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The outer membrane (OM) is an essential component of the Gram-negative bacterial cell envelope. Restricted diffusion of integral OM proteins and lipopolysaccharide (LPS) that constitute the outer leaflet of the OM support a model in which the OM is in a semi-crystalline state. The low fluidity of the OM has been suggested to be an important property of this membrane that even contributes to cell rigidity. The LPS characteristics strongly determine the properties of the OM and the LPS layer fluidity has been measured using different techniques that require specific conditions or are technically challenging. Here, we characterize the Escherichia coli LPS fluidity by evaluating the lateral diffusion of the styryl dye FM4-64FX in fluorescence recovery after photobleaching experiments. This technique allowed us to determine the effect of different conditions and genetic backgrounds on the LPS fluidity. Our results show that a fraction of the LPS can slowly diffuse and that the fluidity of the LPS layer adapts by modifying the diffusion of the LPS and the fraction of mobile LPS molecules.

Small heat-shock proteins (sHSP) are ubiquitous ATP-independent chaperones that prevent irreversible aggregation of heat-damaged denaturing proteins. Lactiplantibacillus plantarum is a widespread Gram-positive bacterium with probiotic claims and vast potential for agro-food, biotechnological and biomedical applications. L. plantarum possesses a family of three sHSP, which were previously demonstrated to be involved in its stress tolerance mechanisms. Here, the three L. plantarum sHSP were heterologously expressed, purified and shown to have a chaperone activity in vitro, measuring their capacity to suppress protein aggregation, as assayed spectrophotometrically by light scattering. Their anti-aggregative capacity was found to be differently influenced by pH. Differences were also found relative to their holdase function and their capacity to modulate liposome membrane fluidity, suggesting interplays between them and indicating diversified activities. This is the first study assessing the chaperone action of sHSP from a probiotic model. The different roles of the three sHSP can increase L. plantarum’s capabilities to survive the various types of stress characterising the diverse habitats of this highly adaptable species. Reported evidence supports the interest in L. plantarum as one of the model species for bacteria that have three different sHSP-encoding genes in their genomes.

Ruscogenin, a kind of steroid saponin, has been shown to have significant anti-oxidant, anti-inflammatory, and anti-thrombotic characteristics. Furthermore, it has the potential to be employed as a medicinal medication to treat a variety of acute and chronic disorders. The interaction of a drug molecule with cell membranes can help to elucidate its system-wide protective and therapeutic effects, and it's also important for its pharmacological activity. The molecular mechanism by which ruscogenin affects membrane architecture is still a mystery. Ruscogenin's interaction with zwitterionic dipalmitoyl phosphatidylcholine (DPPC) and anionic dipalmitoyl phosphatidylglycerol (DPPG) multilamellar vesicles (MLVs) was studied utilizing two non-invasive approaches, including: Fourier Transform Infrared (FTIR) spectroscopy and Differential Scanning Calorimetry. Ruscogenin caused considerable alterations in the phase transition profile, order, dynamics and hydration state of head groups and glycerol backbone of DPPC and DPPG MLVs at all concentrations. The DSC results indicated that the presence of ruscogenin decreased the main phase transition temperature (Tm) and enthalpy (?H) values of both membranes and increased half height width of the main transition (?T1/2). The FTIR results demonstrated that all concentrations (1, 3, 6, 9, 15, 24 and 30 mol percent) of ruscogenin disordered the DPPC MLVs both in the gel and liquid crystalline phases while it increased the order of DPPG MLVs in the liquid crystalline phase. Moreover, ruscogenin caused an increase in the dynamics of DPPC and DPPG MLVs in both phases. Additionally, it enhanced the hydration of the head groups of lipids and the surrounding water molecules implying ruscogenin to interact strongly with both zwitterionic and charged model membranes. © 2022 Elsevier Inc., Ege Üniversitesi: FGA-2021-22592, This work was supported by Ege University Scientific Research Projects Coordination. (Project number is FGA-2021-22592)., This work was supported by Ege University Scientific Research Projects Coordination . (Project number is FGA-2021-22592 ).

Myoblast migration is crucial for myogenesis and muscular tissue homeostasis. However, its spatiotemporal control remains elusive. Here, we explored the involvement of plasma membrane cholesterol and sphingolipids in this process. In resting C2C12 mouse myoblasts, those lipids clustered in sphingomyelin/cholesterol/GM1 ganglioside (SM/chol/GM1)- and cholesterol (chol)-enriched domains, which presented a lower stiffness than the bulk membrane. Upon migration, cholesterol and sphingomyelin polarized at the front, forming cholesterol (chol)- and sphingomyelin/cholesterol (SM/chol)-enriched domains, while GM1-enriched domains polarized at the rear. A comparison of domain proportion suggested that SM/chol- and GM1-enriched domains originated from the SM/chol/GM1-coenriched domains found at resting state. Modulation of domain proportion (through cholesterol depletion, combined or not with actin polymerization inhibition, or sphingolipid synthesis inhibition) revealed that the higher the chol- and SM/chol-enriched domains, the higher the myoblast migration. At the front, chol- and SM/chol-enriched domains were found in proximity with F-actin fibers and the lateral mobility of sphingomyelin in domains was specifically restricted in a cholesterol- and cytoskeleton-dependent manner while domain abrogation impaired F-actin and focal adhesion polarization. Altogether, we showed the polarization of cholesterol and sphingomyelin and their clustering in chol- and SM/chol-enriched domains with differential properties and roles, providing a mechanism for the spatial and functional control of myoblast migration.

