Vesicle fusion

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Vesicle fusion is the merging of a vesicle with other vesicles or a part of a cell membrane. In the latter case, it is the end stage of secretion from secretory vesicles, where their contents are expelled from the cell through exocytosis. Vesicles can also fuse with other target cell compartments, such as a lysosome. Exocytosis occurs when secretory vesicles transiently dock and fuse at the base of cup-shaped structures at the cell plasma membrane called porosome, the universal secretory machinery in cells. Vesicle fusion may depend on SNARE proteins in the presence of increased intracellular calcium (Ca2+) concentration.

Contents

Triggers

Stimuli that trigger vesicle fusion act by increasing intracellular Ca2+.

Model systems

Model systems consisting of a single phospholipid or a mixture have been studied by physical chemists. Cardiolipin is found mainly in mitochondrial membranes, and calcium ions play an important role in the respiratory processes mediated by the mitochondrion. The forces involved have been postulated to explain [3] this process in terms of nucleation for agglomeration of smaller supramolecular entities or phase changes in the structure of the biomembranes. [4]

Mechanisms

Synaptic cleft fusion

In synaptic vesicle fusion, the vesicle must be within a few nanometers of the target membrane for the fusion process to begin. This closeness allows the cell membrane and the vesicle to exchange lipids which is mediated by certain proteins which remove water that comes between the forming junction. Once the vesicle is in position it must wait until Ca2+ enters the cell by the propagation of an action potential to the presynaptic membrane. [5] Ca2+ binds to specific proteins, one of which is Synaptotagmin, in neurons which triggers the complete fusion of the vesicle with the target membrane. [6]

SNARE proteins are also thought to help mediate which membrane is the target of which vesicle. [7]

SNARE protein and pore formation

Molecular machinery driving exocytosis in neuromediator release. The core SNARE complex is formed by four a-helices contributed by synaptobrevin, syntaxin and SNAP-25, synaptotagmin serves as a calcium sensor and regulates intimately the SNARE zipping. Exocytosis-machinery.jpg
Molecular machinery driving exocytosis in neuromediator release. The core SNARE complex is formed by four α-helices contributed by synaptobrevin, syntaxin and SNAP-25, synaptotagmin serves as a calcium sensor and regulates intimately the SNARE zipping.

Assembly of the SNAREs into the "trans" complexes likely bridges the opposing lipid bilayers of membranes belonging to cell and secretory granule, bringing them in proximity and inducing their fusion. The influx of calcium into the cell triggers the completion of the assembly reaction, which is mediated by an interaction between the putative calcium sensor, synaptotagmin, with membrane lipids and/or the partially assembled SNARE complex.

One hypothesis implicates the molecule Complexin within the SNARE complex and its interaction with the molecule synaptotagmin. [9] Known as the "clamp" hypothesis, the presence of complexin normally inhibits the fusion of the vesicle to the cell membrane. However, binding of calcium ions to synaptotagmin triggers the complexin to be released or inactivated, so that the vesicle is then free to fuse. [10]

According to the "zipper" hypothesis, the complex assembly starts at the N-terminal parts of SNARE motifs and proceeds towards the C-termini that anchor interacting proteins in membranes. Formation of the "trans"-SNARE complex proceeds through an intermediate complex composed of SNAP-25 and syntaxin-1, which later accommodates synaptobrevin-2 (the quoted syntaxin and synaptobrevin isotypes participate in neuronal neuromediator release).

Based on the stability of the resultant cis-SNARE complex, it has been postulated that energy released during the assembly process serves as a means for overcoming the repulsive forces between the membranes. There are several models that propose explanation of a subsequent step – the formation of stalk and fusion pore, but the exact nature of these processes remains debated. Two of the most prominent models on fusion pore formation are the lipid-lined and protein-lined fusion pore theories. [11]

Lipid-lined fusion pore theory

In the lipid-lined pore theory both membranes curve toward each other to form the early fusion pore. When the two membranes are brought to a "critical" distance, the lipid head-groups from one membrane insert into the other, creating the basis for the fusion pore. Membrane fusion via stalk formation.jpg
In the lipid-lined pore theory both membranes curve toward each other to form the early fusion pore. When the two membranes are brought to a "critical" distance, the lipid head-groups from one membrane insert into the other, creating the basis for the fusion pore.

