Extracellular Proteins Research Articles

Extracellular vesicles in the pathogenesis and future diagnostics of oral squamous cell carcinoma.

Extracellular vesicles are a group of heterogeneous particles secreted during both physiological and pathological conditions which serve in intercellular communication and play a role in the development and progression of oral squamous cell carcinoma, the most common malignant tumor of the head and neck with a high mortality rate. Extensive research is being conducted in order to determine the precise role of extracellular vesicles in oncogenic processes and to explore the possible application of extracellular vesicles as early tumor biomarkers. In this review, we aimed to systematize observed roles extracellular vesicles might play in organizing of tumor microenvironment, tumor invasion and metastasis, as well as the impact of extracellular vesicles on immune dysregulation and development of resistance to chemotherapeutics. Additionally, we summarized findings involving the potential use of extracellular vesicles cargo proteins as early disease biomarkers.

Enhanced penetration and biofilm eradication by sophorolipid micelles encapsulating Honokiol: a comprehensive solution for biofilm-associated lung infections

BackgroundBiofilm-associated lung infections, particularly those caused by Staphylococcus aureus (S. aureus), pose significant clinical challenges to conventional therapies. S. aureus Biofilm infections are refractory to treatment due to the presence of persister bacterial cells and the barrier effect of unique extracellular polymeric substances (EPS).ResultsThis study describes the development of multifunctional micelles, HK-SL Ms, utilizing sophorolipid (SL) to encapsulate Honokiol (HK). HK-SL Ms potently disrupted the EPS barrier, killed some internal colonizing bacteria, and inhibited further bacterial adhesion. Consequently, the dynamic cycling of biofilms was hindered, achieving a promising removal of S. aureus biofilms. In vitro studies demonstrated that HK-SL Ms exhibited significant antimicrobial reduction of a 6.42 log10CFU/mL. HK-SL Ms eradicated 71.73% of biofilms by targeting extracellular polysaccharides, extracellular proteins, and viable cells within the biofilm. Additionally, 1.66 log10CFU/mL units of S. aureus within biofilms were killed. Moreover, HK-SL Ms inhibited 91.10% of early S. aureus biofilm formation by obstructing initial bacterial adhesion and the formation of extracellular polysaccharides and polysaccharide intercellular adhesins (PIA). Thus, the reestablishment and reinfection of S. aureus biofilms could be resolved promisingly. Biofilm infections are as predominant in acute pneumonia as in chronic cases, inducing similar lung inflammation. In a murine model of pneumonia infected by S. aureus, HK-SL Ms significantly reduced the bacterial load in the lungs, decreased inflammatory factor levels, and repaired lung tissue damage.ConclusionsHK-SL Ms offers a novel strategy for the clinical treatment of biofilm-associated infections by dispersing and removing S. aureus biofilms and preventing new infections.Graphical

Direct development of microalgae-bacterial granular sludge system by seeding pre-made microalgae-dewatered sludge granules: Performance and mechanism analysis.

Microalgae-bacterial granular sludge (MBGS) process has great potential in achieving carbon neutrality and energy neutrality, but rapidly cultivating MBGS remains challenging. To address this challenge, this study proposes a new strategy to develop MBGS systems using pre-made granules from microalgae and dewatered sludge. The results indicate that using pre-made microalgae-dewatered sludge granules (M-DSG) as inoculants can directly develop MBGS system, with M-DSG maintaining a relatively stable granular structure, and ultimately achieving pollutant removal efficiencies of 94.0% for chemical oxygen demand (COD), 99.7% for ammonium nitrogen (NH4+-N), and 86.0% for total inorganic nitrogen (TIN). Extracellular polymeric substances (EPS) play a dominant role in maintaining the structure of granules, while filamentous bacteria/algae provide additional reinforcement. The adhesion of microalgae to granules possibly relies on polysaccharides in tightly bound extracellular polymeric substances (TB-EPS) and proteins in loosely bound extracellular polymeric substances (LB-EPS). Microbial community analysis reveals that the target algae (Chlorella) remain the primary algae, and heterotrophic nitrifying bacteria (HNB) and denitrifying bacteria are enriched.

The enigmatic heptalaminate surface membrane of intravascular schistosomes.

