Search Results (921)

Considering the challenges associated with implementing emerging technologies and bacterial-derived antimicrobial metabolites at an industrial scale in the meat industry, this comprehensive review investigates the interactions between lactic acid bacteria-producing antimicrobial metabolites and emerging food preservation technologies applied to meat systems. By integrating evidence from microbiology, food engineering, and molecular physiology, the review characterizes how metabolites-derived compounds exert inhibitory activity through pH modulation, membrane permeabilization, disruption of proton motive force, and interference with cell wall biosynthesis. These biochemical actions are evaluated in parallel with the mechanistic effects of high-pressure processing, pulsed electric fields, cold plasma, irradiation, pulsed light, ultrasound, ohmic heating and nanotechnology. Across the literature, consistent patterns of synergy emerge: many emerging technologies induce structural and metabolic vulnerabilities in microbial cells, thereby amplifying the efficacy of antimicrobial metabolites while enabling reductions in process intensity. The review consolidates these findings to elucidate multi-hurdle strategies capable of improving microbial safety, extending shelf life, and preserving the physicochemical integrity of meat products. Remaining challenges include optimizing combinational parameters, ensuring metabolite stability within complex matrices, and aligning integrated preservation strategies with regulatory and industrial constraints. Full article

The rising global incidence of Candida parapsilosis infections is increasingly complicated by antifungal resistance, resulting in frequent therapeutic failure. This study investigated the potential of the natural compound catechin to enhance the efficacy of fluconazole through synergistic interaction. We evaluated the susceptibility of C. parapsilosis clinical isolates and a reference strain to combinations of catechin and fluconazole using standardized microbiological assays and molecular techniques. In vivo efficacy was assessed using the Galleria mellonella infection model. Mechanistic studies included the measurement of intracellular reactive oxygen species (ROS) production and plasma membrane permeability. Catechin alone caused growth retardation in all strains. However, the combination of catechin and fluconazole resulted in complete growth inhibition of the reference strain and significant growth reduction in azole-resistant clinical isolates. While the combination slightly increased intracellular ROS production, no significant changes in plasma membrane permeability or membrane potential were observed. Notably, catechin induced the expression of the resistance-associated genes CpTAC1 and CpCDR1B in resistant isolates. In vivo experiments demonstrated that catechin significantly reduced mortality in G. mellonella larvae infected with C. parapsilosis. These findings suggest that catechin is a promising candidate for developing synergistic antifungal therapies against resistant Candida species. Full article

Longan (Dimocarpus longan Lour.) is a commercially valuable tropical fruit crop that contains two antagonistic FLOWERING LOCUS T (FT) homologs involved in regulating flowering time. However, how these FT genes interact with flowering regulators FLOWERING LOCUS D (FD) and APETALA1 (AP1) remains unknown. Four flowering-related genes in longan, DlFT1, DlFT2, DlAP1 and DlFD, were successfully isolated. Expression profiling revealed that all four genes were expressed in leaves and buds across different stages of natural and KClO₃-induced floral bud differentiation. Functional characterization through heterologous overexpression in Arabidopsis thaliana showed that DlAP1 significantly promotes early flowering under long-day conditions and induced morphological changes in floral organs and leaves. In contrast, DlFD overexpression had no effect on flowering time. Subcellular localization assays revealed that DlFT1 and DlFT2 localized to both the nucleus and the plasma membrane, while DlAP1 and DlFD localized exclusively to the nucleus. Yeast two-hybrid and bimolecular fluorescence complementation (BiFC) analyses revealed a novel regulatory node: DlFT1 directly interacts with DlAP1, a finding that expands the classical FT-FD-AP1 flowering model. Additionally, DlFD interacts more strongly with DlFT1 than with DlFT2, whereas DlFT1 only interacts with DlAP1, but not DlFT2. These results demonstrate that DlFT1 promotes flowering not only via the conserved FD-dependent pathway but also through direct association with AP1. These findings advance our understanding of the regulatory mechanisms of flowering in longan and provide valuable insights into flowering pathways of perennial woody species. Full article

