HBP could be a helpful and promising biomarker for sepsis and infection. Full article

Neurodegeneration is increasingly recognized not as a linear trajectory of protein accumulation, but as a multidimensional collapse of biological organization—spanning intracellular signaling, transcriptional identity, proteostatic integrity, organelle communication, and network-level computation. This review intends to synthesize emerging frameworks that reposition neurodegenerative diseases (ND) as progressive breakdowns of interpretive cellular logic, rather than mere terminal consequences of protein aggregation or synaptic attrition. The discussion aims to provide a detailed mapping of how critical signaling pathways—including PI3K–AKT–mTOR, MAPK, Wnt/β-catenin, and integrated stress response cascades—undergo spatial and temporal disintegration. Special attention is directed toward the roles of RNA-binding proteins (e.g., TDP-43, FUS, ELAVL2), m6A epitranscriptomic modifiers (METTL3, YTHDF1, IGF2BP1), and non-canonical post-translational modifications (SUMOylation, crotonylation) in disrupting translation fidelity, proteostasis, and subcellular targeting. At the organelle level, the review seeks to highlight how the failure of ribosome-associated quality control (RQC), autophagosome–lysosome fusion machinery (STX17, SNAP29), and mitochondrial import/export systems (TIM/TOM complexes) generates cumulative stress and impairs neuronal triage. These dysfunctions are compounded by mitochondrial protease overload (LONP1, CLPP), UPR maladaptation, and phase-transitioned stress granules that sequester nucleocytoplasmic transport proteins and ribosomal subunits, especially in ALS and FTD contexts. Synaptic disassembly is treated not only as a downstream event, but as an early tipping point, driven by impaired PSD scaffolding, aberrant endosomal recycling (Rab5, Rab11), complement-mediated pruning (C1q/C3–CR3 axis), and excitatory–inhibitory imbalance linked to parvalbumin interneuron decay. Using insights from single-cell and spatial transcriptomics, the review illustrates how regional vulnerability to proteostatic and metabolic stress converges with signaling noise to produce entropic attractor collapse within core networks such as the DMN, SN, and FPCN. By framing neurodegeneration as an active loss of cellular and network “meaning-making”—a collapse of coordinated signal interpretation, triage prioritization, and adaptive response—the review aims to support a more integrative conceptual model. In this context, therapeutic direction may shift from damage containment toward restoring high-dimensional neuronal agency, via strategies that include the following elements: reprogrammable proteome-targeting agents (e.g., PROTACs), engineered autophagy adaptors, CRISPR-based BDNF enhancers, mitochondrial gatekeeping stabilizers, and glial-exosome neuroengineering. This synthesis intends to offer a translational scaffold for viewing neurodegeneration as not only a disorder of accumulation but as a systems-level failure of cellular reasoning—a perspective that may inform future efforts in resilience-based intervention and precision neurorestoration. Full article

Background/Objectives: Combination therapies using traditional Chinese medicine and Western drugs have gained attention for their enhanced therapeutic effects and reduced side effects. Toujie Quwen Granules (TQG), known for its antiviral properties, particularly against respiratory viruses, could offer new treatment strategies when combined with antiviral drugs like arbidol, especially for diseases such as Coronavirus disease. This study investigates the synergistic mechanisms between arbidol and components from TQG against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease (M^pro). Methods: We identified compounds from TQG via existing data. Multi-ligand molecular docking, pharmacokinetic/toxicity screening, and preliminary simulations were performed to assess potential synergistic compounds with arbidol. UPLC-Q-Exactive Orbitrap-MS verified the presence of these compounds. Extended simulations and in vitro assays, including Luciferase and surface plasmon resonance, validated the findings. Results: Five compounds interacted with arbidol in synergy based on docking and preliminary dynamics simulation results. Only Baicalein (HQA004) could be identified in the herbal remedy by untargeted metabolomics, with ideal pharmacokinetic properties, and as a non-toxic compound. Extended simulations revealed that HQA004 enhanced arbidol’s antiviral activity via a “Far” Addition Mechanism #2, with an optimal 2:1 arbidol:HQA004 ratio. The movements of arbidol (diffusion and intramolecular conformational shifts) in the system were significantly reduced by HQA004, which may be the main reason for the synergism that occurred. In vitro experiments confirmed an increased inhibition of M^pro by the combination. Conclusions: HQA004 demonstrated synergistic potential with arbidol in inhibiting M^pro. The development of combination therapies integrating Western and herbal medicine is supported by these findings for effective antiviral treatments. Full article

