Organization Of Membrane Domains Research Articles

Unraveling the link between neuropathy target esterase NTE/SWS, lysosomal storage diseases, inflammation, abnormal fatty acid metabolism, and leaky brain barrier.

Mutations in Drosophila Swiss cheese (SWS) gene or its vertebrate orthologue neuropathy target esterase (NTE) lead to progressive neuronal degeneration in flies and humans. Despite its enzymatic function as a phospholipase is well established, the molecular mechanism responsible for maintaining nervous system integrity remains unclear. In this study, we found that NTE/SWS is present in surface glia that forms the blood-brain barrier (BBB) and that NTE/SWS is important to maintain its structure and permeability. Importantly, BBB glia-specific expression of Drosophila NTE/SWS or human NTE in the sws mutant background fully rescues surface glial organization and partially restores BBB integrity, suggesting a conserved function of NTE/SWS. Interestingly, sws mutant glia showed abnormal organization of plasma membrane domains and tight junction rafts accompanied by the accumulation of lipid droplets, lysosomes, and multilamellar bodies. Since the observed cellular phenotypes closely resemble the characteristics described in a group of metabolic disorders known as lysosomal storage diseases (LSDs), our data established a novel connection between NTE/SWS and these conditions. We found that mutants with defective BBB exhibit elevated levels of fatty acids, which are precursors of eicosanoids and are involved in the inflammatory response. Also, as a consequence of a permeable BBB, several innate immunity factors are upregulated in an age-dependent manner, while BBB glia-specific expression of NTE/SWS normalizes inflammatory response. Treatment with anti-inflammatory agents prevents the abnormal architecture of the BBB, suggesting that inflammation contributes to the maintenance of a healthy brain barrier. Considering the link between a malfunctioning BBB and various neurodegenerative diseases, gaining a deeper understanding of the molecular mechanisms causing inflammation due to a defective BBB could help to promote the use of anti-inflammatory therapies for age-related neurodegeneration.

Identification of ACBD3 as a new molecular biomarker in pan-cancers through bioinformatic analysis: a preclinical study

BackgroundAcyl-CoA-binding domain-containing 3 (ACBD3) is a multifunctional protein, that plays essential roles in cellular signaling and membrane domain organization. Although the precise roles of ACBD3 in various cancers remain unclear. Thus, we aimed to determine the diverse roles of ACBD3 in pan-cancers.MethodsRelevant clinical and RNA-sequencing data for normal tissues and 33 tumors from The Cancer Genome Atlas (TCGA) database, the Human Protein Atlas, and other databases were applied to investigate ACBD3 expression in various cancers. ACBD3-binding and ACBD3-related target genes were obtained from the STRING and GEPIA2 databases. The possible functions of ACBD3-binding genes were explored using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. We also applied the diagnostic value and survival prognosis analysis of ACBD3 in pan-cancers using R language. The mutational features of ACBD3 in various TCGA cancers were obtained from the cBioPortal database.ResultsWhen compared with normal tissues, ACBD3 expression was statistically upregulated in eleven cancers and downregulated in three cancers. ACBD3 expression was remarkably different among various pathological stages of tumors, immune and molecular subtypes of cancers, cancer phosphorylation levels, and immune cell infiltration. The survival of four tumors was correlated with the expression level of ACBD3, including pancreatic adenocarcinoma, adrenocortical carcinoma, sarcoma, and glioma. The high accuracy in diagnosing multiple tumors and its correlation with prognosis indicated that ACBD3 may be a potential biomarker of pan-cancers.ConclusionAccording to our pan-cancer analysis, ACBD3 may serve as a remarkable prognostic and diagnostic biomarker of pan-cancers as well as contribute to tumor development. ACBD3 may also provide new directions for cancer treatment targets in the future.

