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Traffic (Copenhagen, Denmark) Apr 2015Neuronal communication relies on chemical synaptic transmission for information transfer and processing. Chemical neurotransmission is initiated by synaptic vesicle... (Review)
Review
Neuronal communication relies on chemical synaptic transmission for information transfer and processing. Chemical neurotransmission is initiated by synaptic vesicle fusion with the presynaptic active zone resulting in release of neurotransmitters. Classical models have assumed that all synaptic vesicles within a synapse have the same potential to fuse under different functional contexts. In this model, functional differences among synaptic vesicle populations are ascribed to their spatial distribution in the synapse with respect to the active zone. Emerging evidence suggests, however, that synaptic vesicles are not a homogenous population of organelles, and they possess intrinsic molecular differences and differential interaction partners. Recent studies have reported a diverse array of synaptic molecules that selectively regulate synaptic vesicles' ability to fuse synchronously and asynchronously in response to action potentials or spontaneously irrespective of action potentials. Here we discuss these molecular mediators of vesicle pool heterogeneity that are found on the synaptic vesicle membrane, on the presynaptic plasma membrane, or within the cytosol and consider some of the functional consequences of this diversity. This emerging molecular framework presents novel avenues to probe synaptic function and uncover how synaptic vesicle pools impact neuronal signaling.
Topics: Action Potentials; Animals; Humans; Neurotransmitter Agents; Synaptic Membranes; Synaptic Transmission; Synaptic Vesicles
PubMed: 25620674
DOI: 10.1111/tra.12262 -
Journal of Extracellular Vesicles Jan 2021Extracellular vesicles (EVs) are secreted nano-sized vesicles that contain cellular proteins, lipids, and nucleic acids. Although EVs are expected to be biologically...
Extracellular vesicles (EVs) are secreted nano-sized vesicles that contain cellular proteins, lipids, and nucleic acids. Although EVs are expected to be biologically diverse, current analyses cannot adequately characterize this diversity because most are ensemble methods that inevitably average out information from diverse EVs. Here we describe a single vesicle analysis, which directly visualizes marker expressions of individual EVs using a total internal-reflection microscopy and analyzes their co-localization to investigate EV subpopulations. The single-vesicle imaging and co-localization analysis successfully illustrated the diversity of EVs and revealed distinct patterns of tetraspanin expressions. Application of the analysis demonstrated similarities and dissimilarities between the EV fractions that had been acquired from different conventional EV isolation methods. The analysis method developed in this study will provide a new and reliable tool for investigating characteristics of single EVs, and the findings of the analysis might increase understanding of the characteristics of EVs.
Topics: Biomarkers; Cell Line; Chromatography, Gel; Extracellular Vesicles; HEK293 Cells; Humans; Microscopy; Tetraspanins
PubMed: 33456726
DOI: 10.1002/jev2.12047 -
Journal of Extracellular Vesicles May 2023Membrane-bound vesicles such as extracellular vesicles (EVs) can function as biochemical effectors on target cells. Docking of the vesicles onto recipient plasma...
Membrane-bound vesicles such as extracellular vesicles (EVs) can function as biochemical effectors on target cells. Docking of the vesicles onto recipient plasma membranes depends on their interaction with cell-surface proteins, but a generalizable technique that can quantitatively observe these vesicle-protein interactions (VPIs) is lacking. Here, we describe a fluorescence microscopy that measures VPIs between single vesicles and cell-surface proteins, either in a surface-tethered or in a membrane-embedded state. By employing cell-derived vesicles (CDVs) and intercellular adhesion molecule-1 (ICAM-1) as a model system, we found that integrin-driven VPIs exhibit distinct modes of affinity depending on vesicle origin. Controlling the surface density of proteins also revealed a strong support from a tetraspanin protein CD9, with a critical dependence on molecular proximity. An adsorption model accounting for multiple protein molecules was developed and captured the features of density-dependent cooperativity. We expect that VPI imaging will be a useful tool to dissect the molecular mechanisms of vesicle adhesion and uptake, and to guide the development of therapeutic vesicles.
