Mechanism Of Vesicle Formation Research Articles

Formation of Vesicles from Amphiphilic Random Copolymers in Solution: A Dissipative Particle Dynamics Simulation Study

Five coarse-grained models were built for amphiphilic random copolymers. The self-assembly of amphiphilic random copolymers in selective solvent was investigated via dissipative particle dynamics simulations. The simulation results showed that the content of hydrophilic particles and the repulsive parameter between solvent and copolymer particles were two key factors of the vesicle formation. We report herein on how to control the self-assembled morphology evolution. The two mechanisms of vesicle formation from amphiphilic random copolymers are found through investigating the dynamic processes of vesicle formation, which is in accordance with the experiment and simulation results of amphiphilic block copolymer reported in the literature.

Extensive Vesicle Formation by a Monotopic Membrane Glycosyltransferase

Over-expression of the foreign monotopic glycosyltransferase MGS in Escherichia coli triggers the formation of a large amount of internal vesicles (J. Biol. Chem. 284:33904, 2009). This phenomenon seems to be directly related with the interactions between the protein and the inner membrane lipids. It has been shown that this massive vesiculation process is not induced by the enzymatic activity of the protein, but rather due to the structural and mechanical properties of MGS. In this work we study the specific interactions of MGS with the lipid membrane and we unveil the biophysical mechanisms that lead to the massive formation of internal vesicles in E. coli.We use both molecular dynamics (MD) simulations and analytical modeling to understand the molecular basis for this phenomenon and the relative importance of various potential driving forces that may induce membrane deformations, such as lipid-protein interactions and protein crowding. First we studied the specific interactions between a single MGS and the lipid membrane. By using MD simulations we show that the N-domain (of the double Rossmann fold) has a leading role in modulating the bonds with the lipid bilayer, and that the connection between MGS and lipidic membranes is modulated by both hydrophobic and electrostatic interactions. These conclusions are in perfect agreement with chemical experiments run in parallel.Furthermore, we are using coarse-grained MD simulations to study the effects of asymmetric crowding and determine how several MGS molecules bound to the same bilayer leaflet may induce membrane deformations and trigger the formation of internal vesicles. Both experiments and simulations allow us to gain a deeper insight into the mechanism of vesicle formation by MGS, and potentially by other monotopic proteins.

Biomimetic membrane control of block copolymer vesicles with tunable wall thickness

Our recent simulation study on mechanisms of vesicle formation in ABA triblock copolymer system has revealed the factors or conditions that lead to the transition from mechanism I (a bilayer membrane closing pathway) to mechanism II (a nucleation and growth pathway). The spontaneous vesicle formation involves many metastable states and the two mechanisms do not each lead to a unique intermediate structure, but have a transition period. In this paper, we use dissipative particle dynamics (DPD) simulations to further investigate the effect of vesicle size and hydrophobicity of the blocks on membrane characteristics, with the aim of understanding and controlling the structures of block copolymer vesicle and its formation in different scales, and to try to connect our simulation results to previous experimental studies. We have evaluated the thickness of the hydrophobic layer and observed two types of dependence on the vesicle size. Our results indicate that as the degree of hydrophobicity of the blocks increases, from a totally strong-behavior to a totally weak-behavior relationship, the transformation is observed in large sized vesicles first and then in small sized vesicles. Two characteristics, the chain compaction of the vesicles and the area densities of the inner corona, are thought to be important in controlling the membrane thickness, which are proposed to explain the size-dependent behaviors of bilayer thickness. Finally, the fraction of bridges Φb is also calculated. In our model, Φb increases with an increase in vesicle size for a given system and fixed interaction and decreases with an increase in the degree of hydrophobicity of the blocks.