One possible model for fusion pore formation is the lipid-line pore theory. In this model, once the membranes have been brought into sufficiently close proximity via the "zipper" mechanism of the SNARE complex, membrane fusion occurs spontaneously. It has been shown that when the two membranes are brought within a critical distance, it is possible for hydrophilic lipid headgroups of one membrane to merge with the opposing membrane. [12] In the lipid-lined fusion pore model, the SNARE complex acts as a scaffold, pulling on the membrane, causing both membranes to pucker so they may reach the critical fusion distance. As the two membranes begin to fuse, a lipid-lined stalk is produced, expanding radially outward as fusion proceeds.

While a lipid-lined pore is possible and can achieve all the same properties observed in early pore formation, sufficient data does not exist to prove it is the sole method of formation. [13] There is not currently a proposed mechanism on inter-cellular regulation for fluctuation of lipid-lined pores, and they would have a substantially more difficult time producing effects such as the "kiss-and-run" when compared with their protein-lined counterparts. Lipid-lined pores effectiveness would also be highly dependent on the composition of both membranes, and its success or failure could vary wildly with changes in elasticity and rigidity. [13]

Protein-lined fusion pore theory

Another possible model for fusion pore formation is the protein-lined pore theory. In this model, after activation of synaptotagmin by calcium, several SNARE complexes come together to form a ring structure, with synaptobrevin forming the pore in the vesicle membrane and Syntaxin forming the pore in the cell membrane. [14] As the initial pore expands it incorporates lipids from both bilayers, eventually resulting in complete fusion of the two membranes. The SNARE complex has a much more active role in the protein-lined pore theory; because the pore consists initially entirely of SNARE proteins, the pore is easily able to undergo intercellular regulation, making fluctuation and "kiss-and-run" mechanisms easily attainable. [9]

A protein-lined pore perfectly meets all the observed requirements of the early fusion pore, and while some data does support this theory, [14] sufficient data does not exist to pronounce it the primary method of fusion. A protein-lined pore requires at least five copies of the SNARE complex while fusion has been observed with as few as two. [14]

In both theories the function of the SNARE complex remains largely unchanged, and the entire SNARE complex is necessary to initiate fusion. It has, however, been proven that in vitro Syntaxin per se is sufficient to drive spontaneous calcium independent fusion of synaptic vesicles containing v-SNAREs. [15] This suggests that in Ca2+-dependent neuronal exocytosis synaptotagmin is a dual regulator, in absence of Ca2+ ions to inhibit SNARE dynamics, while in presence of Ca2+ ions to act as agonist in the membrane fusion process.

Kiss-and-run hypothesis

In synaptic vesicles, some neurochemists have suggested that vesicles occasionally may not completely fuse with presynaptic membranes in neurotransmitter release into the synaptic cleft. The controversy lies in whether or not endocytosis always occurs in vesicle reforming after release of the neurotransmitter. Another proposed mechanism for release of vesicle contents into extracellular fluid is called kiss-and-run fusion.

There is some indication that vesicles may only form a small pore in the presynaptic membrane allowing contents to be released by standard diffusion for a short while before retreating back into the presynaptic cell. This mechanism may be a way around clathrin-mediated endocytosis. It is also proposed that the vesicle does not need to return to an endosome to refill, though it is not thoroughly understood by which mechanism it would refill. This does not exclude full vesicle fusion, but only states that both mechanisms may operate in synaptic clefts.

"Kiss and run" has been shown to occur in endocrine cells, though it has not been directly witnessed in synaptic gaps. [16]

See also

Related Research Articles

<span class="mw-page-title-main">Exocytosis</span> Active transport and bulk transport in which a cell transports molecules out of the cell

Exocytosis is a form of active transport and bulk transport in which a cell transports molecules out of the cell. As an active transport mechanism, exocytosis requires the use of energy to transport material. Exocytosis and its counterpart, endocytosis, are used by all cells because most chemical substances important to them are large polar molecules that cannot pass through the hydrophobic portion of the cell membrane by passive means. Exocytosis is the process by which a large amount of molecules are released; thus it is a form of bulk transport. Exocytosis occurs via secretory portals at the cell plasma membrane called porosomes. Porosomes are permanent cup-shaped lipoprotein structures at the cell plasma membrane, where secretory vesicles transiently dock and fuse to release intra-vesicular contents from the cell.

