Membrane Proteins Research Articles

NQO1 Triggers Neutrophil Recruitment and NET formation to Drive Lung Metastasis of Invasive Breast Cancer.

Metastasis to the lungs is a leading cause of death for breast cancer patients. Therefore, effective therapies are urgently needed to prevent and treat breast cancer lung metastasis In this study, we uncovered a mechanism by which NAD(P)H:quinone oxidoreductase 1 (NQO1) orchestrates lung metastasis. NQO1 stabilized and upregulated peptidyl-prolyl cis-trans isomerase A (PPIA), a chaperone that regulates protein conformation and activity, by preventing its oxidation at a critical cysteine residue C161. PPIA subsequently activated CD147, a membrane protein that facilitates cell invasion. Moreover, NQO1-induced secretion of PPIA modulated the immune landscape of both primary and lung metastatic sites. Secreted PPIA engaged CD147 on neutrophils and triggered the release of neutrophil extracellular traps (NET) and neutrophil elastase, which enhanced tumor progression, invasiveness and lung colonization. Pharmacological targeting of PPIA effectively inhibited NQO1-mediated breast cancer lung metastasis. These findings reveal a previously unrecognized NQO1-PPIA-CD147-NET axis that drives breast cancer lung metastasis. Inhibiting this axis is a potential therapeutic strategy to limit lung metastasis in breast cancer patients.

Label-free evanescent imaging of cellular heterogeneity in membrane protein binding kinetics

Quantifying cellular heterogeneity of membrane protein binding kinetics is challenging but important for exploring drug resistance and screening drugs. Label-free analysis methods have emerged as promising tools for in situ binding kinetics analysis, but they have not been used for high throughput single cell analysis in live cells. Here we show that this is possible with Evanescent Scattering Microscopy (ESM). The ESM permits analyzing the kinetics of ligand binding onto membrane proteins in individual fixed and live cells, and provides a throughput of ∼200 cells in a single measurement with a period of ∼7 minutes. The statistical analysis further shows that the dissociation rate constant dominates the heterogeneity of cell responses to ligand binding, providing evidence for a long-standing hypothesis that the drug-target residence time may play a critical role in drug treatment. In addition, the ESM reveals that under some conditions the cells have responses to drug binding at the single cell level, whereas the ensemble measurements may average out the individual differences and present false negative results. We anticipate that the new evanescent imaging method will provide a powerful tool to quantify the functions of cellular proteins, especially their cell-to-cell heterogeneity that can provide fuel for drug resistance.

Proteomic analysis reveals the dominant effect of ipomoeassin F on the synthesis of membrane and secretory proteins in triple-negative breast cancer cells.

Ipomoeassin F (Ipom-F) is a natural compound with embedded carbohydrates that exhibits a potent cytotoxic effect on triple-negative breast cancer (TNBC) cells. The mechanism behind this selective potency remains unclear. To elucidate this mechanism, we analyzed the proteome profiles of the TNBC MDA-MB-231 cells after exposure to Ipom-F at different time points and increasing doses using a quantitative proteomic method. Our proteomic data demonstrate that the major effect of Ipom-F on MDA-MB-231 cells is the inhibition of membrane and secreted protein expression. Our proteomic data are consistent with the recently uncovered molecular mechanism of action of Ipom-F, which binds to Sec61-α and inhibits the co-translational import of proteins into the endoplasmic reticulum. We have defined a subset of membrane and secreted proteins that are particularly sensitive to Ipom-F. Analysis of the expression of these Ipom-F-sensitive proteins in cancer cell lines and breast cancer tissues demonstrates that some of these proteins are upregulated in TNBC cells. Thus, it is likely that TNBC cells may have adapted to the elevated levels of some proteins identified as sensitive to Ipom-F in this study; inhibition of the expression of these proteins leads to a crisis in proliferation and/or survival for the cells.

Irisin preserves mitochondrial integrity and function in tubular epithelial cells after ischemia-reperfusion-induced acute kidney injury.