The Endosomal Sorting Complex ESCRT-II Mediates the Assembly and Architecture of ESCRT-III Helices

The endosomal sorting complexes required for transport (ESCRTs) constitute hetero-oligomeric machines that mediate topologically similar membrane-sculpting processes, including cytokinesis, retroviral egress, and multivesicular body (MVB) biogenesis. Although ESCRT-III drives membrane remodeling that creates MVBs, its structure and the mechanism of vesicle formation are unclear. Using electron microscopy, we visualize an ESCRT-II:ESCRT-III supercomplex and propose how it mediates vesicle formation. We define conformational changes that activate ESCRT-III subunit Snf7 and show that it assembles into spiraling ~9 nm protofilaments on lipid monolayers. A high-content flow cytometry assay further demonstrates that mutations halting ESCRT-III assembly block ESCRT function. Strikingly, the addition of Vps24 and Vps2 transforms flat Snf7 spirals into membrane-sculpting helices. Finally, we show that ESCRT-II and ESCRT-III coassemble into ~65 nm diameter rings indicative of a cargo-sequestering supercomplex. We propose that ESCRT-III has distinct architectural stages that are modulated by ESCRT-II to mediate cargo capture and vesicle formation by ordered assembly.

Glycosylation disorders of membrane trafficking

During evolution from prokaryotic to eukaryotic cells, compartmentalization of cellular functions has been achieved with a high degree of complexity. Notably, all secreted and transmembrane proteins travel through endoplasmic reticulum (ER) and Golgi apparatus, where they are synthesized, folded and subjected to covalent modifications, most particularly glycosylation. N-glycosylation begins in the ER with synthesis and transfer of glycan onto nascent protein and proceeds in Golgi apparatus where maturation occurs. This process not only requires the precise localization of glycosyltransferases, glycosidases and substrates but also an efficient, finely regulated and bidirectional vesicular trafficking among membrane-enclosed organelles. Basically, it is no surprise that alterations in membrane transport or related pathways can lead to glycosylation abnormalities. During the last few years, this has particularly been highlighted in genetic diseases called CDG (Congenital Disorders of Glycosylation). Alterations in mechanisms of vesicle formation due to COPII coat component SEC23B deficiency, or in vesicles tethering, caused by defects of the COG complex, but also impaired Golgi pH homeostasis due to ATP6V0A2 defects have been discovered in CDG patients. This mini review will summarize these fascinating discoveries.

Genesis and hydrocarbon significance of vesicular welded tuffs: A case study from the Fengcheng Formation, Wu-Xia area, Junggar Basin, NW China

Abstract On the basis of a large number of core and thin-section observations, the petrologic and pore evolution characteristics of welded tuffs in the Fengcheng Formation in the Wu-Xia area, Junggar Basin, are evaluated, and the genesis of vesicular welded tuffs and the formation mechanism of vesicles are analyzed. Volcanic activity in the Fengcheng Formation is characterized by welded pyroclastic rocks. There are predominant plastic shards with rounded edges and obvious devitrification in the rocks. Plastic debris (fiamme) is commonly deformed and wrinkled, and crystal fragments are usually corroded. A lot of lithophysae occur in the welded tuffs, and the welded texture is obscured due to the occurrence of these lithophysae features. Vesicles are the cavities of lithophysae, which are formed by solidification contraction. They have little or no late filling cement because the volatile constituent and water vapour in the bubbles have escaped with the help of devitrification. Lithophysae distribution within a cooling unit are characterized by vertical zonation: increasing upwards but decreasing gradually later. The cavities of lithophysae have greatly improved the reservoir quality of welded tuffs. Influenced by the intensive welding process, lithophysae are not subject to collapse or damage. In the study area, deeply buried vesicular welded tuffs are important hydrocarbon reservoir rocks, and the vesicular welded tuff has good petroliferous properties.