Cyclic lipopeptides as membrane fusion inhibitors against SARS-CoV-2: New tricks for old dogs

With the resurgence of the coronavirus pandemic, the repositioning of FDA-approved drugs against coronovirus and finding alternative strategies for antiviral therapy are both important. We previously identified the viral lipid envelope as a potential target for the prevention and treatment of SARS-CoV-2 infection with plant alkaloids (Shekunov et al., 2021). Here, we investigated the effects of eleven cyclic lipopeptides (CLPs), including well-known antifungal and antibacterial compounds, on the liposome fusion triggered by calcium, polyethylene glycol 8000, and a fragment of SARS-CoV-2 fusion peptide (816–827) by calcein release assays. Differential scanning microcalorimetry of the gel-to-liquid-crystalline and lamellar-to-inverted hexagonal phase transitions and confocal fluorescence microscopy demonstrated the relation of the fusion inhibitory effects of CLPs to alterations in lipid packing, membrane curvature stress and domain organization. The antiviral effects of CLPs were evaluated in an in vitro Vero-based cell model, and aculeacin A, anidulafugin, iturin A, and mycosubtilin attenuated the cytopathogenicity of SARS-CoV-2 without specific toxicity.

Optical Manipulation of Gb3 Enriched Lipid Domains: Impact of Isomerization on Gb3 -Shiga Toxin B Interaction.

The plasma membrane is a complex assembly of proteins and lipids that can self-assemble in submicroscopic domains commonly termed "lipid rafts", which are implicated in membrane signaling and trafficking. Recently, photo-sensitive lipids were introduced to study membrane domain organization, and photo-isomerization was shown to trigger the mixing and de-mixing of liquid-ordered (lo ) domains in artificial phase-separated membranes. Here, we synthesized globotriaosylceramide (Gb3 ) glycosphingolipids that harbor an azobenzene moiety at different positions of the fatty acid to investigate light-induced membrane domain reorganization, and that serve as specific receptors for the protein Shiga toxin (STx). Using phase-separated supported lipid bilayers on mica surfaces doped with four different photo-Gb3 molecules, we found by fluorescence microscopy and atomic force microscopy that liquid disordered (ld ) domains were formed within lo domains upon trans-cis photo-isomerization. The fraction and size of these ld domains were largest for Gb3 molecules with the azobenzene group at the end of the fatty acid. We further investigated the impact of domain reorganization on the interaction of the B-subunits of STx with the photo-Gb3 . Fluorescence and atomic force micrographs clearly demonstrated that STxB binds to the lo phase if Gb3 is in the trans-configuration, whereas two STxB populations are formed if the photo-Gb3 is switched to the cis-configuration highlighting the idea of manipulating lipid-protein interactions with a light stimulus.

Ankyrin-R Links Kv3.3 to the Spectrin Cytoskeleton and Is Required for Purkinje Neuron Survival.

Ankyrin scaffolding proteins are critical for membrane domain organization and protein stabilization in many different cell types including neurons. In the cerebellum, Ankyrin-R (AnkR) is highly enriched in Purkinje neurons, granule cells, and in the cerebellar nuclei (CN). Using male and female mice with a floxed allele for Ank1 in combination with Nestin-Cre and Pcp2-Cre mice, we found that ablation of AnkR from Purkinje neurons caused ataxia, regional and progressive neurodegeneration, and altered cerebellar output. We show that AnkR interacts with the cytoskeletal protein β3 spectrin and the potassium channel Kv3.3. Loss of AnkR reduced somatic membrane levels of β3 spectrin and Kv3.3 in Purkinje neurons. Thus, AnkR links Kv3.3 channels to the β3 spectrin-based cytoskeleton. Our results may help explain why mutations in β3 spectrin and Kv3.3 both cause spinocerebellar ataxia.SIGNIFICANCE STATEMENT Ankyrin scaffolding proteins localize and stabilize ion channels in the membrane by linking them to the spectrin-based cytoskeleton. Here, we show that Ankyrin-R (AnkR) links Kv3.3 K+ channels to the β3 spectrin-based cytoskeleton in Purkinje neurons. Loss of AnkR causes Purkinje neuron degeneration, altered cerebellar physiology, and ataxia, which is consistent with mutations in Kv3.3 and β3 spectrin causing spinocerebellar ataxia.

Biosynthesis and Functions of Very-Long-Chain Fatty Acids in the Responses of Plants to Abiotic and Biotic Stresses.