Intravascular schistosomes have an unusual outer, heptalaminate (seven-layered) covering consisting of not one but two lipid bilayers. Here, we present an updated model of the molecular composition of these bilayers in Schistosoma mansoni that places most identified proteins in the outer, and not the inner, membrane. Here, enzymes would have access to their recently described (non-membrane-permeable) substrates. By contrast, nutrient transporter proteins must be in both membranes to facilitate uptake into the worm's inner tissues. Ectoenzyme activities displayed by living worms suggest the presence on their outer surface of several noncanonically extracellular proteins. The advantages of having a double-bilayered covering may relate to impeding host immunological attack and/or to the worm's ability to acquire selected host molecules onto their exterior.

A Bifunctional Lysosome-Targeting Chimera Nanoplatform for Tumor-Selective Protein Degradation and Enhanced Cancer Immunotherapy.

Lysosome-targeting chimeras (LYTACs) have recently emerged as a promising therapeutic strategy for degrading extracellular and membrane-associated pathogenic proteins by hijacking lysosome-targeting receptors. However, the antitumor performance of LYTAC is limited by its insufficient tumor accumulation and nonspecific activation. Additionally, the synergistic effects of LYTACs and other therapeutic modalities are crucial. To address these issues, a bifunctional LYTAC nanoplatform (NLTC) is developed for tumor-selective protein degradation and enhanced cancer immunotherapy. By rationally controlling the surface composition, the NLTC can effectively transport extracellular or membrane proteins into lysosomes for degradation via cation-independent mannose 6-phosphate receptors. With removable surface modification, an NLTC is obtained that efficiently accumulated in tumor tissues and avoided on-target off-tumor toxicity. Moreover, the synthesis method of NLTC is generally applicable to various enzymes. Thus, catalase (CAT) is encapsulated with NLTC to synergistically degrade cancer cell surface programmed death ligand-1 (PD-L1), relieve the immunosuppressive tumor microenvironment for effective cancer immunotherapy, and significantly inhibit tumor growth, recurrence, and metastasis in B16F10-bearing mice. This work presents a bifunctional LYTAC nanoplatform that can not only perform tissue-selective protein degradation but also integrate other therapeutic modalities, providing insights into the design of advanced LYTAC technologies for clinical applications.

Copper(II) carboxylate complexes inhibit Staphylococcus aureus biofilm formation by targeting extracellular proteins.

Three copper(II) complexes of diphenyl acetic acid (DPAA) pyridine (py), 2,2΄-dipyridylamine (dpa), and 4,7-diphenyl, 1,10-phenanthrline (di-phen), [Cu₂(DPAA)₄(py)2] (Cu-1), [Cu(DPAA)₂(dpa)] (Cu-2), and [Cu₂(DPAA)₄(di-phen)₂] (Cu-3) were synthesized and characterized. Their antibacterial activities were evalvated. The minimum inhibitory concentrations (MIC) of these complexes against six tested microbial strains ranged from 1 to 128μg/mL, and that of vancomycin antibiotic ranged from 0.5 to 2μg/mL. The bactericidal effects of Cu-1, Cu-2 and Cu-3 and vancomycin against Staphylococcus aureus (S. aureus) were determined by colony count assay. Cu-1, Cu-2, and vancomycin showed relatively weaker antibiofilm formation activities; however, Cu-3 showed enhanced activity against S. aureus proliferation and biofilm formation as confirmed by microscopic analysis. In antibiofilm assays, Cu-1, Cu-2 and Cu-3 demonstrated high inhibition ability (23-75%), of mature biofilm formation at concentrations of 5 to 15μg/mL, and vancomycin at 15μg/mL inhibited only 47%. Cu-3 also effectively killed S. aureus within biofilms at doses up to 2×MIC μg/mL. Further analysis of extracellular proteins (ECPs) expression revealed, that Cu-3 had significant potential in suppressing ECPs production. Molecular docking (MD) studies with biofilm associated protein (Bap) and SARS-CoV-2 receptors showed high interactions by several bonding types, where Cu-2 found as potent antiviral agent. Collectively, these findings highlighted the copper complexes potential in antibacterial applications, with Cu-3 emerging as a potent candidate for S. aureus biofilm inhibition.

A new weapon: the application of tumor vaccines based on extracellular exosomal heat shock proteins in immunotherapy.

