scholarly journals Clathrin-independent endocytic retrieval of SV proteins mediated by the clathrin adaptor AP-2 at mammalian central synapses

eLife ◽  
2022 ◽  
Vol 11 ◽  
Author(s):  
Tania López-Hernández ◽  
Koh-ichiro Takenaka ◽  
Yasunori Mori ◽  
Pornparn Kongpracha ◽  
Shushi Nagamori ◽  
...  
Keyword(s):  
Cell Surface ◽  
Synaptic Vesicles ◽  
Hippocampal Neurons ◽  
Adaptor Protein ◽  
The Reformation ◽  
Comprehensive Survey ◽  
Vesicular Transporters ◽  
Clathrin Adaptor ◽  
Central Synapses

Neurotransmission is based on the exocytic fusion of synaptic vesicles (SVs) followed by endocytic membrane retrieval and the reformation of SVs. Conflicting models have been proposed regarding the mechanisms of SV endocytosis, most notably clathrin/adaptor protein complex 2 (AP-2)-mediated endocytosis and clathrin-independent ultrafast endocytosis. Partitioning between these pathways has been suggested to be controlled by temperature and stimulus paradigm. We report on the comprehensive survey of six major SV proteins to show that SV endocytosis in mouse hippocampal neurons at physiological temperature occurs independent of clathrin while the endocytic retrieval of a subset of SV proteins including the vesicular transporters for glutamate and GABA depend on sorting by the clathrin adaptor AP-2. Our findings highlight a clathrin-independent role of the clathrin adaptor AP-2 in the endocytic retrieval of select SV cargos from the presynaptic cell surface and suggest a revised model for the endocytosis of SV membranes at mammalian central synapses.

scholarly journals Clathrin-independent endocytic retrieval of SV proteins mediated by the clathrin adaptor AP-2 at mammalian central synapses

2021 ◽  
Author(s):  
Tania Lopez-Hernandez ◽  
Koh-ichiro Takenaka ◽  
Yasunori Mori ◽  
Pornparn Kongpracha ◽  
Shushi Nagamori ◽  
...  
Keyword(s):  
Cell Surface ◽  
Synaptic Vesicles ◽  
Hippocampal Neurons ◽  
Physiological Temperature ◽  
The Reformation ◽  
Comprehensive Survey ◽  
Vesicular Transporters ◽  
Clathrin Adaptor ◽  
Central Synapses

Neurotransmission is based on the exocytic fusion of synaptic vesicles (SVs) followed by endocytic membrane retrieval and the reformation of SVs. Conflicting models have been proposed regarding the mechanisms of SV endocytosis, most notably clathrin/ AP-2-mediated endocytosis and clathrin-independent ultrafast endocytosis. Partitioning between these pathways has been suggested to be controlled by temperature and stimulus paradigm. We report on the comprehensive survey of six major SV proteins to show that SV endocytosis in hippocampal neurons at physiological temperature occurs independent of clathrin while the endocytic retrieval of a subset of SV proteins including the vesicular transporters for glutamate and GABA depend on sorting by the clathrin adaptor AP-2. Our findings highlight a clathrin-independent role of the clathrin adaptor AP-2 in the endocytic retrieval of select SV cargos from the presynaptic cell surface and suggest a unified model for the endocytosis of SV membranes at mammalian central synapses.

scholarly journals The AP2 clathrin adaptor protein complex regulates the abundance of GLR-1 glutamate receptors in the ventral nerve cord of Caenorhabditis elegans

2015 ◽  
Vol 26 (10) ◽  
pp. 1887-1900 ◽  
Author(s):  
Steven D. Garafalo ◽  
Eric S. Luth ◽  
Benjamin J. Moss ◽  
Michael I. Monteiro ◽  
Emily Malkin ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Nerve Cord ◽  
Ventral Nerve Cord ◽  
Secretory Pathway ◽  
Adaptor Protein ◽  
Intracellular Compartments ◽  
Clathrin Adaptor ◽  
Strength And Plasticity