Pollen tube growth entails complex molecular interactions between the cytoskeletal apparatus and membrane trafficking. Tip growth involves polarized distribution of proteins and lipids along the plasma membrane, including liquid-ordered microdomains, rich in sterols and sphingolipids (lipid rafts), in the apical/subapical region of tobacco pollen tubes. Intriguingly, biochemical characterization of detergent-insoluble membranes purified from tobacco pollen tubes revealed the presence of both actin and tubulin. Here, we report that inhibition of sterol biosynthesis altered lipid rafts and lowered the association of tubulin with detergent-insoluble membranes. Our results showed that sterol depletion increased the number of microtubules in the subapical region, altered microtubule distribution and affected microtubule bundling activity. Oryzalin washout experiments also suggested that lipid-ordered domains could play a role in regulating microtubule nucleation/regrowth. Full article

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is caused by mutations in PKD1 or PKD2 genes, encoding polycystin-1 (PC1) or polycystin-2 (PC2), respectively, characterized by excessive cell proliferation and fluid secretion, resulting in renal cyst formation and growth. PC1 and PC2 form a complex localized on the plasma membrane, endoplasmic reticulum, and primary cilia. PC2 is a non-selective cation channel which, in renal epithelial cells, contributes to calcium transport and signaling. It has been previously shown in renal cells that high external calcium increases whole-cell currents likely mediated by PC2. In this study, we explored the possibility that the Calcium Sensing Receptor (CaSR) is involved in the functional regulation of PC2. To test this hypothesis, human conditionally immortalized Proximal Tubular Epithelial cells, isolated from urine sediments, wt or with stably downregulated PKD1 (PC1KD) or PKD2 (PC2KD) were used. Interestingly, CaSR and PC2 co-immunoprecipitated and Proximity Ligation Assay demonstrated a direct physical interaction at endogenous protein levels. Membrane potential measurements demonstrated that selective CaSR activation, elicited by the calcimimetic R568, caused plasma membrane depolarization, consistent with the modulation of PC2-mediated cation currents, which was significantly lower in PC2KD with respect to wt and PC1KD cells. To conclude, this study provides evidence for a functional coupling of CaSR and PC2, which might be relevant for therapeutic strategies to correct dysregulations occurring in ADPKD. Full article

Plasma membrane Ca²⁺-ATPases (PMCA) are activated by calmodulin (CaM) via a C-terminal calmodulin-binding domain, CaMBD. Although specific mutations in this domain have been linked to disease, the broader impact of alternative substitutions across the interface remains unexplored. We applied an integrative in silico workflow to test six substitutions within CaMBD positions 1–18, L5R, N6I, I8T, V14E/D, and F18S, across PMCA isoforms 1–4. CaMBD sequences were aligned across isoforms, and candidates for substitutions were selected by conservation and nucleotide feasibility, prioritizing conserved or co-evolutionarily relevant sites, with substitutions possible by single-nucleotide change. PolyPhen-2 screened the impact of the substitutions on the protein functionality, the DisGeNET database was used to contextualize ATP2B genes with clinical phenotypes, and structural models plus binding free energy changes were estimated with AlphaFold3, FoldX, and MutaBind2. Effects were isoform and subregion dependent, with the strongest weakening toward the CaMBD C-terminus. V14E/D and F18S showed the largest and consistent predicted destabilization, consistent with disruption of conserved hydrophobic anchors. I8T and L5R had mixed outcomes depending on isoform, while N6I presented various scenarios with no clear effect. PolyPhen-2 classified most tested substitutions as damaging. Gene-disease evidence linked ATP2B to neurological, endocrine, and oncologic phenotypes, consistent with roles in Ca²⁺ homeostasis. Overall, CaMBD appears highly sensitive to perturbation, with distal positions 14–18 particularly vulnerable to substitutions that can destabilize CaM binding and potentially impair PMCA-mediated Ca²⁺ clearance in susceptible tissues. Full article

Melanocyte signaling through the MAPK pathway is orchestrated by NRas and relayed downstream via multiple effectors, such as RAF, Ral, and PI3K. In spite of their significance, the molecular mechanisms of signaling relay by NRas, their dynamics, and their potential as therapeutic targets remain unclear. Using multi-color single molecule localization microscopy (PALM and dSTORM), we resolved the mutual nanoscale organization of NRas, PI3K, and BRAF at the plasma membrane of fixed and live melanoma cells. Surprisingly, NRas and its oncogenic mutation Q61R colocalized with PI3K in mutual nanoclusters, where BRAF was also frequently present. In live cells, NRas and PI3K co-clustering declined, yet persisted over minutes. Clinically relevant perturbations revealed unexpected crosstalk within these nanoclusters and consequently, between the MAPK and PI3K pathways. Specifically, overexpression of the Ras binding domain (RBD) and PI3K inhibition by wortmannin disrupted NRAS-PI3K interactions, and reduced both pAKT and pERK levels and cancer cell proliferation. MEK inhibition with trametinib resulted in similar, yet more pronounced effects. Thus, our findings provide novel insights into NRAS-mediated signaling through nanoscale clusters and underscore their potential as therapeutic targets. Full article

