The Adhesion Molecule CHL1 Regulates Uncoating of Clathrin-Coated Synaptic Vesicles

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Background: Cell adhesion molecule CHL1 plays a dual role by either promoting or inhibiting neuritogenesis. Results: Ligand-induced clustering of CHL1 at the cell surface induces lipid raft-dependent endocytosis of CHL1 and neuritogenesis. Conclusion: Ligand-induced remodeling of CHL1 adhesion promotes neurite outgrowth. Significance: High levels of CHL1 adhesion and inhibition of CHL1 endocytosis can interfere with neuronal development and/or regeneration.

Section: Methodsmentioning

confidence: 99%

Section: Cysteine 1102-dependent Targeting Of Chl1 To Lipid Rafts Promentioning

confidence: 99%

Lipid Raft-dependent Endocytosis of Close Homolog of Adhesion Molecule L1 (CHL1) Promotes Neuritogenesis

Tian

Leshchyns’ka

Welch

et al. 2012

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“…Likewise, an increasing number of proteins that influence brain function have been identified to interact with heat shock proteins. In presynaptic terminals, Hsc70 forms a complex with cysteine string protein and small glutamine-rich tetratricopeptide repeat domain protein that plays an important role in synaptic vesicle uncoating during clathrindependent endocytosis (25)(26)(27). Hsc70 also binds gephyrin, induces gephyrin clustering, and regulates the formation of tertiary complexes, including GABA and glycine receptors, at inhibitory synapses (28).…”

Section: Discussionmentioning

confidence: 99%

The Molecular Chaperone Hsc70 Interacts with Tyrosine Hydroxylase to Regulate Enzyme Activity and Synaptic Vesicle Localization

Parra¹,

Baust²,

Smith

et al. 2016

We previously reported that the vesicular monoamine transporter 2 (VMAT2) is physically and functionally coupled with Hsc70 as well as with the dopamine synthesis enzymes tyrosine hydroxylase (TH) and aromatic amino acid decarboxylase, providing a novel mechanism for dopamine homeostasis regulation. Here we expand those findings to demonstrate that Hsc70 physically and functionally interacts with TH to regulate the enzyme activity and synaptic vesicle targeting. Co-immunoprecipitation assays performed in brain tissue and heterologous cells demonstrated that Hsc70 interacts with TH and aromatic amino acid decarboxylase. Furthermore, in vitro binding assays showed that TH directly binds the substrate binding and carboxyl-terminal domains of Hsc70. Immunocytochemical studies indicated that Hsc70 and TH co-localize in midbrain dopaminergic neurons. The functional significance of the Hsc70-TH interaction was then investigated using TH activity assays. In both dopaminergic MN9D cells and mouse brain synaptic vesicles, purified Hsc70 facilitated an increase in TH activity. Neither the closely related protein Hsp70 nor the unrelated Hsp60 altered TH activity, confirming the specificity of the Hsc70 effect. Overexpression of Hsc70 in dopaminergic MN9D cells consistently resulted in increased TH activity whereas knockdown of Hsc70 by short hairpin RNA resulted in decreased TH activity and dopamine levels. Finally, in cells with reduced levels of Hsc70, the amount of TH associated with synaptic vesicles was decreased. This effect was rescued by addition of purified Hsc70. Together, these data demonstrate a novel interaction between Hsc70 and TH that regulates the activity and localization of the enzyme to synaptic vesicles, suggesting an important role for Hsc70 in dopamine homeostasis.Dopaminergic neurons within the substantia nigra and ventral tegmental area are the primary sources of the catecholamine neurotransmitter dopamine (DA).2 Despite the fact that dopaminergic neurons account for less than 0.01% of all neurons, they play a significant role in brain function (1-3). Consequently, DA homeostasis is crucial for the preservation and regulation of physiological functions such as locomotion, cognition, neuroendocrine secretion, and motivated behaviors (4). Thus, it is not surprising that disruptions in the DA system have been implicated in several neurological and psychiatric disorders, including Parkinson disease, depression, schizophrenia, attention deficit hyperactivity disorder, Tourette syndrome, and drug addiction (4 -8).The DA life cycle consists of a series of highly regulated molecular events that are ultimately responsible for controlling DA homeostasis. Synthesis of DA occurs in the presynaptic terminals via two enzymatic reactions. First, tyrosine is converted into L-3,4-dihydroxyphenylalanine through the actions of the rate-limiting enzyme tyrosine hydroxylase (TH) (9, 10). Subsequently, aromatic amino acid decarboxylase (AADC) converts L-DOPA into DA (11). When synthesized, DA is packaged and stored withi...