A myokine secreted by skeletal muscles during exercise called irisin mitigates ischemia-reperfusion (I/R) injury in epithelial cells of various organs by limiting damage to mitochondria. We test whether irisin may preserve the mitochondrial integrity and function in renal tubular epithelial cells and protect against ischemia-reperfusion-induced acute kidney injury (AKI). We correlated serum irisin levels with serum creatinine and BUN levels from both AKI patients and healthy individuals. In mice with irisin administration, various renal injury markers such as serum creatinine, BUN, kidney injury molecule-1 (Kim-1), and neutrophil gelatinase-associated lipocalin (NGAL), and renal histopathology were assessed after I/R. To identify the potential mechanisms of the protective of irisin's protective effect, we perfused proximal tubules under confocal microscopy and analyzed kidney tissues by qPCR, western blot, and immunohistochemistry. Serum irisin correlated inversely with serum creatinine and BUN levels were significantly lower in AKI patients than in healthy subjects. Administering irisin to mice after I/R decreased biomarker levels for AKI including serum creatinine, BUN, Kim-1, NAGL and lessened histological changes. In kidney tissues of mice, irisin upregulated the mitochondrial autophagy marker protein microtubule-associated protein 1 light chain 3 (LC3), the mitochondrial autophagy pathway-related proteins PTEN-induced putative kinase 1 (PINK1) and Parkinson's disease 2 parkin (PARK2) and downregulated the reactive substrate protein sequestosome 1 (P62) and mitochondrial membrane proteins translocase of outer mitochondrial membrane 20 (TOM20) and translocase of inner mitochondrial membrane 23 (TIM23). Irisin protects against renal I/R injury, which may involve the preservation of mitochondrial integrity and function.

TRPML1 activation ameliorates lysosomal phenotypes in CLN3 deficient retinal pigment epithelial cells

Mutations in the lysosomal membrane protein CLN3 cause Juvenile Neuronal Ceroid Lipofuscinosis (JNCL). Activation of the lysosomal ion channel TRPML1 has previously been shown to be beneficial in several neurodegenerative disease models. Here, we tested whether TRPML1 activation rescues disease-associated phenotypes in CLN3-deficient retinal pigment epithelial (ARPE-19 CLN3-KO) cells. ARPE-19 CLN3-KO cells accumulate LAMP1 positive organelles and show lysosomal storage of mitochondrial ATPase subunit C (SubC), globotriaosylceramide (Gb3), and glycerophosphodiesters (GPDs), whereas lysosomal bis(monoacylglycero)phosphate (BMP/LBPA) lipid levels were significantly decreased. Activation of TRPML1 reduced lysosomal storage of Gb3 and SubC but failed to restore BMP levels in CLN3-KO cells. TRPML1-mediated decrease of storage was TFEB-independent, and we identified TRPML1-mediated enhanced lysosomal exocytosis as a likely mechanism for clearing storage including GPDs. Therefore, ARPE-19 CLN3-KO cells represent a human cell model for CLN3 disease showing many of the described core lysosomal deficits, some of which can be improved using TRPML1 agonists.

Assessment of the degree of disability of the Polish NBIA-MPAN population based on a patient survey

Neurodegeneration with brain iron accumulation (NBIA) describes a group of extremely rare hereditary disorders with an estimated prevalence of 1–3/1,000,000. Mitochondrial membrane protein associated neurodegeneration (MPAN) is the third most common form of NBIA, accounting for 10% of NBIA cases globally and up to 50% of cases in Poland. This survey was conducted by the Association NBIA Poland to collect data on the functioning of patients with NBIA/MPAN based on self-reported EQ-5D-5L (EuroQoL 5-Dimension 5-Level) scores. A total of 50 patients living in Poland, including 26 patients with NBIA/MPAN, participated in the study. The mean age of NBIA/MPAN patients was 22 years; the mean age at symptom onset is 10 years. At the time of the survey, most patients were disabled and required assistance with daily activities. More than 50% of participants needed help with hygiene or dressing, and 40% were unable to walk on their own. Nearly 50% of patients declared symptoms indicating lowered mood. The survey showed that NBIA/MPAN leads to significant disability within a short period of time after symptom onset. Intensive work is underway to discover the role of the C19orf12 protein and develop therapeutic strategies. Actions are necessary to improve the comfort of everyday life of patients with NBIA/MPAN and to improve access to rehabilitation, including home-based programmes.