Voltage-gated proton channels (HV 1) have been found in many mammalian cells and play a crucial role in the immune system, male fertility, and cancer progression. Glycosaminoglycans play a significant role in various aspects of cell physiology, including the modulation of membrane receptors and ion channel function. We present here evidence that mechanosensitivity of the dimeric HV 1 channel transduce changes on cell membrane fluidity related to the defective biosynthesis of chondroitin sulphate and heparan sulphate in Chinese Hamster Ovary (CHO-745) cells into a leftward shift in the activation voltage dependence. This effect was accompanied by an increase in the H+ current, and an acceleration of the activation kinetics, under symmetrical or asymmetrical pH gradient (ΔpH) conditions. Similar gating alterations were evoked by two naturally occurring HV 1 N-terminal truncated isoforms expressed in wild-type CHO-K1 and CHO-745 cells. On three different monomeric HV 1 constructs, no alterations in the biophysical parameters were observed. Moreover, we have showed that HV 1 gating can be modulated by manipulating CHO-K1 cell membrane fluidity. Our results suggest that the defective biosynthesis of chondroitin sulphate and heparan sulphate on CHO-745 cell increases membrane fluidity and allosterically modulates the coupling between voltage- and ΔpH-sensing through the dimeric HV 1 channel.

In bacteria, exposure to changes in environmental conditions can alter membrane fluidity, thereby affecting its essential functions in cell physiology. To adapt to these changes, bacteria maintain appropriate fluidity by varying the composition of the fatty acids of membrane phospholipids, a phenomenon known as homeophasic adaptation. In Pseudomonas aeruginosa, this response is achieved mainly by two mechanisms of fatty acid desaturation: the FabA-FabB and DesA-DesB systems. This study analyzed the effect of ultraviolet-A (UVA) radiation-the major fraction of solar UV radiation reaching the Earth's surface-on the homeophasic process. The prototypical strain PAO1 was grown under sublethal UVA doses or in the dark, and the profiles of membrane fatty acids were compared at early logarithmic, logarithmic and stationary growth phases. In the logarithmic growth phase, it was observed that growth under sublethal UVA doses induced the expression of the desaturase-encoding genes desA and desB and increased the proportion of unsaturated fatty acids; in addition, membrane fluidity could also increase, as suggested by the indices used as indicators of this parameter. The opposite effect was observed in the stationary growth phase. These results demonstrate the relevant role of UVA on the homeophasic response at transcriptional level.

Protein phosphatase Z1 (Ppz1) has been shown to take part in important physiological functions in fungi including a contribution to virulence of Candida albicans. Although its involvement in the oxidative stress response has also been documented, the exact mechanism of action of its protective effect against oxidative damage remains unknown. By developing a pipeline to analyze the biophysical properties of the cell membrane in fungi, we demonstrate that the plasma membrane of Ppz1-KO Candida albicans displays increased sensitivity to tert-butyl-hydroperoxide-induced oxidative damage. In particular, the response to the oxidizing agent, characterized by increased lipid peroxidation, reduced lipid order, and inhibited lateral mobility of plasma membrane components, is significantly more pronounced in the Ppz1-KO C. albicans strain than in the wild-type counterpart. Remarkably, membrane constituents became almost completely immobile in the phosphatase deletion mutant exposed to oxidative stress. Furthermore, moderately elevated membrane lipid peroxidation accompanied by the aforementioned changes in the biophysical characteristics of the plasma membrane are already detectable in untreated Ppz1-KO cells indicating latent membrane damage even in the absence of oxidative stress. In conclusion, the hypersensitivity of cells lacking Ppz1 to oxidative damage establishes that potential Ppz1 inhibitors may synergize with oxidizing agents in prospective anti-fungal combination therapies.

1,2-dibehenoyl-sn-glycero-3-phosphocholine (DBPC) is one of the important phospholipids found in cell membrane but not studied well. Importance of curcumin as a dietary supplement and for its medicinal properites is getting widely recoginsed. The present study for the first time explores the effect of curcumin on properties of DBPC liposomes by monitoring the fluorescence properties of curcumin. The phase transition temperature (Tm) of DBPC is assessed which confirms increase in curcumin concentration causes a slight drop in the Tm value. Chitosan is being applied for various drug delivery uses. The study establishes new insight on effect of chitosan oligosaccharide lactate on DBPC liposomes. It is found that in the absence of chitosan oligosaccharide lactate, curcumin partitions more strongly in the liquids crystalline phase than in the solid gel phase, however, the opposite result is obtained with the presence of chitosan oligosaccharide lactate which penetrates into the DBPC liposomes membranes at higher temperature, blocking thus the passage of curcumin into the lipid bilayer. However, addition of chitosan oligosaccharide lactate had no effect on the Tm. Fluorescence quenching study of curcumin establishes that the location of curcumin to be in the hydrophobic cavity of DBPC membrane.

There are still many challenges in understanding the absorption and transport mechanism of liposomes in the gastrointestinal tract of infants, especially for liposome-coentrapped two or more substances. In this study, novel docosahexaenoic acid (DHA)-anthocyanidin-codelivery liposomes (DA-LPs) were fabricated and characterized, and their digestive and absorptive behaviors were evaluated using the in vitro infant digestive method combined with the Caco-2 cell model. The liposomal bilayer structure remained intact with the particles aggregated in simulated infant gastric fluid, while their phospholipid membrane underwent enzymatic lipolysis under simulated intestinal conditions. Compared to single substance-loaded liposomes (DHA- or anthocyanidin-loaded liposomes), the digested DA-LPs showed better cell viability, higher cellular uptake and membrane fluidity, and lower reactive oxygen species (ROS). It can be concluded that DA-LPs are promising carriers for simultaneously transporting hydrophobic and hydrophilic molecules and may be beneficial for improving nutrient absorption and alleviating intestinal stress oxidation.