Regulation of glutamate receptor (GluR) abundance at synapses by clathrin-mediated endocytosis can control synaptic strength and plasticity. We take advantage of viable, null mutations in subunits of the clathrin adaptor protein 2 (AP2) complex in Caenorhabditis elegans to characterize the in vivo role of AP2 in GluR trafficking. In contrast to our predictions for an endocytic adaptor, we found that levels of the GluR GLR-1 are decreased at synapses in the ventral nerve cord (VNC) of animals with mutations in the AP2 subunits APM-2/μ2, APA-2/α, or APS-2/σ2. Rescue experiments indicate that APM-2/μ2 functions in glr-1–expressing interneurons and the mature nervous system to promote GLR-1 levels in the VNC. Genetic analyses suggest that APM-2/μ2 acts upstream of GLR-1 endocytosis in the VNC. Consistent with this, GLR-1 accumulates in cell bodies of apm-2 mutants. However, GLR-1 does not appear to accumulate at the plasma membrane of the cell body as expected, but instead accumulates in intracellular compartments including Syntaxin-13– and RAB-14–labeled endosomes. This study reveals a novel role for the AP2 clathrin adaptor in promoting the abundance of GluRs at synapses in vivo, and implicates AP2 in the regulation of GluR trafficking at an early step in the secretory pathway.

scholarly journals An essential role of acetylcholine-glutamate synergy at habenular synapses in nicotine dependence

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Silke Frahm ◽  
Beatriz Antolin-Fontes ◽  
Andreas Görlich ◽  
Johannes-Friedrich Zander ◽  
Gudrun Ahnert-Hilger ◽  
...  
Keyword(s):  
Synaptic Vesicles ◽  
Critical Role ◽  
Conditioned Reward ◽  
Quantal Size ◽  
Medial Habenula ◽  
Vesicular Transporters ◽  
Circuit Function ◽  
And Behavior ◽  
Presynaptic Facilitation

A great deal of interest has been focused recently on the habenula and its critical role in aversion, negative-reward and drug dependence. Using a conditional mouse model of the ACh-synthesizing enzyme choline acetyltransferase (Chat), we report that local elimination of acetylcholine (ACh) in medial habenula (MHb) neurons alters glutamate corelease and presynaptic facilitation. Electron microscopy and immuno-isolation analyses revealed colocalization of ACh and glutamate vesicular transporters in synaptic vesicles (SVs) in the central IPN. Glutamate reuptake in SVs prepared from the IPN was increased by ACh, indicating vesicular synergy. Mice lacking CHAT in habenular neurons were insensitive to nicotine-conditioned reward and withdrawal. These data demonstrate that ACh controls the quantal size and release frequency of glutamate at habenular synapses, and suggest that the synergistic functions of ACh and glutamate may be generally important for modulation of cholinergic circuit function and behavior.

scholarly journals Ent3p and Ent5p Exhibit Cargo-specific Functions in Trafficking Proteins between the Trans-Golgi Network and the Endosomes in Yeast

2007 ◽  
Vol 18 (5) ◽  
pp. 1803-1815 ◽  
Author(s):  
Alenka Čopič ◽  
Trevor L. Starr ◽  
Randy Schekman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Surface ◽  
Protein Transport ◽  
Adaptor Protein ◽  
Additional Function ◽  
Trans Golgi Network ◽  
Cargo Proteins ◽  
Clathrin Adaptor ◽  
Dependent Transport ◽  
Golgi Network

The phosphoinositide-binding proteins Ent3p and Ent5p are required for protein transport from the trans-Golgi network (TGN) to the vacuole in Saccharomyces cerevisiae. Both proteins interact with the monomeric clathrin adaptor Gga2p, but Ent5p also interacts with the clathrin adaptor protein 1 (AP-1) complex, which facilitates retention of proteins such as Chs3p at the TGN. When both ENT3 and ENT5 are mutated, Chs3p is diverted from an intracellular reservoir to the cell surface. However, Ent3p and Ent5p are not required for the function of AP-1, but rather they seem to act in parallel with AP-1 to retain proteins such as Chs3p at the TGN. They have all the properties of clathrin adaptors, because they can both bind to clathrin and to cargo proteins. Like AP-1, Ent5p binds to Chs3p, whereas Ent3p facilitates the interaction between Gga2p and the endosomal syntaxin Pep12p. Thus, Ent3p has an additional function in Gga-dependent transport to the late endosome. Ent3p also facilitates the association between Gga2p and clathrin; however, Ent5p can partially substitute for this function. We conclude that the clathrin adaptors AP-1, Ent3p, Ent5p, and the Ggas cooperate in different ways to sort proteins between the TGN and the endosomes.

scholarly journals Defective function of GABA-containing synaptic vesicles in mice lacking the AP-3B clathrin adaptor