Despite the significant advancements in cancer research, innovative approaches are still needed to reduce tumor incidence, progression, and dissemination, as well as for prolonging patient survival. Currently, the development of cancer vaccines is gaining attention as a novel preventative and therapeutic strategy. Although the concept of cancer vaccination is not new, a limited number of vaccines have received approval for tumor therapy. Heat shock protein (HSP)-based vaccination represents a promising strategy that harnesses specific tumor antigens to activate immune responses. Exosomes (Exs) are highly heterogeneous bilayer vesicles capable of transporting various types of molecules through extracellular space. Compared with conventional anticancer drugs, exosomes exhibit low toxicity and good biocompatibility, and they can stimulate the immune system either directly or indirectly. Ex-based vaccines may elicit an antitumor immune response that generates memory cells capable of recognizing cancer antigens, thereby inhibiting disease progression. This paper reviews the potential applications of HSPs and exosomes in the prevention and treatment of solid tumors. Finally, we discuss the advantages of the extracellular exosomal heat shock protein (HSP-Ex) vaccine and future research directions aimed at optimizing heat shock protein-based cancer immunotherapy strategies.

Genomic and biochemical investigations in the biomineralizing potential of an isolated marine ureolytic Bacillus sp. N₉.

Microbially Induced Calcium Carbonate Precipitation (MICP) plays a significant role in coastal soil stabilization and erosion prevention. In the present study, the biomineralizing potential of a newly isolated Bacillus sp. N₉ was investigated through MICP. The isolated Bacillus sp. N₉ induced calcium carbonate (CaCO3) precipitation in broth, agar, and real beach rock was evaluated by X-ray Fluorescence, Scanning Electron Microscopy, and X-ray Diffraction. The visual appearance of calcites as white crystals on the rock and the sealing of cracks in the beach sand cubes confirmed the CaCO3 precipitation by the isolated strain. The pH changes, soluble calcium (Ca2+) concentration levels and urease activity during the growth of the isolated Bacillus sp. N₉ were studied. High urease activity was observed, resulting in an increased production of carbonate (CO32-) ions, which in turn promoted a higher rate of CaCO₃ precipitation. Further, the assembly of the 3.27Mb genome of the isolated Bacillus sp. N₉ was evaluated using Nanopore sequencing technology, and various extracellular proteins, including urease, were identified. The gene annotation through PROKKA and RAST predicted 6700 and 7317 protein-coding sequences. The pathway annotation analysis through gene ontology, KEGG, and COG inferred the presence of genes in proteolytic characteristics, carbohydrate metabolism, and amino acid derivatives. The present study provides valuable insights into the isolated native Bacillus sp. N₉, demonstrating its potential to produce high urease activity and calcite precipitation through combined genomics and in vitro techniques.

Biomarkers of RV Dysfunction in HFrEF Identified by Direct Tissue Proteomics: Extracellular Proteins Fibromodulin and Fibulin-5.

Right ventricular dysfunction (RVD) is common in patients with heart failure with reduced ejection fraction, and it is associated with poor prognosis. However, no biomarker reflecting RVD is available for routine clinical use. Proteomic analysis of myocardium from the left ventricle and right ventricle (RV) of patients with heart failure with reduced ejection fraction with (n=10) and without RVD (n=10) who underwent heart transplantation was performed. Concentrations of 2 ECM (extracellular matrix) proteins with the highest myocardial upregulation in RVD, FMOD (fibromodulin) and FBLN5 (fibulin-5), were assayed in the blood and tested in a separate cohort of patients with heart failure with reduced ejection fraction (n=232) to test for the association of the 2 proteins with RV function and long-term outcomes. Multivariable linear regression revealed that plasma concentrations of both FMOD and FBLN5 were significantly associated with RV function regardless of the RV function assessment method. No association of FMOD or FBLN5 with left ventricular dysfunction, cardiac index, body mass index, diabetes status, or kidney function was found. Plasma levels of FMOD and FBLN5 were significantly associated with patient outcomes (P=0.005; P=0.004). Area under the curve analysis showed that the addition of FBLN5 or FMOD to RV function assessment had a significantly higher area under the curve after 4 years of follow-up (0.653 and 0.631, respectively) compared with RV function alone (0.570; P<0.05 for both). Similarly, the combination of MAGGIC (Meta-Analysis Global Group in Chronic Heart Failure) score, FBLN5, and FMOD had a significantly larger area under the curve (0.669) than the combination of MAGGIC score+RVD grade (0.572; P=0.02). The Kaplan-Meier analysis demonstrated that patients with the elevation of both FMOD and FBLN5 (ie, FMOD >64 ng/mL and FMOD >27 ng/mL) had a worse prognosis than those with the elevation of either FBLN5 or FMOD (P=0.03) demonstrating the additive prognostic value of both proteins. Our study proposes that circulating levels of FMOD and FBLN5 may serve as new biomarkers of RVD in patients with heart failure with reduced ejection fraction.