Excessive accumulation of amyloid β-protein (Aβ) is one of the primary mechanisms that leads to neuronal death with phosphorylated tau in the pathogenesis of Alzheimer's disease (AD). Protofibrils, one of the high-molecular-weight Aβ oligomers (HMW-Aβo), are implicated to be important targets of disease modifying therapy of AD. We previously reported that phenolic compounds such as myricetin inhibit Aβ1-40, Aβ1-42, and α-synuclein aggregations, including their oligomerizations, which may exert protective effects against AD and Parkinson's disease. The purpose of this study was to clarify the detailed mechanism of the protective effect of myricetin against the neurotoxicity of HMW-Aβo in SH-SY5Y cells. To assess the effect of myricetin on HMW-Aβo-induced oxidative stress, we systematically examined the level of membrane oxidative damage by measuring cell membrane lipid peroxidation, membrane fluidity, and cell membrane potential, and the mitochondrial oxidative damage was evaluated by mitochondrial permeability transition (MPT), mitochondrial reactive oxygen species (ROS), and manganese-superoxide dismutase (Mn-SOD), and adenosine triphosphate (ATP) assay in SH-SY5Y cells. Myricetin has been found to increased cell viability by suppression of HMW-Aβo-induced membrane disruption in SH-SY5Y cells, as shown in reducing membrane phospholipid peroxidation and increasing membrane fluidity and membrane resistance. Myricetin has also been found to suppress HMW-Aβo-induced mitochondria dysfunction, as demonstrated in decreasing MPT, Mn-SOD, and ATP generation, raising mitochondrial membrane potential, and increasing mitochondrial-ROS generation. These results suggest that myricetin preventing HMW-Aβo-induced neurotoxicity through multiple antioxidant functions may be developed as a disease-modifying agent against AD.

Background Biomolecular condensates are non-stoichiometric assemblies that are characterized by their capacity to spatially concentrate biomolecules and play a key role in cellular organization. Proteins that drive the formation of biomolecular condensates frequently contain oligomerization domains and intrinsically disordered regions (IDRs), both of which can contribute multivalent interactions that drive higher-order assembly. Our understanding of the relative and temporal contribution of oligomerization domains and IDRs to the material properties of in vivo biomolecular condensates is limited. Similarly, the spatial and temporal dependence of protein oligomeric state inside condensates has been largely unexplored in vivo. Methods In this study, we combined quantitative microscopy with number and brightness analysis to investigate the aging, material properties, and protein oligomeric state of biomolecular condensates in vivo. Our work is focused on condensates formed by AUXIN RESPONSE FACTOR 19 (ARF19), a transcription factor integral to the auxin signaling pathway in plants. ARF19 contains a large central glutamine-rich IDR and a C-terminal Phox Bem1 (PB1) oligomerization domain and forms cytoplasmic condensates. Results Our results reveal that the IDR amino acid composition can influence the morphology and material properties of ARF19 condensates. In contrast the distribution of oligomeric species within condensates appears insensitive to the IDR composition. In addition, we identified a relationship between the abundance of higher- and lower-order oligomers within individual condensates and their apparent fluidity. Conclusions IDR amino acid composition affects condensate morphology and material properties. In ARF condensates, altering the amino acid composition of the IDR did not greatly affect the oligomeric state of proteins within the condensate.

Since the phasing out and eventual ban on the production of organohalogen flame retardants, the use of organophosphate flame retardants (OPFRs) has increased rapidly. This has led to the detection of OPFRs in various environments including the Arctic. Two of the most prevalent OPFRs found in the Arctic are tris(2-chloroisopropyl) phosphate (TCPP), and 2-ethylhexyl diphenyl phosphate (EHDPP). The impacts of exposure to OPFRs on Arctic organisms is poorly understood. The objective of the present study was to determine the effects of exposure to TCPP, EHDPP, and a mixture of OPFRs on gene expression patterns in Atlantic cod, Gadus morhua. Precision-cut liver slices from Atlantic cod in vitro were exposed to either TCPP or EHDPP alone or in a mixture and sampled at 2 different time points to quantify gene expression patterns using RNA sequencing. We exposed the liver slices to 2 concentrations of TCPP and EHDPP, one of which was chosen based on the levels found in the Arctic environment. The RNA sequencing results demonstrated differential expression of hundreds of genes in response to exposure. The genes representing cholesterol biosynthesis and lipid metabolism pathway were significantly enriched in all the treatment groups. Almost all the cholesterol biosynthesis genes were significantly down-regulated in response to OPFR exposure. The effects on these pathways could involve various physiological processes including reproduction, growth, and behavior as well as adaptation to changing temperatures. Membrane fluidity is an important adaptive mechanism among aquatic organisms. Altered cholesterol homeostasis could have long-term consequences by altering the adaptive potential of aquatic organisms to changing water temperatures, particularly those living in polar environments. These results suggest that OPFRs could have unique effects on the organisms living in the Arctic compared with other environments. Further studies are needed to understand the long-term impacts of exposure to environmentally realistic concentrations using laboratory and field-based studies. Environ Toxicol Chem 2021;40:1639-1648. © 2021 SETAC.

Listeria monocytogenes is a food-borne pathogen with the ability to grow at low temperatures down to − 0.4 °C. Maintaining cytoplasmic membrane fluidity by changing the lipid membrane composition is important during growth at low temperatures. In Listeria monocytogenes, the dominant adaptation effect is the fluidization of the membrane by shortening of fatty acid chain length. In some strains, however, an additional response is the increase in menaquinone content during growth at low temperatures. The increase of this neutral lipid leads to fluidization of the membrane and thus represents a mechanism that is complementary to the fatty acid-mediated modification of membrane fluidity. This study demonstrated that the reduction of menaquinone content for Listeria monocytogenes strains resulted in significantly lower resistance to temperature stress and lower growth rates compared to unaffected control cultures after growth at 6 °C. Menaquinone content was reduced by supplementation with aromatic amino acids, which led to a feedback inhibition of the menaquinone synthesis. Menaquinone-reduced Listeria monocytogenes strains showed reduced bacterial cell fitness. This confirmed the adaptive function of menaquinones for growth at low temperatures of this pathogen.

