Extracellular Signal-regulated Kinase and Glycogen Synthase Kinase 3β Regulate Gephyrin Postsynaptic Aggregation and GABAergic Synaptic Function in a Calpain-dependent Mechanism

Tyagarajan, Shiva K.; Ghosh, Himanish; Yévenes, Gonzalo E.; Imanishi, Susumu Y.; Zeilhofer, Hanns Ulrich; Gerrits, Bertran; Fritschy, Jean‐Marc

doi:10.1074/jbc.m112.442616

Cited by 93 publications

(150 citation statements)

References 30 publications

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“…The critical involvement of gephyrin in these mechanisms is consistent with several recent data highlighting its implication in the clustering of GABA A Rs. Phosphorylation of gephyrin on Ser270 by glycogen synthase kinase 3β (GSK3β) or on Ser268 by ERK have been shown to modulate the density and size of gephyrin clusters (8,31). Also gephyrin can interact with various partners, including neuroligin 2, collibystin, or even Cdc42 to regulate cluster formation and GABA A R aggregation (7,9).…”

Section: Discussionmentioning

confidence: 99%

Activity-dependent inhibitory synapse remodeling through gephyrin phosphorylation

Flores

Nikonenko

Méndez

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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113

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Maintaining a proper balance between excitation and inhibition is essential for the functioning of neuronal networks. However, little is known about the mechanisms through which excitatory activity can affect inhibitory synapse plasticity. Here we used tagged gephyrin, one of the main scaffolding proteins of the postsynaptic density at GABAergic synapses, to monitor the activity-dependent adaptation of perisomatic inhibitory synapses over prolonged periods of time in hippocampal slice cultures. We find that learning-related activity patterns known to induce N-methyl-daspartate (NMDA) receptor-dependent long-term potentiation and transient optogenetic activation of single neurons induce within hours a robust increase in the formation and size of gephyrin-tagged clusters at inhibitory synapses identified by correlated confocal electron microscopy. This inhibitory morphological plasticity was associated with an increase in spontaneous inhibitory activity but did not require activation of GABA A receptors. Importantly, this activity-dependent inhibitory plasticity was prevented by pharmacological blockade of Ca 2+ /calmodulindependent protein kinase II (CaMKII), it was associated with an increased phosphorylation of gephyrin on a site targeted by CaMKII, and could be prevented or mimicked by gephyrin phosphomutants for this site. These results reveal a homeostatic mechanism through which activity regulates the dynamics and function of perisomatic inhibitory synapses, and they identify a CaMKIIdependent phosphorylation site on gephyrin as critically important for this process.inhibition | gabaergic synapse | plasticity | hippocampus | CaMKII S everal activity-dependent plasticity and homeostatic mechanisms (1, 2) contribute to regulate synaptic strength at excitatory synapses. Similar mechanisms are also expected to finely tune the level of inhibition in response to activity in individual neurons, but the mechanisms remain poorly understood. Different forms of plasticity at GABAergic synapses have been reported based on either presynaptic or postsynaptic mechanisms (3, 4). Similar to receptors at excitatory synapses, GABA A receptors (GABA A Rs), which mediate the fast component of inhibitory transmission, display complex trafficking mechanisms that affect the surface localization and diffusion of receptors (5). The distribution and clustering of GABA A Rs at synapses is tightly regulated through interactions with the scaffolding protein gephyrin, one of the main structural constituent of inhibitory postsynaptic densities. Gephyrin forms multimeric complexes that allow the anchoring of GABA A Rs (6) via molecular mechanisms that include phosphorylation and interactions with the guanine-nucleotide exchange factor collybistin (7-12). In addition to changes in inhibitory strength, more recent in vivo experiments revealed that inhibitory synapses are also dynamic structures that can be formed and eliminated in response to sensory experience (13-15). The mechanisms implicated in the coordinated regulation of excitatory an...