Aqueous Ionic Liquid Mixtures as Minimal Models of Lipid Bilayer Membranes.

We introduce aqueous ionic liquid (IL) mixtures, specifically mixtures of 1-butyl-3-imidazoliumtetrafluoroborate (BMImBF4), with water as a minimal model of lipid bilayer membranes. Imidazolium-based ILs are known to form clustered nanoscale structures in which local inhomogeneities, micellar or lamellar structures, are formed to shield hydrophobic parts of the cation from the polar cosolvent (water). To investigate these nanostructures, dynamic light scattering (DLS) on samples with different mixing ratios of water and BMImBF4 was performed. At mixing ratios of 50% and 45% (v/v), small and homogeneous nanostructures can indeed be detected. To test whether, in particular, these stable nanostructures in aqueous mixtures may mimic the effects of phospholipid bilayer membranes, we further investigated their interaction with myelin basic protein (MBP), a peripheral, intrinsically disordered membrane protein of the myelin sheath. Using dynamic light scattering (DLS), continuous wave (CW) and pulse electron paramagnetic resonance (EPR), and small-angle X-ray scattering (SAXS) on recombinantly produced, "healthy" charge variants rmC1WT and double cysteine variant C1S17CH85C, we find that the size and the shape of the determined nanostructures in an optimum mixture offer model membranes in which the protein exhibits native behavior. SAXS measurements illuminate the size and shape of the nanostructures and indicate IL-rich "beads" clipped together by functional MBP, one of the in vivo roles of the protein in the myelin sheath. All the gathered data combined indicate that the 50% and 45% aqueous IL mixtures can be described as offering minimal models of a lipid mono- or bilayer that allow native processing and potential study of at least peripheral membrane proteins like MBP.

Molecular and serological investigation of Francisella tularensis among wild animals in Yamaguchi prefecture.

Francisella tularensis is an intracellular gram-negative bacterium known as the causative agent of tularemia, which can be transmitted to humans by direct contact with wild animals or by tick bites. Although F. tularensis is highly pathogenic, its recent prevalence in Japan is underreported due to the small number of reported cases. To clarify the current situation of F. tularensis in wild animals, we conducted surveillance on various species of wild animals in Yamaguchi prefecture. In this study, we screened 809 samples collected from 90 Japanese black bears, 105 Japanese monkeys, 168 sika deer, 205 wild boars, and 84 bats. For seroprevalence analysis, we tested 177 serum samples from 75 black bears and 102 monkeys using the microagglutination test. The results showed that serums from five black bears exhibited slight agglutination. Western blot was performed as a confirmatory test on these five samples, but no positive signals were detected. Additionally, molecular surveillance was conducted using DNA extracted from 464 whole blood and 168 tissues, targeting the gene encoding 23 KDa hypothetical protein by real-time PCR and outer membrane protein A gene by conventional PCR. No positive samples of F. tularensis were detected by either real-time or conventional PCR. Although we did not detect any F. tularensis-positive samples through serological and molecular analyses, continuous surveillance studies are necessary since sporadic human cases have been reported in Japan.

The Effect of Reactive Oxygen Species on Respiratory Complex I Activity in Liposomes.

Respiratory complex I (R-CI) is an essential enzyme in the mitochondrial electron transport chain but also a major source of reactive oxygen species (ROS), which are implicated in neurodegenerative diseases and ageing. While the mechanism of ROS production by R-CI is well-established, the feedback of ROS on R-CI activity is poorly understood. Here, we perform EPR spectroscopy on R-CI incorporated in artificial membrane vesicles to reveal that ROS (particularly hydroxyl radicals) reduce R-CI activity by making the membrane more polar and by increasing its hydrogen bonding capability. Moreover, the mechanism that we have uncovered reveals that the feedback of ROS on R-CI activity via the membrane is transient and not permanent; lipid peroxidation is negligible for the levels of ROS generated under these conditions. Our successful use of modular proteoliposome systems in conjunction with EPR spectroscopy and other biophysical techniques is a powerful approach for investigating ROS effects on other membrane proteins.