Biofilm Formation, Modulation, and Transcriptomic Regulation Under Stress Conditions in Halomicronema sp.

In nature, bacteria often form heterogeneous communities enclosed in a complex matrix known as biofilms. This extracellular matrix, produced by the microorganisms themselves, serves as the first barrier between the cells and the environment. It is composed mainly of water, extracellular polymeric substances (EPS), lipids, proteins, and DNA. Cyanobacteria form biofilms and have unique characteristics such as oxygenic photosynthesis, nitrogen fixation, excellent adaptability to various abiotic stress conditions, and the ability to secrete a variety of metabolites and hormones. This work focused on the characterization of the cyanobacterium Halomicronema sp. strain isolated from a brackish environment. This study included microscopic imaging, determination of phenolic content and antioxidant capacity, identification of chemicals interfering with biofilm formation, and transcriptomic analysis by RNA sequencing and real-time PCR. Gene expression analysis was centered on genes related to the production of EPS and biofilm-related transcription factors. This study led to the identification of wza1 and wzt as EPS biomarkers and luxR-05665, along with genes belonging to the TetR/AcrR and LysR families, as potential biomarkers useful for studying and monitoring biofilm formation under different environmental conditions. Moreover, this work revealed that Halomicronema sp. can grow even in the presence of strong abiotic stresses, such as high salt, and has good antioxidant properties.

Tissue fibrosis in cardiorenal syndrome: crosstalk between heart and kidneys.

Cardiorenal syndrome (CRS) is represented as an intricate dysfunctional interplay between the heart and kidneys, marked by cardiorenal inflammation and fibrosis. Unlike other organs, the repair process in cardiorenal injury involves a regenerative phase characterized by proliferation and polyploidization, followed by a subsequent pathogenic phase of fibrosis. In CRS, acute or chronic cardiorenal injury leads to hyperactive inflammation and fibrotic remodeling, associated with injury-mediated immune cell (Macrophages, Monocytes, and T-cells) infiltration and myofibroblast activation. An inflammatory to fibrotic transition corresponds with macrophage transition (M1-M2) associated with increased TGF-β response. Chronic inflammation disrupts hemodynamic pathways, leading to imbalanced oxidative stress and the production of cytokines and growth factors that promote fibrotic stimulation, contributing to pathological cardiorenal remodeling. The inflammatory response paves the pre-fibrotic cardiorenal niche and drives subsequent fibrotic remodeling by activated myofibroblasts. A fibrotic cardiorenal response in CRS is characterized by increased and degradation-resistant deposition of extracellular proteins especially fibrillar Collagen -I, -III, -V, and non-fibrillar Collagen-IV by active myofibroblasts. Recent advances in basic research animal models of CRS have advanced the knowledge of cardiorenal fibrosis. However, a significant need for clinical applications, trials, and evaluation is still needed. Circulating biomarkers like procollagen peptides and TGF-β have clinically been associated with cardiorenal fibrosis diagnosis in CRS. Treatments targeting the fibrotic pathways have also shown efficacy in amelioration of cardiorenal fibrosis in preclinical models. Recent combination therapies targeting multiple fibrotic pathways have been shown to offer promising results. Understanding the heterogenic pathological progression and fibrogenesis could identify novel therapeutic approaches for clinical CRS diagnosis and treatment.

Characterizing the Contaminant-Adhesion of a Dibenzofuran Degrader Rhodococcus sp.