The membrane is the major protective barrier separating the cell from the environment and is thus important for bacteria to survive environmental stress. This study investigates changes in membrane lipid compositions and membrane physiology of meat spoiling bacteria in response to high CO2 (30%) and O2 (70%) concentrations, as commonly used for modified atmosphere packaging of meat. Therefore, the fatty acid profile as well as membrane fluidity, permeability and cell surface were determined and correlated to the genomic settings of five meat spoiling bacteria Brochothrix (B.) thermosphacta, Carnobacterium (C.) divergens, C. maltaromaticum, Leuconostoc (L.) gelidum subsp. gelidum and L. gelidum subsp. gasicomitatum cultivated under different gas atmospheres. We identified different genomic potentials for fatty acid adaptations, which were in accordance with actual measured changes in the fatty acid composition for each species in response to CO2 and/or O2, e.g., an increase in saturated, iso and cyclopropane fatty acids. Even though fatty acid changes were species-specific, the general physiological responses were similar, comprising a decreased membrane permeability and fluidity. Thus, we concluded that meat spoiling bacteria facilitate a change in membrane fatty acids upon exposure to O2 and CO2, what leads to alteration of membrane fluidity and permeability. The observed adaptations might contribute to the resistance of meat spoilers against detrimental effects of the gases O2 and CO2 and thus help to explain their ability to grow under different modified atmospheres. Furthermore, this study provides fundamental knowledge regarding the impact of fatty acid changes on important membrane properties of bacteria.

This review uses gangliosides as an example to characterize various aspects of studies of vertebrate brain lipids directed by Academician E. M. Kreps and continued by his colleagues. Brain gangliosides (like phospholipids) from cold-water stenothermal bony fish species have been shown to be characterized by a higher content of mono- and polyene fatty acids than the analogous brain lipids of warm-blooded stenothermal bony fish species. Changes in the fatty acid composition of fish brain lipids on adaptation to life in cold water (or at greater depth) are directed to maintaining the optimum level of brain cell membrane fluidity and microheterogeneity. Cluster analysis of data on the composition and structure of the carbohydrate component of brain gangliosides from different classes of ectothermal vertebrates were used to construct a dendrogram. This dendrogram was found to be similar to the evolutionary tree corresponding to the classical taxonomy of vertebrates. It is suggested that changes in the molecular organization of gangliosides during the process of evolution in vertebrates contributed to brain differentiation and increases in the complexity of its functions during phylogenetic development. The main gangliosides of the mammalian brain (GM1, GD1a, GD1b, and GT1b) protect neurons and PC12 cells from the actions of excitatory amino acids, hydrogen peroxide, and amyloid β peptide, their protective effects depending on activation of tyrosine kinase Trk receptors and downstream protein kinases (Akt, ERK1/2, protein kinase C). Another defensive mechanism uses gangliosides GM1 and GD1a against the toxic actions of bacterial lipopolysaccharide (LPS). This appears to be linked with changes in the composition of lipid rafts in the plasma membrane of nerve cells due to the inclusion of exogenous gangliosides, which leads to blockade of the translocation of TLR4 LPS receptors within them. Experiments using the Morris water test demonstrated the ability of gangliosides administered to rats with type 2 diabetes mellitus to prevent impairments to spatial memory. This was the first use of intranasal administration of gangliosides and its high efficacy was demonstrated.

For plants, acclimation to low temperatures is fundamental to survival. This process involves the modification of lipids to maintain membrane fluidity. We previously identified a new cold-induced putative desaturase in Physcomitrium (Physcomitrella) patens. Lipid profiles of null mutants of this gene lack sphingolipids containing monounsaturated C24 fatty acids, classifying the new protein as sphingolipid fatty acid denaturase (PpSFD). PpSFD mutants showed a cold-sensitive phenotype as well as higher susceptibility to the oomycete Pythium, assigning functions in stress tolerance for PpSFD. Ectopic expression of PpSFD in the Atads2.1 (acyl coenzyme A desaturase-like 2) Arabidopsis thaliana mutant functionally complemented its cold-sensitive phenotype. While these two enzymes catalyse a similar reaction, their evolutionary origin is clearly different since AtADS2 is a methyl-end desaturase whereas PpSFD is a cytochrome b5 fusion desaturase. Altogether, we suggest that adjustment of membrane fluidity evolved independently in mosses and seed plants, which diverged more than 500 million years ago. Sphingolipid desaturases can modify membrane lipids and change cold tolerance. Two of these enzymes have the same function in Arabidopsis and moss, but their evolutionary origins are different.

After antibiotic treatment, a subpopulation of bacteria often remains and can lead to recalcitrant infections. This subpopulation, referred to as persisters, evades antibiotic treatment through numerous mechanisms such as decreased uptake of small molecules and slowed growth. Membrane perturbing small molecules have been shown to eradicate persisters as well as render these populations susceptible to antibiotic treatment. Chemotype similarities have emerged suggesting amphiphilic heteroaromatic compounds possess ideal properties to increase membrane fluidity and such molecules warrant further investigation as effective agents or potentiators against persister cells.

Transmembrane anion transport is an important biological process in maintaining cellular functions. Thus, synthetic anion transporters are widely developed for their biological applications. Imidazolinium was introduced as anion recognition site to a multiblock amphiphilic structure that consists of octa(ethylene glycol) and aromatic units. Ion transport assay using halide-sensitive lucigenin and pH-sensitive 8-hydroxypyrene-1,3,6-trisulfonate (HPTS) revealed that imidazolinium-based multiblock amphiphile (IMA) transports anions and showed high selectivity for nitrate, which plays crucial roles in many biological events. Temperature-dependent ion transport assay using 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) indicated that IMA works as a mobile carrier. 1 H NMR titration experiments indicated that the C2 proton of the imidazolinium ring recognizes anions via a (C-H)+ ⋅⋅⋅X- hydrogen bond. Furthermore, all-atom molecular dynamics simulations revealed a dynamic feature of IMA within the membranes during ion transportation.