Topics: Extracellular Vesicles; Cell Communication; Cell Membrane; Membrane Proteins
PubMed: 37186457
DOI: 10.1002/jev2.12322 -
Cell Reports Feb 2017Vesicle sharing between synaptic boutons is an important component of the recycling process that synapses employ to maintain vesicle pools. However, the mechanisms...
Vesicle sharing between synaptic boutons is an important component of the recycling process that synapses employ to maintain vesicle pools. However, the mechanisms supporting and regulating vesicle transport during the inter-synaptic exchange remain poorly understood. Using nanometer-resolution tracking of individual synaptic vesicles and advanced computational algorithms, we find that long-distance axonal transport of synaptic vesicles between hippocampal boutons is partially mediated by the actin network, with myosin V as the primary actin-dependent motor that drives this vesicle transport. Furthermore, we find that vesicle exit from the synapse to the axon and long-distance vesicle transport are both rapidly and dynamically regulated by activity. We corroborated these findings with two complementary modeling approaches of vesicle exit, which closely reproduced experimental observations. These findings uncover the roles of actin and myosin V in supporting the inter-synaptic vesicle exchange and reveal that this process is dynamically modulated in an activity-dependent manner.
Topics: Actins; Animals; Axonal Transport; Axons; Cells, Cultured; Hippocampus; Myosin Type V; Neurons; Presynaptic Terminals; Rats; Synapses; Synaptic Transmission; Synaptic Vesicles
PubMed: 28249156
DOI: 10.1016/j.celrep.2017.02.010 -
The Journal of Neuroscience : the... Apr 2021Synaptophysin (syp) is a major integral membrane protein of secretory vesicles. Previous work has demonstrated functions for syp in synaptic vesicle cycling,...
Synaptophysin (syp) is a major integral membrane protein of secretory vesicles. Previous work has demonstrated functions for syp in synaptic vesicle cycling, endocytosis, and synaptic plasticity, but the role of syp in the process of membrane fusion during Ca-triggered exocytosis remains poorly understood. Furthermore, although syp resides on both large dense-core and small synaptic vesicles, its role in dense-core vesicle function has received less attention compared with synaptic vesicle function. To explore the role of syp in membrane fusion and dense-core vesicle function, we used amperometry to measure catecholamine release from single vesicles in male and female mouse chromaffin cells with altered levels of syp and the related tetraspanner protein synaptogyrin (syg). Knocking out syp slightly reduced the frequency of vesicle fusion events below wild-type (WT) levels, but knocking out both syp and syg reduced the frequency 2-fold. Knocking out both proteins stabilized initial fusion pores, promoted fusion pore closure (kiss-and-run), and reduced late-stage fusion pore expansion. Introduction of a syp construct lacking its C-terminal dynamin-binding domain in syp knock-outs (KOs) increased the duration and fraction of kiss-and-run events, increased total catecholamine release per event, and reduced late-stage fusion pore expansion. These results demonstrated that syp and syg regulate dense-core vesicle function at multiple stages to initiate fusion, control the choice of mode between full-fusion and kiss-and-run, and influence the dynamics of both initial and late-stage fusion pores. The transmembrane domain (TMD) influences small initial fusion pores, and the C-terminal domain influences large late-stage fusion pores, possibly through an interaction with dynamin. The secretory vesicle protein synaptophysin (syp) is known to function in synaptic vesicle cycling, but its roles in dense-core vesicle functions, and in controlling membrane fusion during Ca-triggered exocytosis remain unclear. The present study used amperometry recording of catecholamine release from endocrine cells to assess the impact of syp and related proteins on membrane fusion. A detailed analysis of amperometric spikes arising from the exocytosis of single vesicles showed that these proteins influence fusion pores at multiple stages and control the choice between kiss-and-run and full-fusion. Experiments with a syp construct lacking its C terminus indicated that the transmembrane domain (TMD) influences the initial fusion pore, while the C-terminal domain influences later stages after fusion pore expansion.