Very-long-chain fatty acids (i.e., fatty acids with more than 18 carbon atoms; VLCFA) are important molecules that play crucial physiological and structural roles in plants. VLCFA are specifically present in several membrane lipids and essential for membrane homeostasis. Their specific accumulation in the sphingolipids of the plasma membrane outer leaflet is of primordial importance for its correct functioning in intercellular communication. VLCFA are found in phospholipids, notably in phosphatidylserine and phosphatidylethanolamine, where they could play a role in membrane domain organization and interleaflet coupling. In epidermal cells, VLCFA are precursors of the cuticular waxes of the plant cuticle, which are of primary importance for many interactions of the plant with its surrounding environment. VLCFA are also major components of the root suberin barrier, which has been shown to be fundamental for nutrient homeostasis and plant adaptation to adverse conditions. Finally, some plants store VLCFA in the triacylglycerols of their seeds so that they later play a pivotal role in seed germination. In this review, taking advantage of the many studies conducted using Arabidopsis thaliana as a model, we present our current knowledge on the biosynthesis and regulation of VLCFA in plants, and on the various functions that VLCFA and their derivatives play in the interactions of plants with their abiotic and biotic environment.

Differential impact of synthetic antitumor lipid drugs on the membrane organization of phosphatidic acid and diacylglycerol monolayers

Anti-tumour lipids are synthetic analogues of lysophosphatidylcholine. These drugs are both cytotoxic and cytostatic, and more interestingly, exert these effects preferentially in tumour cells. While the exact mechanism of action isn’t fully elucidated, these drugs appear to preferentially partition into rigid lipid domains in cell membranes. Upon insertion, the compounds alter membrane domain organization, disrupt normal signal transduction, and cause cell death. Recently, it has been reported that these drugs induce accumulation of diacylglycerol in yeast cells which in turn sensitizes cells to the drugs. Conversely, phosphatidic acid accumulation appears to protect cells against the drugs. In the current work, the aim was to compare the biophysical effects of the drugs edelfosine, miltefosine and perifosine on monolayers of dimyristoyl phosphatidic acid, dimyristoyl glycerol and an equimolar mixture, to understand how these lipids modulate the mode of action. Surface pressure – area isotherms, compression moduli and Brewster angle microscopy were used to compare drug effects on lipid packing, monolayer compressibility and lateral domain organization of these films. Results suggest that edelfosine and miltefosine have stabilizing effects on all of the monolayers, while perifosine destabilizes dimyristoyl glycerol and the equimolar mixture. Additionally, all three drugs change the morphology of the domains observed. Based on these results the stabilization of diacylgylcerol by edelfosine and miltefosine may contribute to the mode of action as diacylglycerol is a known disruptor of bilayers. Perifosine however does not stabilize diacylglycerol, and therefore cell death may occur through a more direct inhibition of specific signal transduction. These results suggest that perifosine may illicit cytotoxicity through a different mechanism compared to the other antitumor lipid drugs.

Functional lipid pairs as building blocks of phase-separated membranes

Biological membranes exhibit a great deal of compositional and phase heterogeneity due to hundreds of chemically distinct components. As a result, phase separation processes in cell membranes are extremely difficult to study, especially at the molecular level. It is currently believed that the lateral membrane heterogeneity and the formation of domains, or rafts, are driven by lipid-lipid and lipid-protein interactions. Nevertheless, the underlying mechanisms regulating membrane heterogeneity remain poorly understood. In the present work, we combine inelastic X-ray scattering with molecular dynamics simulations to provide direct evidence for the existence of strongly coupled transient lipid pairs. These lipid pairs manifest themselves experimentally through optical vibrational (a.k.a. phononic) modes observed in binary (1,2-dipalmitoyl-sn-glycero-3-phosphocholine [DPPC]-cholesterol) and ternary (DPPC-1,2-dioleoyl-sn-glycero-3-phosphocholine/1-palmitoyl-2-oleoyl-glycero-3-phosphocholine [DOPC/POPC]-cholesterol) systems. The existence of a phononic gap in these vibrational modes is a direct result of the finite size of patches formed by these lipid pairs. The observation of lipid pairs provides a spatial (subnanometer) and temporal (subnanosecond) window into the lipid-lipid interactions in complex mixtures of saturated/unsaturated lipids and cholesterol. Our findings represent a step toward understanding the lateral organization and dynamics of membrane domains using a well-validated probe with a high spatial and temporal resolution.