Chitosan is non-toxic, biodegradable and biocompatible. However, it is insoluble in water, which limits its applications in biomedical areas. Hydroxypropyltrimethyl ammonium chloride chitosan (HACC), a chitosan derivative, can be dissolved in physiological condition and has been widely used in the field of biomedicine and bioengineering. The biological effect of HACC has been extensively studied. However, it is rarely investigated at the subcellular level. To study the biological effect of HACC, mitochondria, energy-producing organelles in eukaryotes, were chosen as a model. The investigation mainly focused on the changes of mitochondrial membrane property in the presence of HACC. Results showed that HACC can induce the collapse of mitochondrial transmembrane potential (∆Ψm), the increase in mitochondrial membrane swelling and the decrease of mitochondrial membrane fluidity, demonstrating that mitochondrial membrane permeability transition pore (mPTP) opening happened. Possible mechanism of mPTP opening investigation indicated that it was occurred in a typical model. In addition, HACC can induce the release of cytochrome C (Cyt c) and affect the respiratory activity of mitochondria. The study will provide a lot of important information on biosafety evaluation of HACC.

Psychrophilic bacteria, living at low and mild temperatures, can contribute significantly to our understanding of microbial responses to temperature, markedly occurring in the bacterial membrane. Here, a newly isolated strain, Pseudomonas sp. B14-6, was found to dynamically change its unsaturated fatty acid and cyclic fatty acid content depending on temperature which was revealed by phospholipid fatty acid (PLFA) analysis. Genome sequencing yielded the sequences of the genes Δ-9-fatty acid desaturase (desA) and cyclopropane-fatty acid-acyl-phospholipid synthase (cfa). Overexpression of desA in Escherichia coli led to an increase in the levels of unsaturated fatty acids, resulting in decreased membrane hydrophobicity and increased fluidity. Cfa proteins from different species were all found to promote bacterial growth, despite their sequence diversity. In conclusion, PLFA analysis and genome sequencing unraveled the temperature-related behavior of Pseudomonas sp. B14-6 and the functions of two membrane-related enzymes. Our results shed new light on temperature-dependent microbial behaviors and might allow to predict the consequences of global warming on microbial communities.

The inhibitory effects of trehalose liposomes (TL) comprising l -α-dimyristoylphosphatidylcholine (DMPC) and α-D-glucopyranosyl-α-D-glucopyranoside monomyritate (TreC14) were investigated on breast cancer MDA-MB-453 cells in vitro and in vivo. The IC50 values of TL for MDA-MB-453 cells were remarkably lower than those of DMPC liposomes. The inhibitory effects of TL on the proliferation of MDA-MB-453 cells mediated via apoptosis induction were observed following their accumulation on MDA-MB-453 cell membranes. The membrane fluidity of MDA-MB-453 cells increased after TL treatment, as evident from a fluorescence depolarization assay. TL induced the apoptosis of MDA-MB-453 cells through caspase activation and mitochondrial membrane potential reduction, and suppressed the nuclear factor kappa B activity. A remarkable reduction in tumor volume was observed in a human breast cancer mouse model topically treated with TL. Induction of apoptosis was evident in TL-treated breast cancer tumors of mice using the TUNEL assay.

Changes in the membrane composition of sub-populations of cells can influence different properties with importance to tumour growth, metastasis and treatment efficacy. In this study, we use correlated fluorescence microscopy and ToF-SIMS with C60+ and (CO2)6k+ ion beams to identify and characterise sub-populations of cells based on successful transfection leading to over-expression of CCTδ, a component of the multi-subunit molecular chaperone named chaperonin-containing tailless complex polypeptide 1 (CCT). CCT has been linked to increased cell growth and proliferation and is known to affect cell morphology but corresponding changes in lipid composition of the membrane have not been measured until now. Multivariate analysis of the surface mass spectra from single cells, focused on the intact lipid ions, indicates an enrichment of phosphatidylethanolamine species in the transfected cells. While the lipid changes in this case are driven by the structural changes in the protein cytoskeleton, the consequence of phosphatidylethanolamine enrichment may have additional implications in cancer such as increased membrane fluidity, increased motility and an ability to adapt to a depletion of unsaturated lipids during cancer cell proliferation. This study demonstrates a successful fluorescence microscopy-guided cell by cell membrane lipid analysis with broad application to biological investigation.Graphical abstract

Myriocin damaged Fon cell membrane. The damage to the cell membrane of Fon was caused by the altered or disrupted expression of some membrane‐related genes and proteins at the mRNA and protein levels; these genes and proteins mainly including those related to sphingolipid metabolism, glycerophospholipid metabolism, steroid biosynthesis, ABC transporters, and protein processing in the endoplasmic reticulum., Summary Myriocin, which is produced by Bacillus amyloliquefaciens LZN01, can inhibit the growth of Fusarium oxysporum f. sp. niveum (Fon). In the present study, the antifungal mechanism of myriocin against Fon was investigated with a focus on the effects of myriocin on the cell membrane. Myriocin decreased the membrane fluidity and destroyed the membrane integrity of Fon. Significant microscopic morphological changes, including conidial shrinkage, the appearance of larger vacuoles and inhomogeneity of electron density, were observed in myriocin‐treated cells. A membrane‐targeted mechanism of action was also supported by transcriptomic and proteomic analyses; a total of 560 common differentially expressed genes (DEGs) and 285 common differentially expressed proteins (DEPs) were identified. The DEGs were further verified by using RT‐qPCR. The combined analysis between the transcriptome and proteome revealed that the expression of some membrane‐related genes and proteins, mainly those related to sphingolipid metabolism, glycerophospholipid metabolism, steroid biosynthesis, ABC transporters and protein processing in the endoplasmic reticulum, was disordered. Myriocin affected the serine palmitoyl transferase (SPT) activity as evidenced through molecular docking. Our results indicate that myriocin has significant antifungal activity owing to its ability to induce membrane damage in Fon.

The biosynthetic efficiency of curcumin, a highly bioactive compound from the plant Curcuma longa, needs to be improved. In this study, we performed host cell and biosynthetic pathway engineering to improve curcumin biosynthesis. Using in vivo-directed evolution, the expression level of curcuminoid synthase (CUS), the rate-limiting enzyme in the curcumin biosynthetic pathway, was significantly improved. Furthermore, as curcumin is a highly hydrophobic compound, two cell membrane engineering strategies were applied to optimize the biosynthetic efficiency. Curcumin storage was increased by overexpression of monoglucosyldiacylglycerol synthase from Acholeplasma laidlawii, which optimized the cell membrane morphology. Furthermore, unsaturated fatty acid supplementation was used to enhance membrane fluidity, which greatly ameliorated the damaging effect of curcumin on the cell membrane. These two strategies enhanced curcumin biosynthesis and demonstrated an additive effect.