2004 ◽  
Vol 167 (2) ◽  
pp. 293-302 ◽  
Author(s):  
Fubito Nakatsu ◽  
Motohiro Okada ◽  
Fumiaki Mori ◽  
Noriko Kumazawa ◽  
Hiroto Iwasa ◽  
...  
Keyword(s):  
Synaptic Vesicles ◽  
Neuronal Excitability ◽  
Critical Role ◽  
Physiological Role ◽  
Adaptor Protein ◽  
Epileptic Seizures ◽  
Gaba Release ◽  
Long Term Potentiation ◽  
Clathrin Adaptor ◽  
And Function

AP-3 is a member of the adaptor protein (AP) complex family that regulates the vesicular transport of cargo proteins in the secretory and endocytic pathways. There are two isoforms of AP-3: the ubiquitously expressed AP-3A and the neuron-specific AP-3B. Although the physiological role of AP-3A has recently been elucidated, that of AP-3B remains unsolved. To address this question, we generated mice lacking μ3B, a subunit of AP-3B. μ3B−/− mice suffered from spontaneous epileptic seizures. Morphological abnormalities were observed at synapses in these mice. Biochemical studies demonstrated the impairment of γ-aminobutyric acid (GABA) release because of, at least in part, the reduction of vesicular GABA transporter in μ3B−/− mice. This facilitated the induction of long-term potentiation in the hippocampus and the abnormal propagation of neuronal excitability via the temporoammonic pathway. Thus, AP-3B plays a critical role in the normal formation and function of a subset of synaptic vesicles. This work adds a new aspect to the pathogenesis of epilepsy.

Synthesis, Packaging, and Termination of Neurotransmitters

2017 ◽  
Author(s):  
Peggy Mason
Keyword(s):  
Molecular Weight ◽  
Synaptic Vesicles ◽  
Drugs Of Abuse ◽  
Low Molecular Weight ◽  
Stiff Person Syndrome ◽  
Catecholamine Synthesis ◽  
Vesicular Transporters ◽  
Psychotropic Effects ◽  
Gaba Synthesis

The synthesis, packaging, and termination of action of neurotransmitters are detailed. There are far more varieties of peptide neurotransmitters than there are of low-molecular-weight neurotransmitters. Yet low-molecular-weight neurotransmitters are the ubiquitous workhorses of the nervous system. Acetylcholine, the catecholamines norepinephrine and dopamine, serotonin, glutamate, and GABA are examined in some depth. The vesicular transporters that carry low-molecular-weight neurotransmitters from the cytoplasm into synaptic vesicles are covered. The role of monoamines in affect and mood and the psychotropic effects of monoaminergic drugs are discussed. Principles of catecholamine synthesis are applied to understand phenylketonuria. Uptake of monoamines into neurons is discussed in the context of amphetamine, cocaine, and other drugs of abuse. Stiff-person syndrome, which results from an impairment of GABA synthesis, is introduced. The modes of action for peptide and gaseous neurotransmitters are briefly covered.

scholarly journals Epithelial sodium channel (ENaC) is multi-ubiquitinated at the cell surface

2007 ◽  
Vol 405 (1) ◽  
pp. 147-155 ◽  
Author(s):  
Dominik Wiemuth ◽  
Ying Ke ◽  
Meino Rohlfs ◽  
Fiona J. Mc Donald
Keyword(s):  
Cell Surface ◽  
Sodium Channel ◽  
Adaptor Protein ◽  
Epithelial Sodium Channel ◽  
Active Sodium ◽  
Rate Limiting Step ◽  
Clathrin Adaptor ◽  
Neuronal Precursor Cell ◽  
Gene 4 ◽  
Protein Ligases

The human ENaC (epithelial sodium channel), a complex of three subunits, provides the rate-limiting step for sodium uptake in the distal nephron, and therefore plays a key role in salt homoeostasis and in regulating blood pressure. The number of active sodium channel complexes present at the plasma membrane appears to be tightly controlled. In Liddle's syndrome, a form of hypertension caused by an increase in the number of active sodium channels at the cell membrane, the βENaC or γENaC subunit gene contains a mutation that disrupts the binding site for the Nedd4 (neuronal precursor cell expressed developmentally down-regulated gene 4) family of ubiquitin-protein ligases. Therefore ubiquitination of channel subunits may be involved in altering cell surface ENaC. Here, we provide evidence that the ENaC subunits located at the cell surface are modified with multiple mono-ubiquitins (multi-ubiquitination) and that Nedd4-2 modulates this ubiquitination. We confirm that ENaC is associated with the μ2 subunit of the AP-2 (adaptor protein 2) clathrin adaptor. Since mono- or multi-ubiquitination of other membrane proteins is a signal for their internalization by clathrin-mediated endocytosis and subsequent trafficking, our results support a model whereby ubiquitin and clathrin adaptor binding sites act in concert to remove ENaC from the cell surface.