<span class="mw-page-title-main">Synaptic vesicle</span> Neurotransmitters that are released at the synapse

In a neuron, synaptic vesicles store various neurotransmitters that are released at the synapse. The release is regulated by a voltage-dependent calcium channel. Vesicles are essential for propagating nerve impulses between neurons and are constantly recreated by the cell. The area in the axon that holds groups of vesicles is an axon terminal or "terminal bouton". Up to 130 vesicles can be released per bouton over a ten-minute period of stimulation at 0.2 Hz. In the visual cortex of the human brain, synaptic vesicles have an average diameter of 39.5 nanometers (nm) with a standard deviation of 5.1 nm.

<span class="mw-page-title-main">SNARE protein</span> Protein family

SNARE proteins – "SNAPREceptors" – are a large protein family consisting of at least 24 members in yeasts, more than 60 members in mammalian cells, and some numbers in plants. The primary role of SNARE proteins is to mediate the fusion of vesicles with the target membrane; this notably mediates exocytosis, but can also mediate the fusion of vesicles with membrane-bound compartments. The best studied SNAREs are those that mediate the release of synaptic vesicles containing neurotransmitters in neurons. These neuronal SNAREs are the targets of the neurotoxins responsible for botulism and tetanus produced by certain bacteria.

<span class="mw-page-title-main">Synaptobrevin</span>

Synaptobrevins are small integral membrane proteins of secretory vesicles with molecular weight of 18 kilodalton (kDa) that are part of the vesicle-associated membrane protein (VAMP) family.

<span class="mw-page-title-main">SNAP25</span> Protein-coding gene in the species Homo sapiens

Synaptosomal-Associated Protein, 25kDa (SNAP-25) is a Target Soluble NSF (N-ethylmaleimide-sensitive factor) Attachment Protein Receptor (t-SNARE) protein encoded by the SNAP25 gene found on chromosome 20p12.2 in humans. SNAP-25 is a component of the trans-SNARE complex, which accounts for membrane fusion specificity and directly executes fusion by forming a tight complex that brings the synaptic vesicle and plasma membranes together.

<span class="mw-page-title-main">Synaptotagmin</span>

Synaptotagmins (SYTs) constitute a family of membrane-trafficking proteins that are characterized by an N-terminal transmembrane region (TMR), a variable linker, and two C-terminal C2 domains - C2A and C2B. There are 17 isoforms in the mammalian synaptotagmin family. There are several C2-domain containing protein families that are related to synaptotagmins, including transmembrane (Ferlins, Extended-Synaptotagmin (E-Syt) membrane proteins, and MCTPs) and soluble (RIMS1 and RIMS2, UNC13D, synaptotagmin-related proteins and B/K) proteins. The family includes synaptotagmin 1, a Ca2+ sensor in the membrane of the pre-synaptic axon terminal, coded by gene SYT1.

<span class="mw-page-title-main">Complexin</span>

Complexin (also known as synaphin) refers to a one of a small set of eukaryotic cytoplasmic neuronal proteins which binds to the SNARE protein complex (SNAREpin) with a high affinity. These are called synaphin 1 and 2. In the presence of Ca2+, the transport vesicle protein synaptotagmin displaces complexin, allowing the SNARE protein complex to bind the transport vesicle to the presynaptic membrane.

<span class="mw-page-title-main">STX1A</span> Protein-coding gene in the species Homo sapiens

Syntaxin-1A is a protein that in humans is encoded by the STX1A gene.

<span class="mw-page-title-main">VAMP2</span> Protein-coding gene in the species Homo sapiens

Vesicle-associated membrane protein 2 (VAMP2) is a protein that in humans is encoded by the VAMP2 gene.

<span class="mw-page-title-main">NAPA (gene)</span> Protein-coding gene in the species Homo sapiens

N-ethylmaleimide-sensitive factor Attachment Protein Alpha, also known as SNAP-α, is a SNAP protein that is involved in the intra-cellular trafficking and fusing of vesicles to target membranes in cells.

<span class="mw-page-title-main">Vesicle-associated membrane protein 8</span> Protein-coding gene in the species Homo sapiens

Vesicle-associated membrane protein 8 is a protein that in humans is encoded by the VAMP8 gene.