Pharmacological Overview of the BGP-15 Chemical Agent as a New Drug Candidate for the Treatment of Symptoms of Metabolic Syndrome.

BGP-15 is a new insulin sensitizer drug candidate, which was developed by Hungarian researchers. In recent years, numerous research groups have studied its beneficial effects. It is effective in the treatment of insulin resistance and it has protective effects in Duchenne muscular dystrophy, diastolic dysfunction, tachycardia, heart failure, and atrial fibrillation, and it can alleviate cardiotoxicity. BGP-15 exhibits chemoprotective properties in different cytostatic therapies, and has also proven to be photoprotective. It can additionally have advantageous effects in mitochondrial-stress-related diseases. Although the precise mechanism of the effect is still unknown to us, we know that the molecule is a PARP inhibitor, chaperone co-inducer, reduces ROS production, and is able to remodel the organization of cholesterol-rich membrane domains. In the following review, our aim was to summarize the investigated molecular mechanisms and pharmacological effects of this potential API. The main objective was to present the wide pharmacological potentials of this chemical agent.

Annexin A2 gene interacting with viral matrix protein to promote bovine ephemeral fever virus release.

Bovine ephemeral fever virus (BEFV) causes bovine ephemeral fever, which can produce considerable economic damage to the cattle industry. However, there is limited experimental evidence regarding the underlying mechanisms of BEFV. Annexin A2 (AnxA2) is a calcium and lipid-conjugated protein that binds phospholipids and the cytoskeleton in a Ca2+-dependent manner, and it participates in various cellular functions, including vesicular trafficking, organization of membrane domains, and virus proliferation. The role of the AnxA2 gene during virus infection has not yet been reported. In this study, we observed that AnxA2 gene expression was up-regulated in BHK-21 cells infected with the virus. Additionally, overexpression of the AnxA2 gene promoted the release of mature virus particles, whereas BEFV replication was remarkably inhibited after reducing AnxA2 gene expression by using the small interfering RNA (siRNA). For viral proteins, overexpression of the Matrix (M) gene promotes the release of mature virus particles. Moreover, the AnxA2 protein interaction with the M protein of BEFV was confirmed by GST pull-down and co-immunoprecipitation assays. Experimental results indicate that the C-terminal domain (268–334 aa) of AxnA2 contributes to this interaction. An additional mechanistic study showed that AnxA2 protein interacts with M protein and mediates the localization of the M protein at the plasma membrane. Furthermore, the absence of the AnxA2-V domain could attenuate the effect of AnxA2 on BEFV replication. These findings can contribute to elucidating the regulation of BEFV replication and may have implications for antiviral strategy development.

Optimization of 2D and 3D cell culture to study membrane organization with STED microscopy

Epithelia cells assemble into sheets that compartmentalize organs and generate tissue barriers. This is achieved by forming polarized membrane domains, which are connected by junctional complexes. While much is known about the organization of the basal membrane due to its easy accessibility by high and super-resolution microscopy, the apical and lateral membrane domains remain poorly characterized. Here we describe our methods to study the molecular organization of apical and lateral membrane domains by combining 2D and 3D epithelial cell culture with super-resolution STED microscopy. We show that inverted cell monolayers enable live cell imaging of the apical membrane with a resolution sufficient to resolve the densely packed micro-villi of human enterocytes. Furthermore, 3D cell culture enables us to resolve adhesion complexes in the lateral domain of kidney derived cells. We envision that these methods will help to reveal the supra-molecular structure of lateral and apical membrane domains in epithelial cells.