Hybrid framework for membrane protein type prediction based on the PSSM

Membrane proteins are considered the major source of drug targets and are indispensable for drug design and disease prevention. However, traditional biomechanical experiments are costly and time-consuming; thus, many computational methods for predicting membrane protein types are gaining popularity. The position-specific scoring matrix (PSSM) method is an excellent method for describing the evolutionary information of protein sequences. In this study, we propose an improved capsule neural network (ICNN) model based on a capsule neural network to acquire sufficient relevant information from the PSSM. Furthermore, accounting for the complementarity between traditional machine learning and deep learning, we propose a hybrid framework that combines both approaches to predict protein types. This framework trains 41 baseline models based on the PSSM. The optimal subset features, selected after traversal, are fused using a two-level decision-level feature fusion approach. Subsequently, comparisons are made using three combined strategies within an ensemble learning framework. The experimental results demonstrate that solely relying on PSSM input, the proposed method not only surpasses the optimal methods by 1.52%\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\%$$\\end{document}, 2.26%\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\%$$\\end{document} and 2.67%\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\%$$\\end{document} on Dataset1, Dataset2, and Datasets3, respectively, but also exhibits superior generalizability. Furthermore, the code and dataset can be free download at https://github.com/ruanxiaoli/membrane-protein-types.

Metabolic perturbation and oxidative damage induced by tris(1,3-dichloro-2-propyl) phosphate (TDCIPP) and tris(2-ethylhexyl) phosphate (TEHP) on Escherichia coli through integrative analyses of metabolome

Organophosphate esters (OPEs) are one of the emerging environmental threats, causing the hazard to ecosystem safety and human health. Yet, the toxic effects and metabolic response mechanism after Escherichia coli (E.coli) exposed to TDCIPP and TEHP is inconclusive. Herein, the levels of SOD and CAT were elevated in a concentration-dependent manner, accompanied with the increase of MDA contents, signifying the activation of antioxidant response and occurrence of lipid peroxidation. Oxidative damage mediated by excessive accumulation of ROS decreased membrane potential and inhibited membrane protein synthesis, causing membrane protein dysfunction. Integrative analyses of GC-MS and LC-MS based metabolomics evinced that significant perturbation to the carbohydrate metabolism, nucleotide metabolism, lipids metabolism, amino acid metabolism, organic acids metabolism were induced following exposure to TDCIPP and TEHP in E.coli, resulting in metabolic reprogramming. Additionally, metabolites including PE(16:1(5Z)/15:0), PA(17:0/15:1(9Z)), PC(20:2(11Z,14Z)/12:0), LysoPC(18:3(6Z,9Z,12Z)/0:0) were significantly upregulated, manifesting that cell membrane protective molecule was afforded by these differential metabolites to improve permeability and fluidity. Overall, current findings generate new insights into the molecular toxicity mechanism by which E.coli respond to TDCIPP and TEHP stress and supply valuable information for potential ecological risks of OPEs on aquatic ecosystems.

Unraveling the impact of deleterious nsSNPs on the MFSD1 protein

MFSD1 (Major facilitator superfamily domain containing 1) protein is a lysosomal multiple transmembrane-spanning protein responsible for the active transport of various compounds. Due to its potential role in maintaining liver homeostasis, any mutation might lead to altered protein expression, thus affecting the functionality. In this study, we explored the impact of deleterious nsSNPs (Nonsynonymous single nucleotide polymorphisms) on the stability, conformation, and functionality of the MFSD1 protein. SNP data and MFSD1 protein sequences were retrieved from NCBI dbSNP and Uniprot respectively. In total, five highly conserved nsSNPs were predicted to be deleterious based on their negative impact on the stability of the protein. Furthermore, the simulation analysis on the 3D structures of both native and mutant proteins revealed a notable impact on the physiological conformation of the protein. The identified variants not only affect the native conformation but also impact the association of MFSD1 with GLMP (Glycosylated Lysosomal Membrane Protein). In conclusion, the analysis revealed that the mutant protein's structural stability was inferior to that of the native protein. This research provides crucial insights for identifying and assessing MFSD1 mutations as potential diagnostic markers for liver-related diseases.