Cover Picture: Proteomics 22'11

AbstractMatching cupsVesicles, as a means of transport and storage of information, have been around longer than humankind. In fact, it seems they have been used as a means of communication as long as eukaryotic cells have felt the need to gossip. They have used general external vesicle systems containing secreted metabolites (cytokines, hormones, and neurotransmitters), systems specifically related to the payload of external vesicles and methods dependent on the mechanism of vesicle formation (budding off, shedding from excetome) to convey information. Koppen et al. looked at the swirling mass of various sized vesicles surrounding Drosophila and asked “Where did these come from?” Not only were the microvesicles found in eukaryotes, but also in Drosophila, nematodes, pathogenic yeast and Leishmania parasites. Cell lines shedding vesicles included Hedgehog, Wnt and Wingless. Sizes range from 40 nm to 1 μm and generally have a common, cup‐like structure.Koppen, T. et al., Proteomics 2011, 11, 4397–4410.Moles, warts, nevi, ugly brown spots, melanomas, mycoses, dermatophytes,...in a word: uglysMost of the uglys of the human race seem to be suffered by the adolescents, regardless of their natural skin color. Most of the human superficial mycoses are caused by a variety of dermatophytes, fungi that live on keratin. Many have been sequenced at the genome level (Microsporum canis, Arthroderma benhamiae). These fungi secrete virulence factors that are neutral or acidic proteases and peptidases including subtilisins, leucine amino peptidases, dipeptidyl‐peptidases,and metallocarboxypeptidases. A particularly interesting feature of all of these fungi is that as they grow, they release metabolites that improve the pH of the growth medium and thus the rate of growth. When 2‐D PAGE/shotgun LC was used to determine approximate numbers of active peptides, M. canis yielded 196 different hydrolases, including lipases, glucosidases, phosphatases, plus miscellaneous other enzymes (Background image obtained from CDC Public Health Image Library).Sriranganadane, D. et al., Proteomics 2011, 11, 4422–4433.Too late for new tricksEven software wears out in time – some of you may remember the millennium panic, when the turn of the calendar from 1999 to 2000 was going to knock the planes out of the air! Now we face the aging of proteomic and genomic analysis packages – our comfortable old shoes that cannot handle the load anymore. As of September 2011, the International Protein Index (IPI) will no longer be supported. Released in 2001, it mediated the differences between specific pairs of genomes – limited to seven specifically chosen models because of the cost of sequencing at that time. Currently there are more genomes sequenced and mutual aid agreements between analysis packages than you can shake a stick at. The replacement for IPI is UniProtKB complete Proteome set, comprising two databases, one manually curated (Swiss‐Prot) and one automatically curated (TrEMBL). Griss et al. cover a number of issues surrounding the recommended substitutions (Image provided courtesy of UniProt Consortium).Griss, J. et al., Proteomics 2011, 11, 4434–4438.

Acrosyringium Is the Main Site of the Vesicle/Pustule Formation in Palmoplantar Pustulosis

Pustulosis palmaris et plantaris or palmoplantar pustulosis (PPP) is a refractory pustular eruption on the palms and soles with unknown etiology. Numerous eccrine sweat pores exist on the palms and soles, suggesting the involvement of eccrine sweating in the pathogenesis of PPP. To the best of our knowledge, however, no definite abnormality in sweating has been documented in PPP. Accordingly, we analyzed the eccrine sweat duct involvement in the mechanism of vesicle formation in PPP. Dermatoscopy showed that PPP vesicles are located on the top of the ridges but not in the furrows. The sweat secretion in the lesional area was much lower than that in the nonlesional area, with or without pain stimulation to induce sweating. Immunostaining of horizontal sections of the lesions using antibodies against gross cystic disease fluid protein-15 (GCDFP-15) and epithelial membrane antigen (EMA) showed that these markers were localized in the cells lining the intraepidermal vesicles. Although the sweat antimicrobial peptides, dermcidin and human cathelicidin antimicrobial peptide 18 (hCAP-18)/LL-37, were detected in the fluid of the vesicles/pustules, neither dermcidin nor hCAP-18/LL-37 were overexpressed by neighboring keratinocytes. These findings suggest that the acrosyringium may be involved as the main site of the vesicle formation in the pathomechanism of PPP.

