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EDITORIAL |
Department of Biomedical Sciences, University of Sheffield, Western Bank, Sheffield S10 2TN, UKEmail: e.p.seward{at}sheffield.ac.uk
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Elementary properties of exocytosis and endocytosis |
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 Top Elementary properties of... Multiple synaptic vesicle...Signals and SNAREs regulating...References |
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Exocytosis, representing the fusion between the vesicle and the plasma membranes, has been a focus for the life sciences for more than 40 years, and still represents an exciting topic, since it involves a highly organized subcellular structure (the vesicle) that allows, in some systems, rather rapid physiological responses. It is thought that the release of vesicle cargo into the extracellular space by exocytosis involves the formation and opening of a fusion pore. Recent studies have revealed that fusion pore formation may lead to the release of vesicle cargo by complete flattening of the vesicle membrane into the plasma membrane. Alternatively, a fusion pore may form transiently, which has ben termed ‘kiss-and-run’ exocytosis. While the latter phenomenon appears to be relatively frequent in some cell types, the mechanisms underlying its occurrence are poorly understood and many open questions remain to be addressed. How do fusion pore diameter and dwell time affect the release of substances stored in vesicles? How is kiss-and-run exocytosis related to the regulation of secretory output from nerve terminals? Is kiss-and-run exocytosis related to the recruitment of the reserve vesicles? These questions are considered in two papers addressing, to some extent, extreme cases of regulated exocytosis.
Vardjan et al. (2007) deal with exocytosis in a relatively slow secretory cell, releasing prolactin. The pituitary lactotroph is devoid of clusters of vesicles beneath the plasma membrane, such as the active zones in neurons. The authors discuss the elementary properties of spontaneous and stimulated fusion of peptidergic vesicles, highlighting that vesicle cargo discharge from a single vesicle is the subject of regulation. Transient kiss-and-run events are the dominant mode of exocytosis in these cells and compared to the stimulated events, spontaneous events occur less frequently with a shorter fusion pore open time and a narrower pore diameter. Therefore, at rest the release of vesicle cargo is likely to be restrained kinetically and/or due to a narrow fusion pore (unproductive exocytosis). Thus, fusion pore gating appears to play an important role in the modulation of peptide release from a single vesicle.
Heidelberger (2007) describes properties of regulated exocytosis of a neuron exhibiting a highly structured presynaptic organization of vesicles, the ribbon synapse. In conventional synapses, neurotransmitter release occurs as a brief burst of exocytosis. In contrast, photoreceptors release neurotransmitters via exocytosis in a graded manner for extended periods of time. Based on their results, the ability of ribbon synapse of rod photoreceptors in salamanders to maintain the vesicle cargo release for extended periods is possible due to a relatively large releasable pool of vesicles in ribbon synapse with a fast rate of refilling from preformed vesicles. The relationship between the secretory activity and cytosolic calcium is unusually shallow in photoreceptors and the rate of exocytosis almost linearly tracks the local calcium concentration, which is likely to contribute to the graded manner of vesicle cargo release.
What the common mechanisms determining exocytosis in these two systems are, however, will have to be determined in the future.
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Multiple synaptic vesicle retrieval pathways in neuronal physiology |
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TopElementary properties of...Multiple synaptic vesicle...Signals and SNAREs regulating...References |
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There are at least three different mechanisms by which a central nerve terminal can retrieve synaptic vesicle (SV) membrane after exocytosis, and three articles in this issue of The Journal of Physiology highlight different aspects of them. The routes are (1) classical clathrin-dependent endocytosis, which retrieves single SVs, (2) a fast reuse pathway (also called kiss-and-run), which immediately replenishes the readily releasable pool of SVs, and (3) a pathway called bulk endocytosis, where large areas of nerve terminal membrane are invaginated to form endosomes from which SVs can bud. Researchers are now dedicated to discovering which pathways are active in which neurones, how these pathways are activated by patterns of neuronal activity and what their basic molecular mechanisms are.
Kavalali (2007) presents evidence that the reuse pathway contributes towards maintenance of exocytosis during continual synaptic activity. This contribution varies between excitatory and inhibitory synapses and also between neuronal types, suggesting different neurones have different strategies for dealing with SV retrieval depending on their need.