The adhesion between dibenzofuran (DF) and degrading bacteria is the first step of DF biodegradation and affects the efficient degradation of DF. However, their efficient adhesion mechanism at the molecular level remains unclear. Therefore, this study first examined the adhesive behaviors and molecular mechanisms of Rhodococcus sp. strain p52 upon exposure to DF. The results showed that the adhesion between strain p52 and DF is mediated by extracellular polymeric substances (EPSs). Compared with sodium acetate as a carbon source, the percentages of glucose and proteins related to electron transfer, toxin-antitoxin, and stress responses were elevated, which were analyzed by polysaccharide composition and proteomics, and the contents of extracellular polysaccharides and proteins were increased. Moreover, biofilm analysis suggested an increase in EPS content, and the change in components increased biofilm yield and promoted loose and porous aggregation between the bacteria; this aggregation caused an increase in the specific surface area in contact with DF. The surface characteristics analysis indicated that the production of EPS reduced the absolute value of the zeta potential and increased the hydrophobicity of strain p52, which was beneficial for the adhesion of strain p52 and DF. These findings help us to enhance the understanding of the adhesion mechanisms and bioremediation of polycyclic aromatic hydrocarbons by degrading bacteria.

Insights into the enhanced uranium reduction efficiency through extracellular polymeric substances from Desulfovibrio vulgaris UR1 induced by mediating materials.

Understanding the role of extracellular polymeric substances (EPS) in the microbial reduction of uranium accelerated by mediating materials is crucial for enhancing the bioremediation of uranium-contaminated wastewater. In this study, biochar- and magnetite-loaded Desulfovibrio vulgaris UR1 exhibited significantly higher uranium reduction efficiency, with increases of 1.52 and 1.44 times respectively within one day. After loading with mediating materials, the charge transfer resistance of EPS was reduced, facilitating the extracellular electron transfer process. The increase of redox components, such as aromatic compounds and flavins, in EPS explained the enhanced extracellular electron transfer capacity. Moreover, the higher α-helix content in extracellular proteins could promote electron hopping. Proteomics analysis showed that extracellular proteins involved in iron-sulfur cluster binding, oxidoreductase activity, and electron transfer were significantly up-regulated, which facilitated the rapid microbial reduction of uranium. These findings provide valuable insights into the in-depth development of bioremediation technology for uranium-contaminated wastewater.

Selection of biofilm-inhibiting ssDNA aptamers against antibiotic-resistant Edwardsiella tarda by inhibition-SELEX and interaction with their binding proteins.

Biofilms can increase bacterial resistance to antibiotic therapies. Edwardsiella tarda with biofilm is highly resistant to antibacterial treatment, especially for the antibiotic-resistant strain. In this study, we obtained biofilm-inhibiting aptamers against antibiotic-resistant E. tarda via a novel systematic evolution of ligands by exponential enrichment (SELEX) technique, called inhibition-SELEX. After four rounds of screening and validation, we identified aptamers IB1, IB2, and IB3, which demonstrated biofilm-inhibition and biofilm-degradation rates of 69%, 75%, and 62% and 51%, 63%, and 45% at 2μmol/L, respectively, against antibiotic-resistant E. tarda. Magnetic separation, SDS-PAGE, and mass spectrometry analyses revealed that all three aptamers could bind to glyceraldehyde-3-phosphate dehydrogenase (GAPDH), while IB2 could also bind to formate C-acetyltransferase (FA). Through molecular docking and molecular dynamics simulations, it was found that the four complexes primarily interact through hydrogen bonding. Among them, IB1-GAPDH exhibited the strongest stability, followed by IB2-FA, then IB2-GAPDH, and IB3-GAPDH was the least stable. Our results suggest that IB1, IB2, and IB3 may inhibit and degrade E. tarda biofilm by interfering with the synthesis, secretion, and transportation of its extracellular polysaccharides and proteins by interacting with GAPDH and FA.

Hepatocyte targeting via the asialoglycoprotein receptor.

This review highlights the potential of asialoglycoprotein receptor (ASGPR)-mediated targeting in advancing liver-specific treatments and underscores the ongoing progress in the field. First, we provide a comprehensive examination of the nature of ASGPR ligands, both natural and synthetic. Next, we explore various drug delivery strategies leveraging ASGPR, with a particular emphasis on the delivery of therapeutic nucleic acids such as small interfering RNAs (siRNAs) and antisense oligonucleotides (ASOs). An in-depth analysis of the current status of RNA interference (RNAi) and ASO-based therapeutics is included, detailing approved therapies and those in various stages of clinical development (phases 1 to 3). Afterwards, we give an overview of other ASGPR-targeted conjugates, such as those with peptide nucleic acids or aptamers. Finally, targeted protein degradation of extracellular proteins through ASGPR is briefly discussed.