Cell membranes contain incredible diversity in the chemical structures of their individual lipid species and the ratios in which these lipids are combined to make membranes. Nevertheless, our current understanding of how each of these components affects the properties of the cell membrane remains elusive, in part due to the difficulties in studying the dynamics of membranes at high spatiotemporal resolution. In this work, we use coarse-grained molecular dynamics simulations to investigate how individual lipid species contribute to the biophysical properties of the neuronal plasma membrane. We progress through eight membranes of increasing chemical complexity, ranging from a simple POPC/CHOL membrane to a previously published neuronal plasma membrane [Ingolfsson, H. I., et al. (2017) Biophys. J. 113 (10), 2271-2280] containing 49 distinct lipid species. Our results show how subtle chemical changes can affect the properties of the membrane and highlight the lipid species that give the neuronal plasma membrane its unique biophysical properties. This work has potential far-reaching implications for furthering our understanding of cell membranes.

Funder: National Collaborative Research Infrastructure Strategy (NCRIS), Funder: ANR/NSF-PIRE, Funder: Science and Technology Facilities Council, Funder: Institut Laue Langevin, Coronavirus disease-2019 (COVID-19), a potentially lethal respiratory illness caused by the coronavirus SARS-CoV-2, emerged in the end of 2019 and has since spread aggressively across the globe. A thorough understanding of the molecular mechanisms of cellular infection by coronaviruses is therefore of utmost importance. A critical stage in infection is the fusion between viral and host membranes. Here, we present a detailed investigation of the role of selected SARS-CoV-2 Spike fusion peptides, and the influence of calcium and cholesterol, in this fusion process. Structural information from specular neutron reflectometry and small angle neutron scattering, complemented by dynamics information from quasi-elastic and spin-echo neutron spectroscopy, revealed strikingly different functions encoded in the Spike fusion domain. Calcium drives the N-terminal of the Spike fusion domain to fully cross the host plasma membrane. Removing calcium, however, reorients the peptide back to the lipid leaflet closest to the virus, leading to significant changes in lipid fluidity and rigidity. In conjunction with other regions of the fusion domain, which are also positioned to bridge and dehydrate viral and host membranes, the molecular events leading to cell entry by SARS-CoV-2 are proposed.

Resistant bacteria are emerging as a critical problem in the treatment of bacterial infections by neutralizing antibiotic activity. The development of new traditional mechanisms of antibiotics is not the optimal solution. A more reasonable approach may be to use relatively safe, plant-based compounds in combination with conventional antibiotics in an effort to increase their efficacy or restore their activity against resistant bacteria. We present our study of mixing Ricini Semen extract, or its constituent fatty acids, with oxacillin and testing the effects of each on the growth of methicillin-resistant Staphylococcus aureus. Changes in the cell membrane fluidity of methicillin-resistant S. aureus were found to be a major component of the mechanism of synergistic antibiotic activity of Ricini Semen extract and its constituent fatty acids. In our model, changes in cellular membrane fluidity disrupted the normal function of bacterial signaling membrane proteins BlaR1 and MecR1, which are known to detect oxacillin, and resulted in the incomplete expression of penicillin-binding proteins 2a and β-lactamase. Utilizing the mechanism presented in this study presents the possibility of developing a method for treating antibiotic-resistant bacteria using traditional antibiotics with plant-based compounds.

Staphylococcus aureus is an opportunistic pathogen and the major causative agent of life-threatening hospital- and community-acquired infections. A combination of antibiotics could be an opportunity to address the widespread emergence of antibiotic-resistant strains, including Methicillin-Resistant S. aureus (MRSA). We here investigated the potential synergy between ampicillin and plant-derived antibiotics (pentacyclic triterpenes, ursolic acid (UA) and oleanolic acid (OA)) towards MRSA (ATCC33591 and COL) and the mechanisms involved. We calculated the Fractional Inhibitory Concentration Index (FICI) and demonstrated synergy. We monitored fluorescence of Bodipy-TR-Cadaverin, propidium iodide and membrane potential-sensitive probe for determining the ability of UA and OA to bind to lipoteichoic acids (LTA), and to induce membrane permeabilization and depolarization, respectively. Both pentacyclic triterpenes were able to bind to LTA and to induce membrane permeabilization and depolarization in a dose-dependent fashion. These effects were not accompanied by significant changes in cellular concentration of pentacyclic triterpenes and/or ampicillin, suggesting an effect mediated through lipid membranes. We therefore focused on membranous effects induced by UA and OA, and we investigated on models of membranes, the role of specific lipids including phosphatidylglycerol and cardiolipin. The effect induced on membrane fluidity, permeability and ability to fuse were studied by determining changes in fluorescence anisotropy of DPH/generalized polarization of Laurdan, calcein release from liposomes, fluorescence dequenching of octadecyl-rhodamine B and liposome-size, respectively. Both UA and OA showed a dose-dependent effect with membrane rigidification, increase of membrane permeabilization and fusion. Except for the effect on membrane fluidity, the effect of UA was consistently higher compared with that obtained with OA, suggesting the role of methyl group position. All together the data demonstrated the potential role of compounds acting on lipid membranes for enhancing the activity of other antibiotics, like ampicillin and inducing synergy. Such combinations offer an opportunity to explore a larger antibiotic chemical space.