Oral diseases pose a major public health challenge, especially in low-income countries where dental care is limited due to high costs. In this context, phytotherapy has gained attention as a complementary approach due to its bacteriostatic, anti-inflammatory, healing, and analgesic properties. These therapeutic effects are mainly attributed to plant-derived bioactive metabolites, which interact with cellular structures, especially the plasma membrane, to modulate inflammation, stimulate tissue regeneration, and support antimicrobial defense. This review systematically examined the scientific literature to identify Latin American medicinal plants with therapeutic potential in dentistry. Based on their clinical and ethnobotanical applications, the analysis focused on species with anti-inflammatory, healing, analgesic, and relaxing effects, particularly in conditions such as dental pain, gingivitis, and periodontitis. Given the close relationship between pain, inflammation, and periodontal disease, these conditions cannot be studied in isolation. Gingivitis and periodontitis often present with painful symptoms and inflammatory responses that overlap with mechanisms of tissue damage and repair. Therefore, broadening the scope of this review allows for a more comprehensive understanding of how Latin American medicinal plants can contribute not only to pain relief but also to periodontal health, inflammation control, and wound healing. Fifty plant species were identified. Among these, 35 exhibited anti-inflammatory activity, 28 had healing properties, 20 showed analgesic effects, and 12 were associated with relaxing properties. Mexico accounted for the highest proportion of species (60%), followed by Colombia and Peru (54%) and then Brazil (32%). These percentages represent the proportion of plant species reported in studies originating from each country, relative to the total number of species identified in the review. The most studied species were Salvia rosmarinus Spenn. (Lamiaceae), Moringa oleifera Lam. (Moringaceae), Aloe vera (L.) Burm.f. (Asphodelaceae), and Ocimum basilicum L. (Lamiaceae). Latin American medicinal plants demonstrate strong potential not only in dental therapy but also in the management of periodontal inflammation and oral diseases. However, further research and clinical validation are needed to ensure their safe integration into conventional treatments. Full article

Background/Objectives: Methicillin-resistant Staphylococcus aureus (MRSA) poses a significant global health threat due to its increasing resistance to conventional antibiotics. Antimicrobial peptides (AMPs) derived from natural sources represent a promising alternative. Fragments of spider membrane-active toxins can serve as AMPs with anti-MRSA activity. Methods: To demonstrate this, amino acid sequences of approximately 2000 linear spider venom peptides were fragmented into 9–22-residue-long moieties (75,235 in total) and pre-trained neural networks were used to predict their anti-MRSA activity. As many as 15 peptides with high predicted activity were synthesized, and three AMPs with high anti-MRSA and low hemolytic activities were selected. One of these peptides was studied using high-resolution ¹H-, ¹³C-, and ¹⁵N-NMR spectroscopy in an aqueous solution and lyso-palmitoylphosphatidylglycerol (LPPG) micelles. Wide-line ³¹P-NMR was applied to multilamellar phospholipid liposomes composed of phosphatidylcholine (PC) or phosphatidylglycerol (PG). Results: Low hemolytic activity is explained by non-specific interaction with PC whereas high antibacterial activity arises from specific interaction with PG accompanied with the formation of a tight complex between the N-terminal tripeptide fragment and PG headgroup. The structure of a such complex, stabilized by an ionic interaction between the N-terminal NH₃⁺ group and the lipid phosphate, was determined based on peptide–LPPG NOEs. The most favorable ratio between anti-MRSA and hemolytic activities, i.e., selectivity of the peptides, is attained when the tripeptide consists exclusively of phenylalanine and tryptophan residues. Confocal microscopy confirmed that the most selective peptide deteriorates the plasma membrane of S. aureus. Conclusions: This approach may enable the production of highly selective AMPs against Stapylococci, including MRSA. Full article