Arabidopsis Trichome Contains Two Plasma Membrane Domains with Different Lipid Compositions Which Attract Distinct EXO70 Subunits

Plasma membrane (PM) lipid composition and domain organization are modulated by polarized exocytosis. Conversely, targeting of secretory vesicles at specific domains in the PM is carried out by exocyst complexes, which contain EXO70 subunits that play a significant role in the final recognition of the target membrane. As we have shown previously, a mature Arabidopsis trichome contains a basal domain with a thin cell wall and an apical domain with a thick secondary cell wall, which is developed in an EXO70H4-dependent manner. These domains are separated by a cell wall structure named the Ortmannian ring. Using phospholipid markers, we demonstrate that there are two distinct PM domains corresponding to these cell wall domains. The apical domain is enriched in phosphatidic acid (PA) and phosphatidylserine, with an undetectable amount of phosphatidylinositol 4,5-bisphosphate (PIP2), whereas the basal domain is PIP2-rich. While the apical domain recruits EXO70H4, the basal domain recruits EXO70A1, which corresponds to the lipid-binding capacities of these two paralogs. Loss of EXO70H4 results in a loss of the Ortmannian ring border and decreased apical PA accumulation, which causes the PA and PIP2 domains to merge together. Using transmission electron microscopy, we describe these accumulations as a unique anatomical feature of the apical cell wall—radially distributed rod-shaped membranous pockets, where both EXO70H4 and lipid markers are immobilized.

Acyl-CoA-Binding Domain-Containing 3 (ACBD3; PAP7; GCP60): A Multi-Functional Membrane Domain Organizer.

Acyl-CoA-binding domain-containing 3 (ACBD3) is a multi-functional scaffolding protein, which has been associated with a diverse array of cellular functions, including steroidogenesis, embryogenesis, neurogenesis, Huntington’s disease (HD), membrane trafficking, and viral/bacterial proliferation in infected host cells. In this review, we aim to give a timely overview of recent findings on this protein, including its emerging role in membrane domain organization at the Golgi and the mitochondria. We hope that this review provides readers with useful insights on how ACBD3 may contribute to membrane domain organization along the secretory pathway and on the cytoplasmic surface of intracellular organelles, which influence many important physiological and pathophysiological processes in mammalian cells.

Mechanical Stretch Redefines Membrane Gαq-Calcium Signaling Complexes.

Muscle cells are routinely subjected to mechanical stretch but the impact of stretch on the organization of membrane domains is unknown. In this study, we characterize the effect of stretch on GPCR-Gαq protein signaling. Activation of this pathway leads to an increase in intracellular calcium. In muscle cells, GPCR-Gαq signals are enhanced when these proteins are localized in caveolae membrane domains whose curved structure can flatten with stretch. When we statically stretch rat aortic smooth muscle A10 cells by 1-5%, cellular calcium appears unperturbed as indicated by a calcium indicator. However, when we activate the bradykinin type 2 receptor (B2R)/Gαq pathway, we observe a loss in calcium that appears to be mediated through perturbations in calcium-activated stretch receptors. In contrast, if we apply oscillating stretch, calcium levels are enhanced. We tested whether the observed changes in B2R-Gαq calcium signals were caused by stretch-induced disruption of caveolae using a combination of silencing RNA technology and growth conditions. We find that stretch changes the ability of monoclonal caveolin antibodies to bind caveolae indicating a change in configuration of the domains. This change is seen by the inability of cells to survive stretch cycles when the level of caveolae is significantly reduced. Our studies show that the effect of calcium signals by mechanical stretch is mediated by the type of stretch and the amount of caveolae.

Lipids and Membrane Microdomains: The Glycerolipid and Alkylphosphocholine Class of Cancer Chemotherapeutic Drugs.