Liposomes that achieve a heterogeneous and spatially organized surface through phase separation have been recognized to be a promising platform for delivery purposes. However, their design and optimization through experimentation can be expensive and time-consuming. To assist with the design and reduce the associated cost, we propose a computational platform for modeling membrane coarsening dynamics based on the principles of continuum mechanics and thermodynamics. This model couples phase separation to lateral flow and accounts for different membrane fluidity within the different phases, which is known to affect the coarsening dynamics on lipid membranes. The simulation results are in agreement with the experimental data in terms of liquid ordered domains area fraction, total domains perimeter over time and total number of domains over time for two different membrane compositions (DOPC:DPPC with a 1:1 molar ratio with 15% Chol and DOPC:DPPC with a 1:2 molar ratio with 25% Chol) that yield opposite and nearly inverse phase behavior. This quantitative validation shows that the developed platform can be a valuable tool in complementing experimental practice. Keywords: Multicomponent Membranes; Membrane fluidity; Membrane Phase Separation; Computational Modeling; Fluorescence Microscopy; Liposomes, 21 pages, 13 figures

PlsX plays a central role in the coordination of fatty acid and phospholipid biosynthesis in Gram-positive bacteria. PlsX is a peripheral membrane acyltransferase that catalyzes the conversion of acyl-ACP to acyl-phosphate, which is in turn utilized by the polytopic membrane acyltransferase PlsY on the pathway of bacterial phospholipid biosynthesis. We have recently studied the interaction between PlsX and membrane phospholipids in vivo and in vitro, and observed that membrane association is necessary for the efficient transfer of acyl-phosphate to PlsY. However, understanding the molecular basis of such a channeling mechanism remains a major challenge. Here, we disentangle the binding and insertion events of the enzyme to the membrane, and the subsequent catalysis. We show that PlsX membrane binding is a process mostly mediated by phospholipid charge, whereas fatty acid saturation and membrane fluidity remarkably influence the membrane insertion step. Strikingly, the PlsX(L254E) mutant, whose biological functionality was severely compromised in vivo but remains catalytically active in vitro, was able to superficially bind to phospholipid vesicles, nevertheless, it loses the insertion capacity, strongly supporting the importance of membrane insertion in acyl-phosphate delivery. We propose a mechanism in which membrane fluidity governs the insertion of PlsX and thus regulates the biosynthesis of phospholipids in Gram-positive bacteria. This model may be operational in other peripheral membrane proteins with an unprecedented impact in drug discovery/development strategies.

Photolithotrophic sulfur bacteria are involved in biota functioning and have a biotechnological potential for bioremediation of contaminated environment, but the mechanisms of xenobiotics, in particular of heavy metal ions damaging action and the pathways of photolithotrophic bacteria adaptation under these conditions have not been established. In this work, the biochemical indicators of green photosynthetic bacteria Chlorobium limicola response to Cu ions were studied. C. limicola cells were incubated during one hour in buffer containing copper (II) sulfate in 0.05–0.5 mM concentrations and grown for 8 days in GSB medium. The content of Cu2+ in cells was estimated by atomic absorption spectroscopy. The activity of enzymes of antioxidant defense, photosynthetic pigments and glutathione content, indexes of lipids unsaturation and membrane viscosity as markers of membrane fluidity were estimated. It was shown that the response of green photosynthetic bacteria C. limicola to Cu2+ action varied depending on cations concentration. Under the influence of metal salt at 0.05 mM concentration, the activity of antioxidant enzymes, GSH/GSSG ratio, the content of photosynthetic pigments and membrane fluidity indexes were higher as compared with control. Under the increase of copper (II) sulfate concentration to 0.25 mM, the activity of antioxidant enzymes was lower compared to the response of the cells under the influence of 0.05 mM copper (II) and the GSSG content was increased. Under the influence of 0.5 mM copper (II) the indexes of membrane fluidity did not differ from the control, but superoxide dismutase and peroxidase activity inhibition and the further decrease of GSH/GSSG ratio were observed followed by the highest Cu2+ cations accumulation in cells and significant decrease of bacteria biomass growth.

In this review some aspects of investigation of vertebrate brain lipids made under the guidance of academician E.M.Kreps and then by his collaborators are characterized using as an example the studies of brain gangliosides. In brain gangliosides and individual phospholipids of cold-water stenothermal teleost fishes higher content of polyenoic and monoenoic fatty acids was revealed than in the brain gangliosides and individual phospholipids of warm-water stenothermal teleosts. The changes in fatty acid composition of lipids during adaptation of fishes to living in cold water and at great water depth are directed to the maintenance of the optimal degree of brain cell membrane fluidity and microheterogeneity. The results of cluster analysis of the data on composition and structure of carbohydrate component of brain gangliosides of various ectothermic vertebrates were used to create a dendrogram. This dendrogram was found to correspond appreciably to the tree of classical taxonomy of vertebrates. The changes in molecular organization of brain gangliosides in the course of evolution of vertebrates are suggested to contribute to differentiation of brain and complication of its functions in phylogenesis. The main mammalian brain gangliosides (GM1, GD1a, GD1b и GT1b) protect nerve cells against the action of excitatory amino acids, hydrogen peroxide, amyloid β-peptide and other toxins, their protective effect depending on activation of Trk receptor tyrosine kinase and downstream protein kinases (Akt, ERK1/2, protein kinase C). Another mechanism of protection is used by GM1 and GD1a gangiosides against bacterial lipopolysaccharide (LPS)-induced toxicity. It appears to be due to changes of lipid composition of plasma membrane rafts of nerve cells due to exogenous ganglioside incorporation, which prevents LPS receptors TLR4 translocation to them. Using Morris water test intranasal administration of gangliosides was shown to prevent the impairment of spatial memory in rats with 2-nd type diabetes mellitus. Intranasal administration of gangliosides was used for the first time, its high efficiency was shown.