Excitation of Reactive Oxygen Species and Damage to the Cell Membrane, Protein, and DNA are Important Inhibition Mechanisms of CO2 on Shewanella putrefaciens at 4 °C.

To explore whether oxidative stress caused by 100% CO2 is an inhibitory mechanism against Shewanella putrefaciens, the oxidative stress reaction, antioxidant activity, and damage to the cell membrane, protein, and DNA of CO2-incubated S. putrefaciens at 4 °C were evaluated. Research demonstrated that CO2 caused more severe reactive oxygen species (ROS) accumulation. Simultaneously, weaker •OH/H2O2/O2•--scavenging activity and decreased T-VOC and GSH content were also observed. The activities of antioxidant enzymes (SOD, POD, CAT, and GPX) continuously declined, which might be attributed to the CO2-mediated decrease in the pH value. Correspondingly, the cell membrane was damaged with hyperpolarization, increased permeability, and more severe lipid peroxidation. The expression of total and membrane protein decreased, and the synthesis and activity of extracellular protease were inhibited. DNA was also subjected to oxidative damage and expressed at a lower level. All results collaboratively confirmed that ROS excitation and inhibition of antioxidant activity were important inhibition mechanisms of CO2 on S. putrefaciens.

CPT2 Deficiency Modeled in Zebrafish: Abnormal Neural Development, Electrical Activity, Behavior, and Schizophrenia-Related Gene Expression.

Carnitine palmitoyltransferase 2 (CPT2) is an inner mitochondrial membrane protein of the carnitine shuttle and is involved in the beta-oxidation of long chain fatty acids. Beta-oxidation provides an alternative pathway of energy production during early development and starvation. CPT2 deficiency is a genetic disorder that we recently showed can be associated with schizophrenia. We hypothesize that CPT2 deficiency during early brain development causes transcriptional, structural, and functional abnormalities that may contribute to a CNS environment that is susceptible to the emergence of schizophrenia. To investigate the effect of CPT2 deficiency on early vertebrate development and brain function, CPT2 was knocked down in a zebrafish model system. CPT2 knockdown resulted in abnormal lipid utilization and deposition, reduction in body size, and abnormal brain development. Axonal projections, neurotransmitter synthesis, electrical hyperactivity, and swimming behavior were disrupted in CPT2 knockdown zebrafish. RT-qPCR analyses showed significant increases in the expression of schizophrenia-associated genes in CPT2 knockdown compared to control zebrafish. Taken together, these data demonstrate that zebrafish are a useful model for studying the importance of beta-oxidation for early vertebrate development and brain function. This study also presents novel findings linking CPT2 deficiency to the regulation of schizophrenia and neurodegenerative disease-associated genes.

Disulfide-Driven Charge and Hydrophobicity Rearrangement of Remodeled Membrane Proteins toward Amyloid-Type Aggregation.

As a common pathological hallmark, protein aggregation into amyloids is a highly complicated phenomenon, attracting extensive research interest for elucidating its structural details and formation mechanisms. Membrane deposition and disulfide-driven protein misfolding play critical roles in amyloid-type aggregation, yet the underlying molecular process remains unclear. Here, we employed sum frequency generation (SFG) vibrational spectroscopy to comprehensively investigate the remodeling process of lysozyme, as the model protein, into amyloid-type aggregates at the cell membrane interface. It was discovered that disulfide reduction concurrently induced the transition of membrane-bound lysozyme from predominantly α-helical to antiparallel β-sheet structures, under a mode switch of membrane interaction from electrostatic to hydrophobic, and subsequent oligomeric aggregation. These findings shed light on the systematic understanding of dynamic molecular mechanisms underlying membrane-interactive amyloid oligomer formation.