Synthetic antitumor lipids are metabolically stable lysophosphatidylcholine derivatives, encompassing a class of non-mutagenic drugs that selectively target cancerous cells. In this chapter we review the literature as relates to the clinical efficacy of these antitumor lipid drugs and how our understanding of their mode of action has evolved alongside key advances in our knowledge of membrane structure, organization, and function. First, the history of the development of this class of drugs is described, providing a summary of clinical outcomes of key members including edelfosine, miltefosine, perifosine, erufosine, and erucylphosphocholine. A detailed description of the biophysical properties of these drugs and specific drug-lipid interactions which may contribute to the selectivity of the antitumor lipids for cancer cells follows. An updated model on the mode of action of these lipid drugs as membrane disorganizing agents is presented. Membrane domain organization as opposed to targeting specific proteins on membranes is discussed. By altering membranes, these antitumor lipids inhibit many survival pathways while activating pro-apoptotic signals leading to cell demise.

Annexin A2 in Virus Infection.

Viral life cycles consist of three main phases: (1) attachment and entry, (2) genome replication and expression, and (3) assembly, maturation, and egress. Each of these steps is intrinsically reliant on host cell factors and processes including cellular receptors, genetic replication machinery, endocytosis and exocytosis, and protein expression. Annexin A2 (AnxA2) is a membrane-associated protein with a wide range of intracellular functions and a recurrent host factor in a variety of viral infections. Spatially, AnxA2 is found in the nucleus and cytoplasm, vesicle-bound, and on the inner and outer leaflet of the plasma membrane. Structurally, AnxA2 exists as a monomer or in complex with S100A10 to form the AnxA2/S100A10 heterotetramer (A2t). Both AnxA2 and A2t have been implicated in a vast array of cellular functions such as endocytosis, exocytosis, membrane domain organization, and translational regulation through RNA binding. Accordingly, many discoveries have been made involving AnxA2 in viral pathogenesis, however, the reported work addressing AnxA2 in virology is highly compartmentalized. Therefore, the purpose of this mini review is to provide information regarding the role of AnxA2 in the lifecycle of multiple epithelial cell-targeting viruses to highlight recurrent themes, identify discrepancies, and reveal potential avenues for future research.

Annexin A2 Mediates the Localization of Measles Virus Matrix Protein at the Plasma Membrane.

Annexins are a family of structurally related proteins that bind negatively charged membrane phospholipids in a Ca2+-dependent manner. Annexin A2 (AnxA2), a member of this family, has been implicated in a variety of cellular functions, including the organization of membrane domains, vesicular trafficking, and cell-cell adhesion. AnxA2 generally forms a heterotetrameric complex with a small Ca2+-binding protein, S100A10. Measles virus (MV), a member of the family Paramyxoviridae, is an enveloped virus with a nonsegmented negative-strand RNA genome. Knockdown of AnxA2 greatly reduced MV growth in cells without affecting its entry and viral RNA production. In MV-infected, AnxA2 knockdown cells, the expression level of the matrix (M) protein, but not other viral proteins, was reduced compared with that in control cells, and the distribution of the M protein at the plasma membrane was decreased. The M protein lines the inner surface of the envelope and plays an important role in virus assembly by connecting the nucleocapsid to the envelope proteins. The M protein bound to AnxA2 independently of AnxA2's phosphorylation or its association with S100A10 and was colocalized with AnxA2 within cells. Truncation of the N-terminal 10 amino acid residues, but not the N-terminal 5 residues, compromised the ability of the M protein to interact with AnxA2 and localize at the plasma membrane. These results indicate that AnxA2 mediates the localization of the MV M protein at the plasma membrane by interacting with its N-terminal region (especially residues at positions 6 to 10), thereby aiding in MV assembly.IMPORTANCE MV is an important human pathogen, still claiming ∼100,000 lives per year despite the presence of effective vaccines, and it causes occasional outbreaks even in developed countries. Replication of viruses largely relies on the functions of host cells. Our study revealed that the reduction of the host protein annexin A2 compromises the replication of MV within the cell. Further studies demonstrated that annexin A2 interacts with the MV M protein and mediates the localization of the M protein at the plasma membrane where MV particles are formed. The M protein lines the inner surface of the MV envelope membrane and plays a role in MV particle formation. Our results provide useful information for the understanding of the MV replication process and potential development of antiviral agents.

Modeling of annexin A2-Membrane interactions by molecular dynamics simulations.