Topics: Animals; Animals, Newborn; Catecholamines; Chromaffin Cells; Dynamins; Electrophysiological Phenomena; Exocytosis; Female; Membrane Fusion; Mice; Mice, Knockout; Pregnancy; Primary Cell Culture; Synaptic Vesicles; Synaptogyrins; Synaptophysin
PubMed: 33664131
DOI: 10.1523/JNEUROSCI.2833-20.2021 -
Applied and Environmental Microbiology Dec 2022The exchange of bacterial extracellular vesicles facilitates molecular exchange between cells, including the horizontal transfer of genetic material. Given the...
The exchange of bacterial extracellular vesicles facilitates molecular exchange between cells, including the horizontal transfer of genetic material. Given the implications of such transfer events on cell physiology and adaptation, some bacterial cells have likely evolved mechanisms to regulate vesicle exchange. Past work has identified mechanisms that influence the formation of extracellular vesicles, including the production of small molecules that modulate membrane structure; however, whether these mechanisms also modulate vesicle uptake and have an overall impact on the rate of vesicle exchange is unknown. Here, we show that membrane-binding molecules produced by microbes influence both the formation and uptake of extracellular vesicles and have the overall impact of increasing the vesicle exchange rate within a bacterial coculture. In effect, production of compounds that increase vesicle exchange rates encourage gene exchange between neighboring cells. The ability of several membrane-binding compounds to increase vesicle exchange was demonstrated. Three of these compounds, nisin, colistin, and polymyxin B, are antimicrobial peptides added at sub-inhibitory concentrations. These results suggest that a potential function of exogenous compounds that bind to membranes may be the regulation of vesicle exchange between cells. The exchange of bacterial extracellular vesicles is one route of gene transfer between bacteria, although it was unclear if bacteria developed strategies to modulate the rate of gene transfer within vesicles. In eukaryotes, there are many examples of specialized molecules that have evolved to facilitate the production, loading, and uptake of vesicles. Recent work with bacteria has shown that some small molecules influence membrane curvature and induce vesicle formation. Here, we show that similar compounds facilitate vesicle uptake, thereby increasing the overall rate of vesicle exchange within bacterial populations. The addition of membrane-binding compounds, several of them antibiotics at subinhibitory concentrations, to a bacterial coculture increased the rate of horizontal gene transfer via vesicle exchange.
Topics: Bacteria; Gene Transfer, Horizontal; Extracellular Vesicles; Membranes; Eukaryota
PubMed: 36342184
DOI: 10.1128/aem.01346-22 -
Cancers Sep 2022Extracellular vesicles are membrane-bound vesicles released by cells to mediate intercellular communication and homeostasis. Various external stimuli as well as inherent... (Review)
Review
Extracellular vesicles are membrane-bound vesicles released by cells to mediate intercellular communication and homeostasis. Various external stimuli as well as inherent abnormalities result in alterations in the extracellular vesicle milieu. Changes to cells result in alterations in the content of the extracellular vesicle biogenesis, which may affect proximal and distal cells encountering these altered extracellular vesicles. Therefore, the examination of changes in the extracellular vesicle signature can be used to follow disease progression, reveal possible targets to improve therapy, as well as to serve as mediators of therapy. Furthermore, recent studies have developed methods to alter the cargo of extracellular vesicles to restore normal function or deliver therapeutic agents. This review will examine how extracellular vesicles from cancer cells differ from normal cells, how these altered extracellular vesicles can contribute to cancer progression, and how extracellular vesicles can be used as a therapeutic agent to target cancer cells and cancer-associated stroma. Here we present extracellular vesicles as a novel tool in nanomedicine.
PubMed: 36139610
DOI: 10.3390/cancers14184450 -
Traffic (Copenhagen, Denmark) Sep 2007Two models of synaptic vesicle recycling have been intensely debated for decades: kiss-and-run, in which the vesicle opens and closes transiently, presumably through a... (Review)
Review
Two models of synaptic vesicle recycling have been intensely debated for decades: kiss-and-run, in which the vesicle opens and closes transiently, presumably through a small fusion pore, and full fusion, in which the vesicle collapses into the plasma membrane and is retrieved by clathrin-coat-dependent processes. Conceptually, it seems that kiss-and-run would be faster and would retrieve vesicles with greater fidelity. Is this the case? This review discusses recent evidence for both models. We conclude that both mechanisms allow for high fidelity of vesicle recycling. Also, the presence in the plasma membrane of a depot of previously fused vesicles that are already interacting with the endocytotic machinery (the 'readily retrievable' vesicles) allows full fusion to trigger quite fast endocytosis, further blurring the efficiency differences between the two models.