ALS-linked VapB P56S mutation alters neuronal mitochondrial turnover at the synapse.

Mitochondrial population maintenance in neurons is essential for neuron function and survival. Contact sites between mitochondria and the endoplasmic reticulum (ER) are poised to regulate mitochondrial homeostasis in neurons. These contact sites can function to facilitate transfer of calcium and lipids between the organelles and have been shown to regulate aspects of mitochondrial fission and fusion dynamics. VapB is an ER membrane protein present at a subset of ER-mitochondria contact sites. Mutations in VapB cause neurodegenerative disease. Specifically, a proline-to-serine mutation at amino acid 56 (P56S), correlates with susceptibility to amyotrophic lateral sclerosis (ALS) type 8. Given the relationship between failed mitochondrial health and neurodegenerative disease, we investigated the function of VapB in mitochondrial population maintenance. We demonstrate that transgenic expression of VapBP56S in zebrafish larvae (sex undetermined) increased mitochondrial biogenesis, causing increased mitochondrial population size in the axon terminal. Expression of wild type VapB did not alter biogenesis but, instead, increased mitophagy in the axon terminal. Using genetic manipulations to independently increase mitochondrial biogenesis in zebrafish neurons, we show that biogenesis is normally balanced by mitophagy to maintain a constant mitochondrial population size. VapBP56S transgenics fail to increase mitophagy to compensate for the increase in mitochondrial biogenesis, suggesting an impaired mitophagic response. Finally, using a synthetic ER-mitochondria tether, we show that VapB's function in mitochondrial turnover is likely independent of ER-mitochondrial tethering by contact sites. Our findings demonstrate that VapB can control mitochondrial turnover in the axon terminal, and this function is altered by the P56S ALS-linked mutation.Significance statement Mitochondrial population dysfunction is tightly tied to neurodegenerative diseases, including ALS. Maintenance of the mitochondrial population in neurons requires the birth of new mitochondria and the degradation of damaged organelles. ER-mitochondrial contact site proteins are in a position to regulate both processes in neurons. Our work demonstrates that an ALS-associated mutation in the contact site protein VapB disrupts both processes, identifying VapB as a mediator of regulated mitochondrial turnover to maintain a steady-state mitochondrial population.

EMP3 inhibits the formation of heterotypic cell-in-cell structures in hepatocellular carcinoma

Heterotypic cell-in-cell (heCIC) structures represent a unique intercellular interaction where tumor cells internalize immune cells to enhance the killing efficiency of immune cells. However, the mechanism of heCIC structure formation remains to be fully elucidated. In this study, we explored the role of epithelial membrane protein 3 (EMP3), a PMP-22/EMP/MP20 protein family member highly expressed in the patients with hepatocellular carcinoma and poor prognosis, in the formation of the heCIC structure formed by natural killer cells and hepatocellular carcinoma cells. The analysis of monoclonal hepatocellular carcinoma cell lines revealed that EMP3 presented low expression in the cells with high capability to form heCIC structure and high expression in those with low capability. Knocking down the expression of EMP3 by gene editing promoted the formation of heCIC structures, while overexpression of EMP3 significantly inhibited this process. Additionally, the expression of factors involved in the heCIC structure formation suggested that EMP3 inhibited the formation of heCIC structures by modulating the adhesion ability and cytoskeleton of tumor cells. The findings lay a foundation for enhancing the heCIC-mediated tumor immunotherapy by targeting EMP3.

SLC45A4 encodes a mitochondrial putrescine transporter that promotes GABA de novo synthesis.

Solute carriers (SLC) are membrane proteins that facilitate the transportation of ions and metabolites across either the plasma membrane or the membrane of intracellular organelles. With more than 450 human genes annotated as SLCs, many of them are still orphan transporters without known biochemical functions. We developed a metabolomic-transcriptomic association analysis, and we found that the expression of SLC45A4 has a strong positive correlation with the cellular level of γ-aminobutyric acid (GABA). Using mass spectrometry and the stable isotope tracing approach, we demonstrated that SLC45A4 promotes GABA de novo synthesis through the Arginine/Ornithine/Putrescine (AOP) pathway. SLC45A4 functions as a putrescine transporter localized to the mitochondrial membrane to facilitate GABA production. Taken together, our results revealed a new biochemical mechanism where SLC45A4 controls GABA production.