Toona sinensis (A. Juss.) Roem, classified under the Toona genus of the Meliaceae family, is a fast-growing, woody species endemic to China, valued as both a vegetable crop and medicinal plant. Its seeds achieve rapid germination through a cascade of interconnected physiological, metabolic, and hormonal adaptations. Initially, physiological hydration is driven and accelerated by only two distinct phases of water imbibition. This hydration surge triggers storage reserve mobilization, with soluble sugars, proteins, and lipids undergoing rapid degradation during imbibition, while starch catabolism proceeds gradually—a pattern mirrored by progressive increases in enzymatic activities (amylase, protease, and acid phosphodiesterase (ACP)) that correlate with reserve reallocation. Concurrently, a metabolic shift from glycolysis to the pentose phosphate pathway (PPP) optimizes energy utilization, supporting germination acceleration. These biochemical changes are orchestrated by hormonal coordination: elevated gibberellin A₃ (GA₃), zeatin riboside (ZR), and indole-3-acetic acid (IAA) levels, coupled with rising GA₃/ABA, IAA/ABA, and ZR/ABA ratios, temporally aligned with germination progression. Finally, structural evidence confirms successful germination completion, as cotyledon lipid droplet breakdown and starch granule synthesis directly correlate with embryonic elongation. Together, these mechanisms underscore T. sinensis’ adaptive strategy, integrating physiological plasticity, metabolic flexibility, and endocrine precision to ensure efficient germination. Full article

The coronavirus disease 2019 (COVID-19) pandemic has been linked to long-term neurological effects with multifaceted complications of neurodegenerative diseases. Several studies have found that pathological changes in transactive response DNA-binding protein of 43 kDa (TDP-43) are involved in these cases. This review explores the causal interactions between severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and TDP-43 from multiple perspectives. Some viral proteins of SARS-CoV-2 have been shown to induce pathological changes in TDP-43 through its cleavage, aggregation, and mislocalization. SARS-CoV-2 infection can cause liquid−liquid phase separation and stress granule formation, which accelerate the condensation of TDP-43, resulting in host RNA metabolism disruption. TDP-43 has been proposed to interact with SARS-CoV-2 RNA, though its role in viral replication remains to be fully elucidated. This interaction potentially facilitates viral replication, while viral-induced oxidative stress and protease activity accelerate TDP-43 pathology. Evidence from both clinical and experimental studies indicates that SARS-CoV-2 infection may contribute to long-term neurological sequelae, including amyotrophic lateral sclerosis-like and frontotemporal dementia-like features, as well as increased phosphorylated TDP-43 deposition in the central nervous system. Biomarker studies further support the link between TDP-43 dysregulation and neurological complications of long-term effects of COVID-19 (long COVID). In this review, we presented a novel integrative framework of TDP-43 pathology, bridging a gap between SARS-CoV-2 infection and mechanisms of neurodegeneration. These findings underscore the need for further research to clarify the TDP-43-related neurodegeneration underlying SARS-CoV-2 infection and to develop therapeutic strategies aimed at mitigating long-term neurological effects in patients with long COVID. Full article

Vertebrate mast cells are the first cells to initiate the inflammatory response. The origin of these highly specialised innate immunity cells in chordates is an intriguing unanswered question, and tunicates represent the best candidates to address this question for their close relationship with vertebrates. In the colonial ascidian Botrylloides leachii, a particular cell type circulates in the haemolymph, namely, ‘granular cell’, which is a distinct immunocyte from both phagocytic and cytotoxic lines. Like mast cells and unlike basophils, granular cells were labelled with anti-c-kit antibody on their plasmalemma and exhibited a high content of heparin in their granules, as revealed by various histochemical techniques. Immunohistochemistry revealed the presence of heparin and histamine inside the same granules resembling the granules of mast cells. Histoenzymatic assays revealed the presence of mast cell enzymes such as β-glucuronidase, arylsulphatase, chloroacetyl esterase, and proteases. These cells degranulated after exposure to bacteria, compound 48/80, or heterologous plasma. During exposure to bacteria, they crowd into the perivisceral sinus and then infiltrate the epithelium of the postbranchial gut, where they release the content of their granules, a behaviour remarkably similar to that of the gastric leukopedesis of mast cells. Full article