The annexins are a family of Ca2+-regulated phospholipid binding proteins that are involved in membrane domain organization and membrane trafficking. Although they are widely studied and crystal structures are available for several soluble annexins their mode of membrane association has never been studied at the molecular level. Here we obtained molecular information on the annexin-membrane interaction that could serve as paradigm for the peripheral membrane association of cytosolic proteins by Molecular Dynamics simulations. We analyzed systems containing the monomeric annexin A2 (AnxA2), a membrane with negatively charged phosphatidylserine (POPS) lipids as well as Ca2+ ions. On the atomic level we identify the AnxA2 orientations and the respective residues which display the strongest interaction with Ca2+ ions and the membrane. The simulation results fully agree with earlier experimental findings concerning the positioning of bound Ca2+ ions. Furthermore, we identify for the first time a significant interaction between lysine residues of the protein and POPS lipids that occurs independently of Ca2+ suggesting that AnxA2-membrane interactions can also occur in a low Ca2+ environment. Finally, by varying Ca2+ concentrations and lipid composition in our simulations we observe a calcium-induced negative curvature of the membrane as well as an AnxA2-induced lipid ordering.

Axonal Membranes and Their Domains: Assembly and Function of the Axon Initial Segment and Node of Ranvier.

Neurons are highly specialized cells of the nervous system that receive, process and transmit electrical signals critical for normal brain function. Here, we review the intricate organization of axonal membrane domains that facilitate rapid action potential conduction underlying communication between complex neuronal circuits. Two critical excitable domains of vertebrate axons are the axon initial segment (AIS) and the nodes of Ranvier, which are characterized by the high concentrations of voltage-gated ion channels, cell adhesion molecules and specialized cytoskeletal networks. The AIS is located at the proximal region of the axon and serves as the site of action potential initiation, while nodes of Ranvier, gaps between adjacent myelin sheaths, allow rapid propagation of the action potential through saltatory conduction. The AIS and nodes of Ranvier are assembled by ankyrins, spectrins and their associated binding partners through the clustering of membrane proteins and connection to the underlying cytoskeleton network. Although the AIS and nodes of Ranvier share similar protein composition, their mechanisms of assembly are strikingly different. Here we will cover the mechanisms of formation and maintenance of these axonal excitable membrane domains, specifically highlighting the similarities and differences between them. We will also discuss recent advances in super resolution fluorescence imaging which have elucidated the arrangement of the submembranous axonal cytoskeleton revealing a surprising structural organization necessary to maintain axonal organization and function. Finally, human mutations in axonal domain components have been associated with a growing number of neurological disorders including severe cognitive dysfunction, epilepsy, autism, neurodegenerative diseases and psychiatric disorders. Overall, this review highlights the assembly, maintenance and function of axonal excitable domains, particularly the AIS and nodes of Ranvier, and how abnormalities in these processes may contribute to disease.

Antimicrobial Peptides Targeting Gram-Positive Bacteria.

Antimicrobial peptides (AMPs) have remarkably different structures as well as biological activity profiles, whereupon most of these peptides are supposed to kill bacteria via membrane damage. In order to understand their molecular mechanism and target cell specificity for Gram-positive bacteria, it is essential to consider the architecture of their cell envelopes. Before AMPs can interact with the cytoplasmic membrane of Gram-positive bacteria, they have to traverse the cell wall composed of wall- and lipoteichoic acids and peptidoglycan. While interaction of AMPs with peptidoglycan might rather facilitate penetration, interaction with anionic teichoic acids may act as either a trap for AMPs or a ladder for a route to the cytoplasmic membrane. Interaction with the cytoplasmic membrane frequently leads to lipid segregation affecting membrane domain organization, which affects membrane permeability, inhibits cell division processes or leads to delocalization of essential peripheral membrane proteins. Further, precursors of cell wall components, especially the highly conserved lipid II, are directly targeted by AMPs. Thereby, the peptides do not inhibit peptidoglycan synthesis via binding to proteins like common antibiotics, but form a complex with the precursor molecule, which in addition can promote pore formation and membrane disruption. Thus, the multifaceted mode of actions will make AMPs superior to antibiotics that act only on one specific target.