“…Adhesion molecules are indispensable components of synapses that play a role not only in synapse formation and stabilization but also in regulation of synapse functioning after maturation (4,6,33,34,36). The neural cell adhesion molecule NCAM and, in particular, its largest NCAM180 isoform, is one of such components (14,25).…”

Section: Discussionmentioning

confidence: 99%

“…To identify postsynaptic regions in NCAM180GFP transfected neurons, synaptic boutons of hippocampal neurons maintained in culture for 2 weeks were loaded with the lipophilic dye FM4-64 by a brief incubation in high K ϩ solution (47 mM K ϩ , 90 s) in the presence of the dye (12,25,33,34). NCAM180GFP accumulations in dendrites apposed to FM4-64 clusters were then identified as postsynaptic ( Fig.…”

Section: Ncam180gfp Fluorescence Recovers Nonuniformly In the Postsynmentioning

confidence: 99%

Immobilized Pool of NCAM180 in the Postsynaptic Membrane Is Homeostatically Replenished by the Flux of NCAM180 from Extrasynaptic Regions

Leshchyns’ka

Tanaka

Schachner

et al. 2011

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Homeostatic mechanisms maintaining high levels of adhesion molecules in synapses over prolonged periods of time remain incompletely understood. We used fluorescence recovery after photobleaching experiments to analyze the steady state turnover of the immobile pool of green fluorescent protein-labeled NCAM180, the largest postsynaptically accumulating isoform of the neural cell adhesion molecule (NCAM). We show that there is a continuous flux of NCAM180 to the postsynaptic membrane from nonsynaptic regions of dendrites by diffusion. In the postsynaptic membrane, the newly delivered NCAM180 slowly intermixes with the immobilized pool of NCAM180. Preferential immobilization and accumulation of NCAM180 in the postsynaptic membrane is reduced after disruption of the association of NCAM180 with the spectrin cytoskeleton and in the absence of the homophilic interactions of NCAM180 in synapses. Our observations indicate that the homophilic interactions and binding to the cytoskeleton promote immobilization of NCAM180 and its accumulation in the postsynaptic membrane. Flux of NCAM180 from extrasynaptic regions and its slow intermixture with the immobile pool of NCAM180 in the postsynaptic membrane may be important for the continuous homeostatic replenishment of NCAM180 protein at synaptic contacts without compromising the long term synaptic contact stability.Neurons are assembled into circuits via specialized contacts, called synapses, that are essential for information processing, learning, and storage in the brain. Synapses are stable long lasting structures (1-3). This stability is at least partially rendered by a number of cell adhesion molecules accumulating in the pre-and postsynaptic membranes (4 -9). During development, cell adhesion molecules are delivered to nascent synapses in intracellular organelles or as cell surface aggregates (10 -12). A similar type of quantal delivery of adhesion molecules has been also observed during induction of long term potentiation (13). Long lasting stability of synapses (months or years), however, suggests that synaptic pools of cell adhesion molecules (protein life time hours or days) have to be also homeostatically and continuously replenished over prolonged periods of time without compromising synapse stability and composition. Mechanisms of such replenishment remain poorly understood.Among synaptic adhesion molecules, the neural cell adhesion molecule (NCAM), 2 a member of the immunoglobulin superfamily of adhesion molecules, was shown to be involved in mechanical stabilization of interneuronal contacts and their transformation into synapses (12,14). In mice, deficiency in NCAM results in impaired long term potentiation, increased aggression and anxiety, deficits in spatial learning, and exploratory behavior (15-21). In humans, genetic variations in the NCAM gene have been reported to confer risk factors associated with bipolar affective disorders and schizophrenia (22)(23)(24). The largest isoform of NCAM, NCAM180, accumulates in the postsynaptic membrane and recruits the s...