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Section: Discussionmentioning

confidence: 99%

Activity-dependent inhibitory synapse remodeling through gephyrin phosphorylation

Flores

Nikonenko

Méndez

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

113

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“…Phosphorylation [29][30][31][32] , palmitoylation 33,34 and acetylation 35,36 of gephyrin have been reported and these post-translational modifications have been shown to affect the structure, trafficking, half-life and importantly the ability of gephyrin to interact with partner proteins. To test whether the identified short GABA A Rderived peptide fragments a9-11 also mediate binding to native full-length gephyrin comprising post-translational modifications, the peptides were covalently immobilized on iodoacetyl-activated beads and subsequently incubated with mouse brain lysates.…”

Section: Resultsmentioning

confidence: 99%

Molecular basis of the alternative recruitment of GABAA versus glycine receptors through gephyrin

et al. 2014

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g-Aminobutyric acid type A and glycine receptors (GABA A Rs, GlyRs) are the major inhibitory neurotransmitter receptors and contribute to many synaptic functions, dysfunctions and human diseases. GABA A Rs are important drug targets regulated by direct interactions with the scaffolding protein gephyrin. Here we deduce the molecular basis of this interaction by chemical, biophysical and structural studies of the gephyrin-GABA A R a3 complex, revealing that the N-terminal region of the a3 peptide occupies the same binding site as the GlyR b subunit, whereas the C-terminal moiety, which is conserved among all synaptic GABA A R a subunits, engages in unique interactions. Thermodynamic dissections of the gephyrinreceptor interactions identify two residues as primary determinants for gephyrin's subunit preference. This first structural evidence for the gephyrin-mediated synaptic accumulation of GABA A Rs offers a framework for future investigations into the regulation of inhibitory synaptic strength and for the development of mechanistically and therapeutically relevant compounds targeting the gephyrin-GABA A R interaction.

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“…Puzzlingly, one study showed that protein phosphatase 1 directly interacted with gephyrin and that gephyrin clustering was decreased by the application of broad-spectrum phosphatase inhibitors, 45 whereas two other studies conversely found that the dephosphorylation of gephyrin increased gephyrin clustering. 43,44 In this context, it seems noteworthy that acute application of tumor necrosis factor-α downregulates inhibitory synaptic transmission via the protein phosphatase 1-dependent trafficking of GABA A receptors. 46 In addition to phosphorylation, gephyrin is subject to palmitoylation 47 (Figure 1).…”

Section: Posttranslational Modificationsmentioning

confidence: 99%

“…However, gephyrin has been identified as a phosphoprotein when copurified with the GlyR (which has kinase activity), 40 and mass spectrometric analyses of rat and mouse brains revealed that gephyrin has 22 phosphorylation sites, most of which are located within the C domain (except for threonine 324, which lies in the E domain 25,[41][42][43] ) (Figure 1). These modifications might induce conformational changes by affecting the structure of the C domain or the neighboring G and E domains, thereby altering the clustering, trafficking and binding properties of gephyrin.…”

Section: Posttranslational Modificationsmentioning

confidence: 99%

“…Interestingly, lithium chloride (an inhibitor of glycogen synthase kinase-β (GSK3β)) enhances gephyrin clustering in cultured neurons, and it has been speculated that the mood-stabilizing effect of lithium may be mediated at least partly via this mechanism. 44 Serine 268 of gephyrin is phosphorylated by extracellular signal-regulated kinases 1 and 2; this synergistically alters gephyrin clustering and alters the amplitude and frequency of miniature inhibitory postsynaptic currents 43 (Figure 1). In addition, cyclin-dependent kinase 5 mediates the collybistindependent phosphorylation of Ser270, suggesting that multiple signaling pathways converge at this residue of gephyrin.…”

Section: Posttranslational Modificationsmentioning

confidence: 99%

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Gephyrin: a central GABAergic synapse organizer

2015

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Gephyrin is a central element that anchors, clusters and stabilizes glycine and γ-aminobutyric acid type A receptors at inhibitory synapses of the mammalian brain. It self-assembles into a hexagonal lattice and interacts with various inhibitory synaptic proteins. Intriguingly, the clustering of gephyrin, which is regulated by multiple posttranslational modifications, is critical for inhibitory synapse formation and function. In this review, we summarize the basic properties of gephyrin and describe recent findings regarding its roles in inhibitory synapse formation, function and plasticity. We will also discuss the implications for the pathophysiology of brain disorders and raise the remaining open questions in this field.

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Extracellular Signal-regulated Kinase and Glycogen Synthase Kinase 3β Regulate Gephyrin Postsynaptic Aggregation and GABAergic Synaptic Function in a Calpain-dependent Mechanism

Cited by 93 publications

References 30 publications

Activity-dependent inhibitory synapse remodeling through gephyrin phosphorylation

Activity-dependent inhibitory synapse remodeling through gephyrin phosphorylation

Molecular basis of the alternative recruitment of GABAA versus glycine receptors through gephyrin

Gephyrin: a central GABAergic synapse organizer

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