Topics: Animals; Clathrin-Coated Vesicles; Endocytosis; Exocytosis; Humans; Membrane Fusion; Models, Biological; Synaptic Membranes; Synaptic Vesicles; Transport Vesicles
PubMed: 17645434
DOI: 10.1111/j.1600-0854.2007.00614.x -
Developmental Biology Jun 2020Arl13b is a gene known to regulate ciliogenesis. Functional alterations in this gene's activity have been associated with Joubert syndrome. We found that in Arl13 null...
Arl13b is a gene known to regulate ciliogenesis. Functional alterations in this gene's activity have been associated with Joubert syndrome. We found that in Arl13 null mouse embryos the orientation of the optic cup is inverted, such that the lens is abnormally surrounded by an inverted optic cup whose retina pigmented epithelium is oddly facing the surface ectoderm. Loss of Arl13b leads to the disruption of optic vesicle's patterning and expansion of ventral fates. We show that this phenotype is consequence of miss-regulation of Sonic hedgehog (Shh) signaling and demonstrate that the Arl13b eye phenotype can be rescued by deletion of Gli2, a downstream effector of the Shh pathway. This work identified an unexpected role of primary cilia during the morphogenetic movements required for the formation of the eye.
Topics: ADP-Ribosylation Factors; Animals; Body Patterning; Bone Morphogenetic Protein 4; Cilia; Embryonic Development; Eye; Eye Proteins; Gene Expression Regulation, Developmental; Hedgehog Proteins; Homeodomain Proteins; Humans; Lens, Crystalline; Male; Mice; Mice, Knockout; Morphogenesis; Nerve Tissue Proteins; Organogenesis; Retinal Pigment Epithelium; Signal Transduction; Wnt1 Protein; Zinc Finger Protein Gli2; Homeobox Protein SIX3
PubMed: 32169553
DOI: 10.1016/j.ydbio.2020.02.016 -
Journal of Neurochemistry Jun 2007Synaptic vesicles are key organelles in neurotransmission. Vesicle integral or membrane-associated proteins mediate the various functions the organelle fulfills during... (Review)
Review
Synaptic vesicles are key organelles in neurotransmission. Vesicle integral or membrane-associated proteins mediate the various functions the organelle fulfills during its life cycle. These include organelle transport, interaction with the nerve terminal cytoskeleton, uptake and storage of low molecular weight constituents, and the regulated interaction with the pre-synaptic plasma membrane during exo- and endocytosis. Within the past two decades, converging work from several laboratories resulted in the molecular and functional characterization of the proteinaceous inventory of the synaptic vesicle compartment. However, up until recently and due to technical difficulties, it was impossible to screen the entire organelle thoroughly. Recent advances in membrane protein identification and mass spectrometry (MS) have dramatically promoted this field. A comparison of different techniques for elucidating the proteinaceous composition of synaptic vesicles revealed numerous overlaps but also remarkable differences in the protein constituents of the synaptic vesicle compartment, indicating that several protein separation techniques in combination with differing MS approaches are required to identify and characterize the synaptic vesicle proteome. This review highlights the power of various gel separation techniques and MS analyses for the characterization of the proteome of highly purified synaptic vesicles. Furthermore, the newly detected protein assignments to synaptic vesicles, especially those proteins which are new to the inventory of the synaptic vesicle proteome, are critically discussed.
Topics: Animals; Clathrin-Coated Vesicles; GTP-Binding Proteins; Humans; Membrane Proteins; Nerve Tissue Proteins; Neurotransmitter Transport Proteins; Proteasome Endopeptidase Complex; Protein Isoforms; Proteome; Synaptic Vesicles
PubMed: 17355250
DOI: 10.1111/j.1471-4159.2007.04453.x