scholarly journals UNC-11, a Caenorhabditis elegans AP180 Homologue, Regulates the Size and Protein Composition of Synaptic Vesicles

1999 ◽  
Vol 10 (7) ◽  
pp. 2343-2360 ◽  
Author(s):  
Michael L. Nonet ◽  
Andrea M. Holgado ◽  
Faraha Brewer ◽  
Craig J. Serpe ◽  
Betty A. Norbeck ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Synaptic Vesicle ◽  
Neurotransmitter Release ◽  
Synaptic Vesicles ◽  
Adaptor Protein ◽  
Protein Composition ◽  
Mammalian Brain ◽  
Snare Protein ◽  
Vesicle Biogenesis ◽  
Clathrin Adaptor

The unc-11 gene of Caenorhabditis elegans encodes multiple isoforms of a protein homologous to the mammalian brain-specific clathrin-adaptor protein AP180. The UNC-11 protein is expressed at high levels in the nervous system and at lower levels in other tissues. In neurons, UNC-11 is enriched at presynaptic terminals but is also present in cell bodies. unc-11mutants are defective in two aspects of synaptic vesicle biogenesis. First, the SNARE protein synaptobrevin is mislocalized, no longer being exclusively localized to synaptic vesicles. The reduction of synaptobrevin at synaptic vesicles is the probable cause of the reduced neurotransmitter release observed in these mutants. Second,unc-11 mutants accumulate large vesicles at synapses. We propose that the UNC-11 protein mediates two functions during synaptic vesicle biogenesis: it recruits synaptobrevin to synaptic vesicle membranes and it regulates the size of the budded vesicle during clathrin coat assembly.

scholarly journals The Yeast Adaptor Protein Complex, AP-3, Is Essential for the Efficient Delivery of Alkaline Phosphatase by the Alternate Pathway to the Vacuole

1997 ◽  
Vol 139 (7) ◽  
pp. 1761-1774 ◽  
Author(s):  
J. David Stepp ◽  
Kristen Huang ◽  
Sandra K. Lemmon
Keyword(s):  
Alkaline Phosphatase ◽  
Adaptor Protein ◽  
Carboxypeptidase Y ◽  
Loss Of Function ◽  
Synthetic Lethal ◽  
Efficient Delivery ◽  
Cargo Proteins ◽  
Clathrin Adaptor ◽  
Class E

A novel clathrin adaptor-like complex, adaptor protein (AP)-3, has recently been described in yeast and in animals. To gain insight into the role of yeast AP-3, a genetic strategy was devised to isolate gene products that are required in the absence of the AP-3 μ chain encoded by APM3. One gene identified by this synthetic lethal screen was VPS45. The Vps pathway defines the route that several proteins, including carboxypeptidase Y, take from the late Golgi to the vacuole. However, vacuolar alkaline phosphatase (ALP) is transported via an alternate, intracellular route. This suggested that the apm3-Δ vps45 synthetic phenotype could be caused by a block in both the alternate and the Vps pathways. Here we demonstrate that loss of function of the AP-3 complex results in slowed processing and missorting of ALP. ALP is no longer localized to the vacuole membrane by immunofluorescence, but is found in small punctate structures throughout the cell. This pattern is distinct from the Golgi marker Kex2p, which is unaffected in AP-3 mutants. We also show that in the apm3-Δ mutant some ALP is delivered to the vacuole by diversion into the Vps pathway. Class E vps mutants accumulate an exaggerated prevacuolar compartment containing membrane proteins on their way to the vacuole or destined for recycling to the Golgi. Surprisingly, in AP-3 class E vps double mutants these proteins reappear on the vacuole. We suggest that some AP-3–dependent cargo proteins that regulate late steps in Golgi to vacuole transport are diverted into the Vps pathway allowing completion of transfer to the vacuole in the class E vps mutant.

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