Identification of crucial extracellular genes as potential biomarkers in newly diagnosed Type 1 diabetes via integrated bioinformatics analysis.

In this study, we aimed to study the role of extracellular proteins as biomarkers associated with newly diagnosed Type 1 diabetes (NT1D) diagnosis and prognosis. We retrieved and analyzed the GSE55098 microarray dataset from the Gene Expression Omnibus (GEO) database. Using R software, we screened out the extracellular protein-differentially expressed genes (EP-DEGs) through several protein-related databases. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were applied to describe the role and function of these EP-DEGs. We used the STRING database to construct the interaction of proteins, Cytoscape software to visualize the protein-protein interaction (PPI) networks, and its plugin CytoHubba to identify the crucial genes between PPI networks. Finally, we used the comparative toxicogenomics database (CTD) to evaluate the connection between NT1D with the potential crucial genes and we validated our conclusions with another dataset (GSE33440) and some clinical samples. We identified 422 DEGs and 122 EP-DEGs from a dataset that includes (12) NT1D patients compared with (10) healthy people. Protein digestion and absorption, toll-like receptor signaling, and T cell receptor signaling were the most meaningful pathways defined by KEGG enrichment analyses. We recognized nine important extracellular genes: GZMB, CCL4, TNF, MMP9, CCL5, IFNG, CXCL1, GNLY, and LCN2. CTD analyses showed that LCN2, IFNG, and TNF had higher levels in NT1D and hypoglycemia; while TNF, IFNG and MMP9 increased in hyperglycemia. Further verification showed that LCN2, MMP9, TNF and IFNG were elevated in NT1D patients. The nine identified key extracellular genes, particularly LCN2, IFNG, TNF, and MMP9, may be potential diagnostic biomarkers for NT1D. Our findings provide new insights into the molecular mechanisms and novel therapeutic targets of NT1D.

Circulating large extracellular vesicles as diagnostic biomarkers of indeterminate thyroid nodules: multi-platform omics analysis.

While most thyroid nodules are benign, 7-15% are malignant. Patients with indeterminate thyroid nodules (specifically Bethesda IV/Thy3f) often undergo diagnostic hemithyroidectomy to reach a diagnosis on final histology. The aim of this study was to assess the feasibility of circulating large extracellular vesicles as diagnostic biomarkers in patients presenting with Thy3f thyroid nodules. This was a two-gate diagnostic accuracy study; patients with Thy3f thyroid nodules were age, sex and body mass index matched to healthy individuals. Final histology confirmed benign and malignant diagnoses. Plasma large extracellular vesicle counts were quantified using flow cytometry. Large extracellular vesicle microRNA and protein profiles were identified using next generation sequencing and mass spectrometry, respectively. A total of 42 patients with Thy3f nodules (22 with cancer, 20 with non-cancer diagnosis) and 16 healthy controls were included. Total large extracellular vesicle concentrations and the concentrations of extracellular vesicles expressing epithelial cell adhesion molecule and the cancer markers atypical chemokine receptor type 7, extracellular matrix metalloproteinase inducer and syndecan-4 were significantly higher in patients with Thy3f nodules (cancer and non-cancer) compared with healthy individuals. In patients with cancerous versus non-cancer Thy3f nodules, one microRNA was upregulated: mir-195-3p (P < 0.001). Five were downregulated: mir-3176 (P < 0.001), mir-205-5p (P < 0.001), novel-hsa-mir-208-3p (P < 0.001), mir-3529-3p (P = 0.01) and let-7i-3p (P = 0.02). Furthermore, three large extracellular vesicle proteins (kallikrein-related peptidase11 (KLK11) (P = 0.001), alpha-1-acid glycoprotein 2 (A1AG2) (P <0.001) and small integral membrane protein 1 (SMIM1) (P = 0.04)) were significantly upregulated, while 20 large extracellular vesicle proteins were significantly downregulated (most downregulated: chemokine (C-X-C motif) ligand 7 (CXCL7), tubulin beta chain 1 (TBB1), binding immunoglobulin protein (BIP) and actinin alpha 1 (ACTN1) (P < 0.001)) in cancerous compared with non-cancer Thy3f nodules. Circulating large extracellular vesicle miRNA and protein profiles have a high diagnostic value to discriminate between benign and malignant nodules for patients with Thy3f cytology. Further validation for clinical performance will be needed.