Redox switched transition of vesicles self-assembled from AOT and ferrocene derivative molecules

Vesicles prepared in an aqueous solution through self-assembly of the oxidized form of N,N-dimethyl-aminomethylferrocene (Fc +M) and an anionic surfactant, sodium bis(2-ethyl-1-hexyl) sulfosuccinate (AOT), could be reversibly transformed by redox reactions. Adding the hydroquinone as a reducing agent will cause the dissociation of vesicles and change the solution into emulsion. Subsequent oxidization by adding Ce(SO 4) 2 will regenerate vesicles. The vesicle structure and morphology were characterized by transmission electron microscopy (TEM) and dynamic light scattering (DLS), respectively and the mean particle size was about 200 nm. The mechanism of vesicle formation and disruption caused by redox reactions is discussed along with the data obtained from cyclic voltammetry (CV) and UV–vis spectroscopy measurements. The redox state changes of the ferrocenyl moiety may influence the noncovalent interactions between FcM and AOT. Meanwhile, the π–π stacking and amphiphilic hydrophobic association are also included during such a redox modulated process. These results provide guidance for the design of surfactant and small molecular systems that permit active control of self-assembly.

A lipase-like gene from Heliothis virescens ascovirus (HvAV-3e) is essential for virus replication and cell cleavage

A unique feature of ascovirus infection is cleavage of host cells into virus containing vesicles. It has been suggested that the virus induces apoptosis, either by expression of a caspase or other means, which is then diverted toward vesicle formation. There is little known about the mechanism of vesicle formation. Recent genome sequences of three ascoviruses indicated the presence of several putative open reading frames coding for proteins that could be involved in lipid metabolism. These proteins may play a role in rearrangement of membranes in infected host cells leading to formation of vesicles. Here, we analyzed a lipase-like gene (ORF19) from Heliothis virescens ascovirus (HvAV-3e) expressed from 8 h after infection and essential for virus replication and cell cleavage. In addition, ORF19 knock down by RNA interference inhibited virus replication indicating that the gene is indispensable for HvAV-3e replication. However, under enzymatic assays tested, we did not detect any lipase or esterase activity from ORF19.

Self-assembly microstructures of amphiphilic polyborate in aqueous solutions

A new type of amphiphilic homo-polyborate was synthesized with the surface tension ( γ CMC) in deionized water 31.2 mN m −1 (pH = 7, 25 °C), which is similar to low-molecule surfactants. Self-assembly behaviors of the polyborate in aqueous solutions were investigated by TEM. Self-aggregation morphologies of the polyborate can be controlled by the concentration of NaCl. With the increase of the concentration of NaCl, micromorphologies from the amphiphilic polyborate transformed from spherical polymeric vesicles to tree-like or even vertebra-like morphologies. It was also found that, the polymeric vesicles show a strong stability against ethanol addition, even when the concentration of ethanol increases to a volume ratio of 33%, they still maintain good micromorphologies. A possible mechanism of vesicle formation was proposed based on packing parameter theory.

Dynamics of vesicle formation from lipid droplets: Mechanism and controllability

A coarse-grained model developed by Marrink et al. [J. Phys. Chem. B 111, 7812 (2007)] is applied to investigate vesiculation of lipid [dipalmitoylphosphatidylcholine (DPPC)] droplets in water. Three kinds of morphologies of micelles are found with increasing lipid droplet size. When the initial lipid droplet is smaller, the equilibrium structure of the droplet is a spherical micelle. When the initial lipid droplet is larger, the lipid ball starts to transform into a disk micelle or vesicle. The mechanism of vesicle formation from a lipid ball is analyzed from the self-assembly of DPPC on the molecular level, and the morphological transition from disk to vesicle with increasing droplet size is demonstrated. Importantly, we discover that the transition point is not very sharp, and for a fixed-size lipid ball, the disk and vesicle appear with certain probabilities. The splitting phenomenon, i.e., the formation of a disk/vesicle structure from a lipid droplet, is explained by applying a hybrid model of the Helfrich membrane theory. The elastic module of the DPPC bilayer and the smallest size of a lipid droplet for certain formation of a vesicle are successfully predicted.