The ribbon synapse is a type of neuronal synapse characterized by the presence of an electron-dense structure, the synaptic ribbon, that holds vesicles close to the active zone. It is characterized by a tight vesicle-calcium channel coupling that promotes rapid neurotransmitter release and sustained signal transmission. Ribbon synapses undergo a cycle of exocytosis and endocytosis in response to graded changes of membrane potential. It has been proposed that most ribbon synapses undergo a special type of exocytosis based on coordinated multivesicular release. This interpretation has recently been questioned at the inner hair cell ribbon synapse, where it has been instead proposed that exocytosis is described by uniquantal release shaped by a flickering vesicle fusion pore.

<span class="mw-page-title-main">Syntaxin</span> Group of proteins

Syntaxins are a family of membrane integrated Q-SNARE proteins participating in exocytosis.

Munc-18 proteins are the mammalian homologue of UNC-18 and are a member of the Sec1/Munc18-like (SM) protein family. Munc-18 proteins have been identified as essential components of the synaptic vesicle fusion protein complex and are crucial for the regulated exocytosis of neurons and neuroendocrine cells.

<span class="mw-page-title-main">Active zone</span>

The active zone or synaptic active zone is a term first used by Couteaux and Pecot-Dechavassinein in 1970 to define the site of neurotransmitter release. Two neurons make near contact through structures called synapses allowing them to communicate with each other. As shown in the adjacent diagram, a synapse consists of the presynaptic bouton of one neuron which stores vesicles containing neurotransmitter, and a second, postsynaptic neuron which bears receptors for the neurotransmitter, together with a gap between the two called the synaptic cleft. When an action potential reaches the presynaptic bouton, the contents of the vesicles are released into the synaptic cleft and the released neurotransmitter travels across the cleft to the postsynaptic neuron and activates the receptors on the postsynaptic membrane.

Kiss-and-run fusion is a type of synaptic vesicle release where the vesicle opens and closes transiently. In this form of exocytosis, the vesicle docks and transiently fuses at the presynaptic membrane and releases its neurotransmitters across the synapse, after which the vesicle can then be reused.

Membrane vesicle trafficking in eukaryotic animal cells involves movement of biochemical signal molecules from synthesis-and-packaging locations in the Golgi body to specific release locations on the inside of the plasma membrane of the secretory cell. It takes place in the form of Golgi membrane-bound micro-sized vesicles, termed membrane vesicles (MVs).

<span class="mw-page-title-main">Soluble NSF attachment protein</span> Protein family

Soluble N-ethylmaleimide-Sensitive Factor Attachment Proteins are a family of cytosolic adaptor proteins involved in vesicular fusion at membranes during intracellular transport and exocytosis. SNAPs interact with proteins of the SNARE complex and NSF to play a key role in recycling the components of the fusion complex. SNAPs are involved in the priming of the vesicle fusion complex during assembly, as well as in the disassembly following a vesicle fusion event. Following membrane fusion, the tethering SNARE proteins complex disassembles in response to steric changes originating from the ATPase NSF. The energy provided by NSF is transferred throughout the SNARE complex and SNAP, allowing the proteins to untangle, and recycled for future fusion events. Mammals have three SNAP genes: α-SNAP, β-SNAP, and γ-SNAP. α- and γ-SNAP are expressed throughout the body, while β-SNAP is specific to the brain. The yeast homolog of the human SNAP is Sec17, the structural diagram of which is included on this page.

Zero ionic layer is the main site of interaction in the core SNARE complex. Dipole-dipole interactions take place between 3 glutamine (Q) residues and 1 arginine (R) residue exposed in this layer. Despite that, the majority of the SNARE complex is hydrophobic because of the leucine zipper. Extensively studied layers within the SNARE alpha-helical bundle are designated from "-7" to "+8". Zero ionic layer is at the center of the bundle, and thus designated as "0" layer.

Edwin R. Chapman is an American biochemist known for his work on Ca2+-triggered exocytosis. He currently serves as the Ricardo Miledi Professor of Neuroscience at the University of Wisconsin–Madison, where he is also an investigator of the Howard Hughes Medical Institute (HHMI).