Granseth et al. (2007) show that at mild stimulation frequencies, only one SV retrieval route appears to be present, which has a time constant of 15 s and is totally dependent on clathrin. At first glance this may argue against a contribution from the reuse pathway, but another option is possible, which is that reuse is also a clathrin-dependent process and the fast recycling of SVs back to the readily releasable pool occurs after SV retrieval from the plasma membrane.
Clayton et al. (2007) show that during intense physiological stimulation the bulk endocytosis pathway can be recruited to facilitate SV retrieval. This recruitment is reliant on the activity-dependent stimulation of the calcium-dependent protein phosphatase calcineurin, which dephosphorylates a subset of endocytosis proteins called the dephosphins. This suggests that that dephosphorylation of the dephosphins is the key molecular event in the recruitment of bulk endocytosis in central nerve terminals.
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Signals and SNAREs regulating vesicle exocytosis |
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TopElementary properties of...Multiple synaptic vesicle...Signals and SNAREs regulating...References |
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Over the past decade there has been tremendous progress in our understanding of the molecular mechanisms underlying exocytosis and in particular in the function of SNAREs (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptors) in regulating fusion of neurotransmitter-storing vesicles with the plasma membrane. More recently the focus of many researchers in the field has shifted towards identifying molecules that regulate SNAREs.
Lang (2007) has addressed the contentious issue of the role of ‘lipid-rafts’ in organizing SNAREs. If seeing is believing, then the STED-microscopic analysis of syntaxin and SNAP-25 organization in the plasma membrane strongly suggests that these proteins are indeed organized into ‘membrane rafts’. Whether the observed clusters of syntaxin define docking and fusion sites remains to be established, but if they do, then segregation of different syntaxins by specific cluster formation would provide a simple and efficient method to spatially separate different types of exocytotic events within a cell and hence compartmentalize cellular processes.
Membrane lipids may not only regulate the spatial organization of SNARE proteins but also play a critical role in regulating their function. Darios et al. (2007) describe how phospholipid-derived polyunsaturated fatty acids may activate syntaxin by promoting the ‘open’ configuration necessary for interactions with its SNARE partners. Remarkably, this activation mechanism can occur even on syntaxins ‘clamped’ shut by the protein Munc18, indicating that abolition of syntaxin–Munc18 association may not be a necessary prerequisite for SNARE complex formation. Production of fatty acids at the membrane is tightly regulated by a number of well known phospholipases, which are themselves regulated by a variety of signalling cascades, and thus the pathway described by Darios et al. provides a novel mechanism by which cell surface receptors may regulate the formation of SNARE complexes, vesicle fusion and the secretory output of a cell.
The importance of regulating syntaxin function is again highlighted in the report by Gracheva et al. (2007). Analysis of neurotransmission in tom-1 mutant C. elegans reveal that tomosyn, a syntaxin-binding protein, negatively regulates exocytosis by sequestering syntaxin and limiting its availability to form functional SNARE complexes. Ultrastructural analysis indicated that tomosyn may be acting at the level of vesicle priming. Thus is would appear that syntaxin plays a key role not just in vesicle fusion but also docking and priming. Moreover the results of these three papers show that, like ion channels, SNARE proteins are also subject to complex regulation by a variety of molecules.
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References |
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TopElementary properties of...Multiple synaptic vesicle...Signals and SNAREs regulating...References |
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Darios F, Connell E & Davletov B (2007). Phospholipases and fatty acid signalling in exocytosis. J Physiol 585, 699–704.
Gracheva EO, Burdina AO, Touroutine D, Berthelot-Grosjean M, Parekh H & Richmond JE (2007). Tomosyn negatively regulates both synaptic transmitter and neuropeptide release at the C. elegans neuromuscular junction. J Physiol 585, 705–709.
Granseth B, Odermatt B, Royle S & Lagnado L (2007). Clathrin-mediated endocytosis: the physiological mechanism of vesicle retrieval at hippocampal synapses. J Physiol 585, 681–685.
Heidelberger R (2007). Mechanisms of tonic, graded release: lessons from the vertebrate photoreceptor. J Physiol 585, 663–667.
Kavalali ET (2007). Multiple vesicle recycling pathways in central synapses and their impact on neurotransmission. J Physiol 585, 669–679.
Lang T (2007). SNARE proteins and ‘membrane rafts’. J Physiol 585, 693–698.
Vardjan N, Stenovec M, Jorgacevski J, Kreft M & Zorec R (2007). Elementary properties of spontaneous fusion of peptidergic vesicles: fusion pore gating. J Physiol 585, 655–661.
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