Spontaneous Vesicle Formation of Poly(ethylene oxide)−Poly(propylene oxide)−Poly(ethylene oxide) Triblock Copolymer

A novel method has been developed to prepare vesicles from aqueous solutions of poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) triblock copolymer, by adding anionic surfactant sodium dodecyl sulfate (SDS) and inorganic salt NaF. As determined by TEM and dynamic light scattering (DLS) measurements, the average diameter of vesicles is about 800 nm having 50 nm outer shell thickness. Identifying hydrophobic interactions between the block copolymers and the microenvironments around the vesicles using FTIR, 1H NMR, and fluorescence spectroscopy techniques revealed the vesicle formation mechanism. The spontaneously formed vesicles were further cross-linked by converting the terminal hydroxyl groups of block copolymers into aldehydes, and then chemically bridging the polymer chains by the reaction between aldehydes and diamine compounds. The cross-linked vesicles are proved much more stable than free vesicles even at higher dilutions. The obtained vesicles with good stability and biocompatibility are promising candidates for widespread applications.

Fluctuations and destabilization of single phospholipid bilayers

Supported phospholipid bilayers are interesting model systems for biologists and present fascinating physical properties. The authors present an extensive experimental study of the dynamic properties of supported bilayers. The structure and the equilibrium properties of single and double supported bilayers were investigated with neutron reflectivity. The submicronic fluctuation spectrum of a nearly free "floating" bilayer was determined using off-specular x-ray scattering: the surface tension of the bilayer, its bending modulus, and the intermembrane potential could be determined. Using fluorescence microscopy, the authors showed that this well-controlled single bilayer can form vesicles. Destabilization occurred either at the main gel-fluid transition of the lipids and could be interpreted in terms of a decrease in the bending rigidity or under an ac low-frequency electric field applied in the fluid phase. In the latter case, the authors also studied the effect of the electric field at the molecular length scale by neutron reflectivity. In both cases, destabilization leads to the formation of relatively monodisperse vesicles. This could give further understanding on the vesicle formation mechanism and on the parameters that determine the vesicle size.

Proteomics in gram‐negative bacterial outer membrane vesicles

Gram-negative bacteria constitutively secrete outer membrane vesicles (OMVs) into the extracellular milieu. Recent research in this area has revealed that OMVs may act as intercellular communicasomes in polyspecies communities by enhancing bacterial survival and pathogenesis in hosts. However, the mechanisms of vesicle formation and the pathophysiological roles of OMVs have not been clearly defined. While it is obvious that mass spectrometry-based proteomics offers great opportunities for improving our knowledge of bacterial OMVs, limited proteomic data are available for OMVs. The present review aims to give an overview of the previous biochemical, biological, and proteomic studies in the emerging field of bacterial OMVs, and to give future directions for high-throughput and comparative proteomic studies of OMVs that originate from diverse Gram-negative bacteria under various environmental conditions. This article will hopefully stimulate further efforts to construct a comprehensive proteome database of bacterial OMVs that will help us not only to elucidate the biogenesis and functions of OMVs but also to develop diagnostic tools, vaccines, and antibiotics effective against pathogenic bacteria.

Impact of mechanism of formation on encapsulation in block copolymer vesicles

Vesicles prepared from block copolymers have been mooted for the encapsulation of water-soluble molecules. This is because the membranes of polymer vesicles have been shown to be more stable than those in vesicles formed from lipids, with the membrane properties being tuned by the length and nature of the hydrophobic block in the polymer. The generally accepted mechanisms of vesicle formation involve either wrap-up of a lamellar sheet or formation via a sequence of micelle to worm to disks to vesicles. These should lead to efficient encapsulation. Alternatively, a method involving phase separation followed by re-structuring has been recently suggested. Here, we show that this final mechanism holds for vesicles formed from a PEO- b-PDEAMA copolymer by a pH switch and that this mechanism leads to highly inefficient encapsulation on vesicle formation.