Interrelations between the plasma membrane and cytoskeleton are of crucial importance for essential cellular processes such as endocytosis, formation of intercellular junctions, cell morphology, etc. Many studies validate the beneficial effects of polyphenols as antioxidant and protective agents, but a molecular mechanism of their interaction and transition through the plasma membranes of different cell lines is still missing. In this study, we examined the affinity of fractions enriched in flavonoid glycosides (FGs) and caffeoylquinic acids (CQAs), obtained from the methanol extract of the medicinal plant Inula oculus-christi L., to reorganize the plasma membrane structure and actin cytoskeleton by using confocal microscopy. Assessment of the degree of membrane ordering aiming to distinguish the ordered from disordered regions of the cellular membranes was performed using the fluorescent dye Di-4-ANEPPDHQ, and visualization of F-actin was by TRITC-phalloidin. Two epithelial cell lines with clear differences in their origin and plasma membrane organization were chosen: the non-malignant MDCK II and the cancerous A549. Our results showed that flavonoid glycosides exhibited an ordering effect on plasma membranes of cancerous cells and fluidized one on non-malignant cells. Different patterns of actin reorganization were observed for both cell lines after treatment. Our results indicate the potential of plant-derived polyphenols as modulators of the membrane’s structural organization, offering valuable insights for the development of membrane-targeted therapeutic strategies. Full article

Milk coffee is a composite beverage in which interactions among dairy proteins, lipids, and coffee polyphenols govern flavor, texture, and stability. This review synthesizes recent research to guide formulation and processing, covering conventional Ultra-high temperature sterilization (UHT) and innovative routes including blending-after-sterilization (BAS), high-pressure homogenization (HPH), ultrasound/pulsed electric field (PEF)/cold plasma (CP), microencapsulation, and plant-based matrices. Key findings show that non-covalent protein–polyphenol complexes and interfacial partitioning at fat-globule membranes control volatile retention, astringency, droplet structure, and phenolic bioaccessibility; appropriate fat levels and HPH refine microstructure; BAS better preserves aroma; and matrix or decaffeination choices modulate antioxidant capacity. Guided by these insights, we propose a concise “process–activity–stability” framework linking parameters to functionality and shelf life to accelerate the development of high-quality, nutritious, enjoyable, and more sustainable milk coffee products. Full article

The present study demonstrates encapsulation of Coriandrum sativum essential oil in chitosan nanoemulsion and its effectiveness against fungal infestation and fumonisin B₁ (FB₁)- and B₂ (FB₂)-mediated biodeterioration of stored rice samples. Mycoflora analysis of different rice varieties revealed fungal occurrence and Fusarium proliferatum-BRC-R2 as the most toxigenic strain with highest FB₁- and FB₂-producing potentiality. GC-MS analysis of Coriandrum sativum essential oil (CEO) revealed linalool as the major component. The CEO-loaded chitosan nanoemulsion (Ne-CEO) was characterized by Scanning electron microscopy, X-ray diffractometry, Dynamic light scattering, and Fourier transform infrared spectroscopy. The Ne-CEO showed better antifungal and anti-fumonisin effectiveness as compared to unencapsulated CEO. The antifungal mechanism was associated with reduced ergosterol content, efflux of ions, proteins, nucleic acids, and destruction of plasma membrane integrity. The in silico interaction of linalool with Fum 1 protein confirmed the molecular action of anti-fumonisin activity. Additionally, the Ne-CEO displayed improved antioxidant activity and promising antifungal and anti-fumonisin activity during in situ investigation in rice samples (Gobindobhog variety) along with inhibition of the deterioration of carbohydrate, protein content, and lipid peroxidation without altering organoleptic properties and seed germination potentiality. Overall, the investigation strengthens the potentiality of Ne-CEO as a novel preservative of stored food commodities. Full article