References

  1. 1 2 3 Page 237 in: Costanzo, Linda S. (2007). Physiology . Hagerstwon, MD: Lippincott Williams & Wilkins. ISBN   978-0-7817-7311-9.
  2. ↑ Walter F., PhD. Boron (2003). Medical Physiology: A Cellular And Molecular Approaoch. Elsevier/Saunders. p. 1300. ISBN   978-1-4160-2328-9.
  3. ↑ Papahadjopoulos, Demetrios (1990). "Molecular mechanisms of calcium-induced membrane fusion". Journal of Bioenergetics and Biomembranes. 22 (2): 157–179. doi:10.1007/BF00762944. PMID   2139437. S2CID   1465571.
  4. ↑ Ortiz, Antonio; Killian, J. Antoinette; Verkleij, Arie J.; Wilschut, Jan (October 1999). "Membrane Fusion and the Lamellar-to-Inverted-Hexagonal Phase Transition in Cardiolipin Vesicle Systems Induced by Divalent Cations". Biophysical Journal. 77 (4): 2003–2014. Bibcode:1999BpJ....77.2003O. doi:10.1016/S0006-3495(99)77041-4. PMC   1300481 . PMID   10512820.
  5. ↑ Pigino, Gustavo; Morfini, Gerardo; Brady, Scott (2006). "Chapter 9: Intracellular Trafficking". In Siegal, George J.; Albers, R. Wayne; Brady, Scott T.; et al. (eds.). Basic Neurochemistry: Molecular, Cellular and Medical Aspects (Textbook) (7th ed.). Burlington, MA: Elsevier Academic Press. p. 143. ISBN   978-0-12-088397-4.
  6. ↑ Pigino et al. p 158
  7. ↑ Pigino et al. p.143
  8. ↑ Georgiev, Danko D .; James F . Glazebrook (2007). "Subneuronal processing of information by solitary waves and stochastic processes". In Lyshevski, Sergey Edward (ed.). Nano and Molecular Electronics Handbook. Nano and Microengineering Series. CRC Press. pp. 17–1–17–41. doi:10.1201/9781315221670-17. ISBN   978-0-8493-8528-5. S2CID   199021983.
  9. 1 2 Kümmel, D.; Krishnakumar, S. S.; Radoff, D. T.; Li, F.; Giraudo, C. G.; Pincet, F.; Rothman, J. E.; Reinisch, K. M. (2011). "Complexin cross-links prefusion SNAREs into a zigzag array". Nature Structural & Molecular Biology. 18 (8): 927–933. doi:10.1038/nsmb.2101. PMC   3410656 . PMID   21785414.
  10. ↑ Richmond, Janet. "Synapse Function".
  11. ↑ Jackson, Meyer B.; Chapman, Edwin R. (2006). "Fusion Pores and Fusion Machines in Ca2+-Triggered Exocytosis". Annual Review of Biophysics and Biomolecular Structure. 35 (1): 135–160. doi:10.1146/annurev.biophys.35.040405.101958. PMID   16689631.
  12. ↑ Marrink, Siewert J.; Mark, Alan E. (2003-09-01). "The Mechanism of Vesicle Fusion as Revealed by Molecular Dynamics Simulations" (PDF). Journal of the American Chemical Society. 125 (37): 11144–11145. doi:10.1021/ja036138+. ISSN   0002-7863. PMID   16220905.
  13. 1 2 Nanavati, C; Markin, V S; Oberhauser, A F; Fernandez, J M (1992-10-01). "The exocytotic fusion pore modeled as a lipidic pore". Biophysical Journal. 63 (4): 1118–1132. Bibcode:1992BpJ....63.1118N. doi:10.1016/s0006-3495(92)81679-x. ISSN   0006-3495. PMC   1262250 . PMID   1420930.
  14. 1 2 3 Chang, Che-Wei; Hui, Enfu; Bai, Jihong; Bruns, Dieter; Chapman, Edwin R.; Jackson, Meyer B. (2015-04-08). "A Structural Role for the Synaptobrevin 2 Transmembrane Domain in Dense-Core Vesicle Fusion Pores". The Journal of Neuroscience. 35 (14): 5772–5780. doi:10.1523/JNEUROSCI.3983-14.2015. ISSN   0270-6474. PMC   4388931 . PMID   25855187.
  15. ↑ Woodbury DJ, Rognlien K (2000). "The t-SNARE syntaxin is sufficient for spontaneous fusion of synaptic vesivles to planar membranes" (PDF). Cell Biology International. 24 (11): 809–818. doi:10.1006/cbir.2000.0631. PMID   11067766. S2CID   37732173. Archived from the original (PDF) on 2011-07-19. Retrieved 2009-05-31.
  16. ↑ Piginio et al. pp. 161-162
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