Self‐Assembly Vesicles Made from a Cyclodextrin Supramolecular Complex

Self-assembly vesicles have been made from a cyclodextrin (CD) supramolecular complex, which is cooperatively formed with natural beta-CD, 1-naphthylammonium chloride (NA), and sodium bis(2-ethyl-1-hexyl)sulfosuccinate (AOT) by weak noncovalent interactions. In the complex structure, a NA molecule is included inside a beta-CD molecule while it is coupled with an AOT molecule on one side. The supramolecular structure and morphology of the vesicles were characterized by transmission electron microscopy (TEM) and dynamic light scattering (DLS), respectively. The mechanism of vesicle formation and transition is discussed along with the data obtained from induced circular dichroism (ICD) and UV/visible spectroscopy, polarized optical microscopy (POM), and (1)H NMR spectroscopy. Both the fabrication and the transition of vesicles are controlled by the inclusion equilibria and the cooperative binding of noncovalent interactions, which include the "key-lock" principle, electrostatic interactions, pi-pi stacking, and amphiphilic hydrophobic association.

Proteomic Analysis of Microvesicles Derived from Human Colorectal Cancer Cells

Microvesicles (MV) are membrane vesicles secreted from the plasma and endosomal membrane compartment by various cell types such as hematopoietic, epithelial, and tumor cells. Actively growing tumor cells shed MV, and the rate of shedding increases in malignant tumors. Although recent progress in this area has revealed that tumor-derived MV play multiple roles in tumor growth and metastasis via immune escape, tumor invasion, and angiogenesis, the mechanism of vesicle formation and the biological roles of tumor-derived MV are not understood. Here, we report the first global proteomic analysis of highly purified MV from human colorectal cancer cells. Using 1D SDS gel electrophoresis and nano-LC-MS/MS analyses, we identified a total of 547 microvesicular proteins from three independent experiments with high confidence; 416 proteins were identified at least in two trials, including 181 as yet unreported proteins. We identified 49 proteins involved in the biogenesis of MV, including annexins, ADP-ribosylation factors, and Rab proteins. We also identified 28 proteins that may function in tumorigenesis via promotion of migration, invasion, and growth of tumor cells, immune modulation, metastasis, and angiogenesis. Taken together with previously reported results, our observations suggest that tumor-derived MV may act as communicasomes, that is, extracellular organelles that play diverse roles in intercellular communication. This information will help elucidate the biogenesis and functions of tumor-derived MV, and aid in the development of effective vaccines for various cancers, including colorectal cancer.

Global proteomic profiling of native outer membrane vesicles derived from Escherichia coli

Gram-negative bacteria constitutively secrete native outer membrane vesicles (OMVs) into the extracellular milieu. Although recent progress in this area has revealed that OMVs are essential for bacterial survival and pathogenesis, the mechanism of vesicle formation and the biological roles of OMVs have not been clearly defined. Using a proteomics approach, we identified 141 protein components of Escherichia coli-derived native OMVs with high confidence; two separate analyses yielded identifications of 104 and 117 proteins, respectively, with 80 proteins overlapping between the two trials. In the group of identified proteins, the outer membrane proteins were highly enriched, whereas inner membrane proteins were lacking, suggesting that a specific sorting mechanism for vesicular proteins exists. We also identified proteins involved in vesicle formation, the removal of toxic compounds and attacking phage, and the elimination of competing organisms, as well as those involved in facilitating the transfer of genetic material and protein to other bacteria, targeting host cells, and modulating host immune responses. This study provides a global view of native bacterial OMVs. This information will help us not only to elucidate the biogenesis and functions of OMV from nonpathogenic and pathogenic bacteria but also to develop vaccines and antibiotics effective against pathogenic strains.

Density functional simulation of spontaneous formation of vesicle in block copolymer solutions

The author carries out numerical simulations of vesicle formation based on the density functional theory for block copolymer solutions. It is shown by solving the time evolution equations for concentrations that a polymer vesicle is spontaneously formed from the homogeneous state. The vesicle formation mechanism obtained by this simulation agrees with the results of other simulations based on the particle models as well as experiments. By changing parameters such as the volume fraction of polymers or the Flory-Huggins interaction parameter between the hydrophobic subchains and solvents, the spherical micelles, cylindrical micelles, or bilayer structures can also be obtained. The author also shows that the morphological transition dynamics of the micellar structures can be reproduced by controlling the Flory-Huggins interaction parameter.