Development of Aspergillus oryzae BCC7051 as a Robust Cell Factory Towards the Transcriptional Regulation of Protease-Encoding Genes for Industrial Applications.

Enzyme-mediated protein degradation is a major concern in industrial fungal strain improvement, making low-proteolytic strains preferable for enhanced protein production. Here, we improved food-grade Aspergillus oryzae BCC7051 by manipulating the transcriptional regulation of protease-encoding genes. Genome mining of the transcription factor AoprtR and computational analysis confirmed its deduced amino acid sequence sharing evolutionary conservation across Aspergillus and Penicillium spp. The AoPrtR protein, which is classified into the Zn(II)2-Cys6-type transcription factor family, manipulates both intra- and extracellular proteolytic enzymes. Our transcriptional analysis indicated that the regulation of several protease-encoding genes was AoPrtR-dependent, with AoPrtR acting as a potent activator for extracellular acid-protease-encoding genes and a likely repressor for intracellular non-acid-protease-encoding genes. An indirect regulatory mechanism independent of PrtR may enhance proteolysis. Moreover, AoPrtR disruption increased extracellular esterase production by 2.55-fold, emphasizing its role in protein secretion. Our findings highlight the complexity of AoPrtR-mediated regulation by A. oryzae. Manipulation of regulatory processes through AoPrtR prevents secreted protein degradation and enhances the quantity of extracellular proteins, suggesting the low-proteolytic variant as a promising platform for the production of these proteins. This modified strain has biotechnological potential for further refinement and sustainable production of bio-based products in the food, feed, and nutraceutical industries.

Novel Approaches in Targeting Cell Surface and Secreted Proteins for Lysosomal Degradation.

Protein degradation is pivotal for all biochemical aspects of cellular function. In mammalian cells, protein degradation is mediated mainly by the ubiquitin proteasome system (UPS) and the autophagic-lysosomal system (ALS). Over the last two decades, different types of targeted protein degradation approaches have been developed including proteolysis targeting chimeras (PROTACs) and lysosome targeting chimeras (LYTACs), which employ the UPS to degrade intracellular proteins and the ALS to degrade extracellular and membrane proteins respectively. Nevertheless, Current targeted membrane protein degradation approaches face some inherent challenges including limited target protein degradation efficacy and cell type specific applicability. Herein, we highlight some recent developments of novel targeted membrane protein degradation modalities that exhibit wide-applicability and high protein degradation efficiency. These novel membrane protein degraders hold tremendous promise as new pharmacological and biochemical tools in targeting membrane and secretory proteins for lysosomal degradation.

N-Acetylcysteine in Agriculture: New Insights into Its Effects for Sustainable Crop Management

N-Acetylcysteine (NAC), an amino acid naturally found in Allium plants, is well-known for its health benefits in humans. However, its potential applications in plant health remain underexplored, especially considering that while thousands of studies are published annually in clinical research, fewer than 10 focus on its use in plants each year. This review consolidates recent findings on the role of NAC as a potent antioxidant molecule in mitigating both biotic and abiotic stresses in plants, highlighting its valuable contributions to sustainable agriculture. NAC reduces oxidative stress by directly scavenging reactive oxygen species (ROS) and indirectly increasing cysteine and glutathione levels, both critical for cellular defense responses. Moreover, NAC breaks intramolecular and intermolecular disulfide bonds in thiolated proteins, potentially further enhancing its antioxidant effects by releasing free thiols. It also functions as a chelating agent, binding to heavy metals and reducing their toxicity. Furthermore, NAC’s efficacy in controlling bacterial infections in humans and animals—through mechanisms such as disrupting disulfide bonds in extracellular proteins and breaking down biofilm matrices—has been shown to extend to plant pathosystems, where it aids in suppressing bacterial pathogens. In addition to stress alleviation, NAC has been demonstrated to enhance crop productivity, improve food quality, and offer ecological benefits by promoting the health of pollinators and free-living soil organisms. These multifaceted attributes highlight NAC’s potential as a key tool in agriculture, helping to improve plant resilience, support ecosystem balance, and advance sustainable crop management practices.