Tryptase is a tetrameric serine protease and a key component of mast cell granules. Here, we explored an integrated approach to characterize tryptase ligands, combining novel experimental binding studies using Surface Plasmon Resonance, with in silico analysis through the Exscalate platform. For this, we focused on three inhibitors previously reported in the literature, including a bivalent inhibitor and its corresponding monovalent compound. All three ligands showed concentration-dependent binding to immobilized human tryptase with the bivalent inhibitor showing the highest affinity. Furthermore, Rmax values were similar, indicating that the compounds occupy all four binding pockets of the tryptase tetramer. This hypothesis was supported by in silico computational analysis that revealed the binding mode of the monovalent ligand, one in each monomer pocket, compared with crystal structure of the bivalent one, which simultaneously occupies two binding pockets. Additionally, we solved the 2.06 Å X-ray crystal structures of human Tryptase beta-2 (hTPSB2), in both its apo form and in complex with compound #1, experimentally confirming the binding mode and the key molecular interactions predicted by docking studies for this compound. This integrated approach offers a robust framework for elucidating both the strength and mode of interaction of potential tryptase inhibitors. Full article

Solid-state fermentation (SSF) has emerged as an effective method for wheat bran valorization, providing advantages like cost reduction, decreased water usage, and enhanced product quality. In this study, wheat bran was fermented using Rhizopus oryzae to evaluate the extraction yield of soluble dietary fiber, the activities of protease and amylase, and the physicochemical characteristics of wheat bran during SSF. The findings demonstrated that the maximum yield of soluble dietary fiber was achieved after 120 h of fermentation at a moisture content of 55%. Simultaneously, protease activity peaked at 45% moisture content after 120 h, while amylase activity was maximized at 55% moisture content after 96 h. The microstructure result indicated that most of the starch granules degraded after 144 h of fermentation at a moisture content of 55%, exhibiting a smooth outer layer of wheat bran. Furthermore, fermented bran showed a significant rise in total phenols, peaking at 96 h at a moisture content of 55%. Flavonoid content also reached its maximum after 72 h of fermentation at 55% moisture content. The content of alkylresorcinols in fermented wheat bran changed slightly under different moisture content and fermentation time conditions, which was consistent with the change in pH value. The DPPH radical scavenging rate was optimal when the moisture content was 55% after 96 h. The ABTS radical scavenging rate, hydroxyl radical scavenging rate, and reducing ability were optimal at 55% moisture content after 120 h. These findings demonstrate that the optimal conditions for the SSF of wheat bran using Rhizopus oryzae involve maintaining the moisture at 55%, suggesting that this method is effective for enhancing the value of wheat bran. Full article

Quantitative transcriptomics offers a new way to obtain a detailed picture of freshly isolated cells. By direct isolation, the cells are unaffected by in vitro culture, and the isolation at cold temperatures maintains the cells relatively unaltered in phenotype by avoiding activation through receptor cross-linking or plastic adherence. Simultaneous analysis of several cell types provides the opportunity to obtain detailed pictures of transcriptomic differences between them. Here, we present such an analysis focusing on four human blood cell populations and compare those to isolated human skin mast cells. Pure CD19⁺ peripheral blood B cells, CD14⁺ monocytes, and CD4⁺ and CD8⁺ T cells were obtained by fluorescence-activated cell sorting, and KIT+ human connective tissue mast cells (MCs) were purified by MACS sorting from healthy skin. Detailed information concerning expression levels of the different granule proteases, protease inhibitors, Fc receptors, other receptors, transcription factors, cell signaling components, cytoskeletal proteins, and many other protein families relevant to the functions of these cells were obtained and comprehensively discussed. The MC granule proteases were found exclusively in the MC samples, and the T-cell granzymes in the T cells, of which several were present in both CD4⁺ and CD8⁺ T cells. High levels of CD4 were also observed in MCs and monocytes. We found a large variation between the different cell populations in the expression of Fc receptors, as well as for lipid mediators, proteoglycan synthesis enzymes, cytokines, cytokine receptors, and transcription factors. This detailed quantitative comparative analysis of more than 780 proteins of importance for the function of these populations can now serve as a good reference material for research into how these entities shape the role of these cells in immunity and tissue homeostasis. Full article