Background: DHA can regulate various physiological functions of cells. Our group has clarified the immunomodulatory activity and molecular mechanism of DHA on RAW264.7 cells. However, the effect of DHA on the membrane fatty acid environment and the activation of signaling pathways on the cell membrane is still not clear. Methods: In this study, we evaluated the fluidity, the potential and the fatty acid, phospholipid and protein composition of the RAW264.7 cell membrane by DHA treatment. Results: The fluidity of the RAW264.7 cell membrane was increased by DHA treatment. The results of membrane potential analysis suggested that DHA (2.4 μM) significantly reduced the surface potential of the cell membrane, which might influence the fluidity of cell membranes. In addition, the fatty acids and phospholipids were measured and the results indicated that DHA treatment (2.4 μM) altered the lipid environment and the composition of phospholipids on the RAW264.7 cell membrane. Then the LC-MS/MS-based label free quantitative proteomics approach was applied to identify a total of 86 differential proteins in the 2.4 μM DHA and control groups (>2.0-fold change or

Ubiquinone 8 (coenzyme Q8 or Q8) mediates electron transfer within the aerobic respiratory chain, mitigates oxidative stress, and contributes to gene expression inEscherichia coli. In addition, Q8 was proposed to confer bacterial osmotolerance by accumulating during growth at high osmotic pressure and altering membrane stability. The osmolyte trehalose and membrane lipid cardiolipin accumulate inE. colicells cultivated at high osmotic pressure. Here, Q8 deficiency impairedE. coligrowth at low osmotic pressure and rendered growth osmotically sensitive. The Q8 deficiency impeded cellular O2uptake and also inhibited the activities of two proton symporters, the osmosensing transporter ProP and the lactose transporter LacY. Q8 supplementation decreased membrane fluidity in liposomes, but did not affect ProP activity in proteoliposomes, which is respiration-independent. Liposomes and proteoliposomes prepared withE. colilipids were used for these experiments. Similar oxygen uptake rates were observed for bacteria cultivated at low and high osmotic pressures. In contrast, respiration was dramatically inhibited when bacteria grown at the same low osmotic pressure were shifted to high osmotic pressure. Thus, respiration was restored during prolonged growth ofE. coliat high osmotic pressure. Of note, bacteria cultivated at low and high osmotic pressures had similar Q8 concentrations. The protection of respiration was neither diminished by cardiolipin deficiency nor conferred by trehalose overproduction during growth at low osmotic pressure, but rather might be achieved by Q8-independent respiratory chain remodeling. We conclude that osmotolerance is conferred through Q8-independent protection of respiration, not by altering physical properties of the membrane.

To unravel the underlying principles of membrane adaptation in small systems like bacterial cells, robust approaches to characterize membrane fluidity are needed. Currently available relevant methods require advanced instrumentation and are not suitable for high‐throughput settings needed to elucidate the biochemical pathways involved in adaptation. We developed a fast, robust, and financially accessible quantitative method to measure the microviscosity of lipid membranes in bulk suspension using a commercially available plate reader. Our approach, which is suitable for high‐throughput screening, is based on the simultaneous measurements of absorbance and fluorescence emission of a viscosity‐sensitive fluorescent dye, 9‐(2,2‐dicyanovinyl)julolidine (DCVJ), incorporated into a lipid membrane. We validated our method using artificial membranes with various lipid compositions over a range of temperatures and observed values that were in good agreement with previously published results. Using our approach, we were able to detect a lipid phase transition in the ruminant pathogen Mycoplasma mycoides., Membrane adaptation: We developed a robust and financially accessible quantitative method to measure the microviscosity of lipid membranes that is suitable for high‐throughput screening. The approach is based on the simultaneous measurements of absorbance and fluorescence emission of a fluorescent dye and allows monitoring of the temperature dependence of viscosity in bacterial membranes.

Naja spp. venom is a natural source of active compounds with therapeutic application potential. Phospholipase A2 (PLA2) is abundant in the venom of Naja spp. and can perform neurotoxicity, cytotoxicity, cardiotoxicity, and hematological disorders. The PLA2s from Naja spp. venoms are Asp 49 isoenzymes with the exception of PLA2 Cys 49 from Naja sagittifera. When looking at the functional aspects, the neurotoxicity occurs by PLA2 called β-toxins that have affinity for phosphatidylcholine in nerve endings and synaptosomes membranes, and by α-toxins that block the nicotinic acetylcholine receptors in the neuromuscular junctions. In addition, these neurotoxins may inhibit K+ and Ca++ channels or even interfere with the Na+/K+/ATPase enzyme. The disturbance in the membrane fluidity also results in inhibition of the release of acetylcholine. The PLA2 can act as anticoagulants or procoagulant. The cytotoxicity exerted by PLA2s result from changes in the cardiomyocyte membranes, triggering cardiac failure and hemolysis. The antibacterial activity, however, is the result of alterations that decrease the stability of the lipid bilayer. Thus, the understanding of the structural and functional aspects of PLA2s can contribute to studies on the toxic and therapeutic mechanisms involved in the envenomation by Naja spp. and in the treatment of pathologies.

Alkaliphilic Bacillus pseudofirmus OF4, which has a broad pH growth range of 7.5 to above 10.5, is yellow-pigmented due to carotenoids. Carotenoids contribute to membrane rigidity and can alleviate cellular oxidative stress. This study was undertaken to gain insight into the roles carotenoids play in alkaliphile physiology. Carotenoid content was high in stationary phase and in cells grown nonfermentatively at pH 10.5 A colourless mutant was isolated by the in-frame deletion of a key carotenogenic gene, crtM. In cells grown to stationary phase in a pH 10.5 medium with a suboptimal concentration of Na+, the ∆crtM strain exhibited lower resistance to paraquat and hydrogen peroxide. Preincubation of the mutant in a nutrient-free pH 10.5 buffer revealed a pronounced sensitivity to hydrogen peroxide in growth at pH 7.5. In growth curves in media with optimal or suboptimal nutrient concentrations conducted at 37°, the mutant grew identically to the wild-type at pH 7.5 but its lag time was longer than the wild-type at pH 10.5 and growth was slower when the carbon source, malate, was limiting. When the temperature of the growth curves was lowered to 25°, the mutant no longer had a pH 10.5 phenotype, implicating the effect of carotenoids on membrane rigidity for the pH 10.5 growth phenotype. These results suggest that carotenoids in B. pseudofirmus OF4 play a role in managing oxidative stress when cells are adapting to other stressful conditions such as nutrient limitation while also helping to maintain membrane fluidity/rigidity balance for membrane-linked functions.