Serotonin (5-HT) is a vital intercellular messenger with diverse signaling functions throughout the human body. We have characterized and implemented a novel, in vitro fluorescence-based method of measuring 5-HT binding to gain a fuller understanding of the interactions between 5-HT and its receptors. This method involves expression of 5-HT receptor proteins in cultured cells with the fluorescent, non-canonical amino acid l-3-(6-acetylnaphthalen-2-ylamino)-2-aminopropanoic acid (ANAP) incorporated into the ligand binding site. ANAP fluorescence was quenched in solution by both 5-HT and dopamine. Time-resolved photoluminescence and transient absorption spectroscopy confirmed that ANAP quenching by 5-HT occurs via a charge-transfer process that recovers through back-electron transfer on the nanosecond timescale. Supported by density functional theory calculations, this process likely involved an ANAP reduction by 5-HT. To test this method on intact receptors in a cellular context, we expressed 5-HT_3A receptors (5-HT-gated ion channels) in HEK293T cells with ANAP inserted co-translationally into the transmitter binding site. Fluorescently labeled 5-HT_3A receptors were functional and activated by 5-HT, as assessed by whole-cell patch clamp. Addition of 5-HT caused a concentration-dependent quenching of fluorescence from ANAP-tagged channels in intact cells and unroofed plasma membranes, demonstrating the utility of this method for measuring 5-HT binding to its receptors. Collectively, these results delineate a technique for measuring transmitter binding that can be widely adopted to explore 5-HT binding not only to 5-HT₃ receptors, but to any 5-HT receptor, transporter, or binding protein in heterologous expression systems. Full article

Charcot–Marie–Tooth disease type 1A (CMT1A) is a hereditary condition caused by the duplication of the PMP22 gene. Overexpression of peripheral myelin protein 22 in Schwann cells leads to myelin sheath defects and axonal loss. We have produced a cell model to facilitate studies of the molecular mechanisms involved in PMP22 accumulation and clearance. Our model is a stably transfected, clonal, immortalised human Schwann cell line with overexpressed levels of PMP22 fusion protein. A control-transfected cell line (vector lacking PMP22) was also produced. PMP22-transfected cells had reduced levels of mitosis, with the PMP22 fusion protein concentrated in punctate aggregates in the cytoplasm and expressed at the plasma membranes, which were often irregular and spindly. In contrast, control cells (control-transfected and parent cell lines) generally had smooth and regular plasma membrane morphology. Culturing in the presence of NRG1 and forskolin lead to upregulation of markers of myelination potential in the control cells. These markers were more variable in the cells stably transfected with PMP22, including decreased levels of transcripts of SOX10, JUN, S100B and NGFR, but increased levels of MPZ and EGR2 compared to controls. Using proximity-dependent biotin identification (BioID2), several hundred proteins were identified in the proximity of the overexpressed PMP22, of which 291 significant proteins were only detected in the proximity of PMP22 and not in that of control pull-downs. Among the most significantly enriched PMP22-interacting proteins were integrins alpha-2 (ITGA2) and alpha-7 (ITGA7), which play a role in myelination via their interactions with the extracellular matrix. The presence of ITGA2 in just the PMP22-transfected fraction was confirmed by western blot. Some of the proteins were associated with several enriched molecular pathways, including molecular transport and protein trafficking, and may represent potential therapeutic targets for CMT1A by promoting the degradation and enhanced trafficking of PMP22. Full article

Mancozeb is a broad-spectrum fungicide used extensively in agriculture to protect crops against a wide range of plant diseases. Although its capacity to induce oxidative stress is well documented, the cytotoxic effects of mancozeb on red blood cells (RBCs) remain poorly characterized. The present study aimed to investigate the cytotoxic effects of mancozeb on isolated RBCs, with particular focus on oxidative stress-induced cellular and molecular alterations. Human RBCs were exposed to mancozeb (0.5–100 µM) for 24 h. No hemolytic activity was observed across the tested concentrations. However, 10 and 100 µM mancozeb induced a significant increase in intracellular reactive oxygen species (ROS), leading to lipid and protein oxidation and impaired Na⁺/K⁺-ATPase and anion exchanger 1 (AE1) function. These changes resulted in altered RBC morphology, reduced deformability, and increased methemoglobin levels. Alterations in glycophorin A distribution, anion exchanger 1 (AE1) clustering and phosphorylation, and α/β-spectrin and band 4.1 re-arrangement indicated disrupted membrane–cytoskeleton interactions. A release of extracellular vesicles (EVs) positive for glycophorin A and annexin-V was also observed, consistent with plasma membrane remodeling. Despite increased intracellular calcium, eryptosis remained minimal, possibly due to activation of protective estrogen receptor (ER)-mediated pathways involving ERK1/2 and AKT signaling. Activation of the cellular antioxidant system and the glutathione redox system (GSH/GSSG) occurred, with catalase (CAT) playing a predominant role, while superoxide dismutase (SOD) activity remained largely unchanged. These findings offer mechanistic insights regarding the potential health impact of oxidative stress induced by pesticide exposure. Full article