Mechanosensitive Channels: Utilising the Bacterial Cell’s Defence Mechanism against It

Mechanosensitive channels are integral membrane proteins in bacterial cell membranes as well as archaea, and eukaryotes. The role of these bacterial mechanosensitive channels is essential for protecting cells from structural damage during hypoosmotic shock. Mechanosensitive channel of small conductance (MscS) and mechanosensitive channels of large conductance (MscL) are the predominant channels in E. coli. Activation of mechanosensitive channels typically occurs when the bacterial membrane senses tension or distortion. Overall, the ecological niche of bacteria shapes the selective pressures acting on mechanosensitive channels, leading to their adaptation to specific environmental conditions. Bacterial mechanosensitive channels contribute to the ability of bacterial pathogens to adapt to and survive within the host environment, as well as to modulate interactions with host cells and tissues during infection. They could also act as entrance gates for specific antibiotic classes into bacterial cells. Accordingly, it was discovered that nitrofuran nifuroxazide and tuberactinomycin viomycin depended on both Ec-MscS and Ec-MscL for enhanced efficacy. Several compounds have been identified that directly target mechanosensitive channels. For example, ramizol has been shown to reduce the gating threshold of MscL and MscS channels, while styrylbenzene inhibits MscL channels in S. aureus, Streptococcus pneumoniae, and Clostridium difficile. Bacterial mechanosensitive channels are versatile drug targets due to their role in promoting bacterial virulence and host colonization, serving as entry points for antibiotics, and being structurally distinct from mammalian counterparts. These special structures which are meant to protect the cell can also be harnessed as drug targets, thereby increasing the susceptibility of the bacterial cell to target antibiotics. This feature has been harnessed to reduce the burden of antibiotics resistance and its attendant effect on global healthcare complications. The aim of this review is to examine the various physiological, drug resistance and pathogenicity roles of these mechanosensitive channels as well as their usefulness as potential drug targets.

Unlocking the potential of higher-molecular-weight 5-HT7R ligands: Synthesis, affinity, and ADMET examination

An increasing number of drugs introduced to the market and numerous repositories of compounds with confirmed activity have posed the need to revalidate the state-of-the-art rules that determine the ranges of properties the compounds should possess to become future drugs. In this study, we designed a series of two chemotypes of aryl-piperazine hydantoin ligands of 5-HT7R, an attractive target in search for innovative CNS drugs, with higher molecular weight (close to or over 500). Consequently, 14 new compounds were synthesised and screened for their receptor activity accompanied by extensive docking studies to evaluate the observed structure–activity/properties relationships. The ADMET characterisation in terms of the biological membrane permeability, metabolic stability, hepatotoxicity, cardiotoxicity, and protein plasma binding of the obtained compounds was carried out in vitro. The outcome of these studies constituted the basis for the comprehensive challenge of computational tools for ADMET properties prediction. All the compounds possessed high affinity to the 5-HT7R (Ki below 250 nM for all analysed structures) with good selectivity over 5-HT6R and varying affinity towards 5-HT2AR, 5-HT1AR and D2R. For the best compounds of this study, the expression profile of genes associated with neurodegeneration, anti-oxidant response and anti-inflammatory function was determined, and the survival of the cells (SH-SY5Y as an in vitro model of Alzheimer’s disease) was evaluated. One 5-HT7R agent (32) was characterised by a very promising ADMET profile, i.e. good membrane permeability, low hepatotoxicity and cardiotoxicity, and high metabolic stability with the simultaneous high rate of plasma protein binding and high selectivity over other GPCRs considered, together with satisfying gene expression profile modulations and neural cell survival. Such encouraging properties make it a good candidate for further testing and optimisation as a potential agent in the treatment of CNS-related disorders.