The present study was carried out to evaluate the survival, growth, and digestive ontogeny of C. striatus larvae fed with different experimental diets from 4 days post-hatch (dph) to 32 dph at three-day intervals. A total of 24,000 larvae, with 1600 larvae per tank in triplicate and an initial mean weight of 0.64 ± 0.01 mg at 4 days post hatch (dph) were subjected to five different early weaning diets, namely Artemia nauplii (T1), co-feed diet comprising Artemia nauplii and formulated micro diet (T2), formulated micro diet (T3), formulated micro diet with protease supplementation (T4), and a commercial diet (T5). All the early weaning diets significantly affected the survival, growth, and ontogeny of the digestive system. Initially at 8 dph, C. striatus fed with T1 showed better survival and growth performance compared to other treatments. By 12 dph, the larvae fed with T1 and T2 showed similar results in terms of survival and growth performance, outperforming other treatments. However, the larvae fed with T2 and T4 outperformed T1 in survival and growth performance at 16 dph. By 24–32 dph, the larvae fed with all treatments met the basic nutritional needs for survival, with T4 fed larvae showing better growth compared to other treatments. At the end of the trial, cumulative mortality was lowest in larvae fed with T1 and highest in the larvae fed with T3 and T5. Similarly, the larvae fed with T4 showed significantly higher weight gain, specific growth rate (SGR), and average daily growth (ADG), while T1 fed larvae exhibited better feed conversion ratio (FCR) and protein efficiency ratio (PER). The enzyme activity fluctuated throughout the experimental duration. Lavae fed with T1 and T2 showed higher enzyme activities initially. However, T4 fed larvae showed higher trypsin and chymotrypsin specific activity at 16 dph along with well-developed intestinal folds with dense microvilli, higher pepsin-specific activity at 20 dph onwards with fully developed gastric glands and thicker gastric mucosal epithelium, and higher amylase and lipase activity at 16 dph with large and prominent zymogen granules in the exocrine pancreas. Peaking at 4 dph, the activity of protein metabolic enzymes (AST and ALT) sharply declined at 8 dph and increased until 32 dph. Larvae fed with T1 showed higher AST and ALT activity along with increased lipid deposits, followed by those fed with T2 and the larvae fed with T4 showing higher activity without fat accumulation but significantly lower than those fed T1 and T2. From the present research findings, it is recommended to initiate weaning for Channa striatus larvae with Artemia nauplii (from 4 dph to 8 dph) followed by a co-feeding regime (Artemia nauplii and formulated diet) between 9 and 16 dph and transition to protease-supplemented micro diet (T4) from 17 dph onwards. Full article

The skin of amphibians is a rich source of peptides with a wide range of biological activities. They are stored in secretory granules in an inactive form. Upon stimulation, they are secreted together with proteases into the skin. Once activated, they rapidly exert their biological effects, including fighting microorganisms and predators, while their excess is immediately destroyed by the released proteases. To keep bioactive peptides in their initial form, it is necessary to inhibit these enzymes. Several inhibitors for this purpose have previously been mentioned; however, there has not been any reliable comparison of their efficiency so far. Here, we studied the efficiency of methanol and hydrochloric and formic acids, as well as phenylmethylsulfonyl fluoride, in the inhibition of nine frog peptides with the known sequence, belonging to five families in the secretion of Pelophylax esculentus. The results demonstrated that methanol had the highest inhibitory efficiency, while phenylmethylsulfonyl fluoride was the least efficient, probably due to its instability in aqueous media. Possible cleavages between certain amino acid residues in the sequence were established for each of the inhibitors. These results may be helpful for future studies on the nature of proteases and on prediction of the possible cleavage sites in novel peptides. Full article

Neutrophil elastase (HNE), like other members of the so-called GASPIDs (Granule-Associated Serine Peptidases of Immune Defense), is activated during protein biosynthesis in myeloid precursors and stored enzymatically active in cytoplasmic granules of resting neutrophils until secreted at sites of host defense and inflammation. Inhibitors thus could bind to the fully formed active site of the protease intracellularly in immature progenitors, in circulating neutrophils, or to HNE secreted into the extracellular space. Here, we have compared the ability of a panel of diverse inhibitors to inhibit HNE in the U937 progenitor cell line, in human blood-derived neutrophils, and in solution. Most synthetic inhibitors and, surprisingly, even a small naturally occurring proteinaceous inhibitor inhibit HNE intracellularly, but the extent and dynamics differ markedly from classical enzyme kinetics describing extracellular inhibition. Intracellular inhibition of HNE potentially affects neutrophil functions and has side effects, but it avoids competition of inhibitors with extracellular substrates that limit its efficacy. As both intra- and extracellular inhibition have advantages and disadvantages, the quantification of intracellular inhibition, in addition to classical enzyme kinetics, will aid the design of novel, clinically applicable HNE inhibitors with targeted sites of action. Full article