Collybistin activation by GTP-TC10 enhances postsynaptic gephyrin clustering and hippocampal GABAergic neurotransmission

Abstract

In many brain regions, gephyrin and GABA A receptor clustering at developing inhibitory synapses depends on the guanine nucleotide exchange factor collybistin (Cb). The vast majority of Cb splice variants contain an autoinhibitory src homology 3 domain, and several synaptic proteins are known to bind to this SH3 domain and to thereby activate gephyrin clustering. However, many functional GABAergic synapses form independently of the known Cb-activating proteins, indicating that additional Cb activators must exist. Here we show that the small Rho-like GTPase TC10 stimulates Cb-dependent gephyrin clustering by binding in its active, GTP-bound state to the pleckstrin homology domain of Cb. Overexpression of a constitutively active TC10 variant in neurons causes an increase in the density of synaptic gephyrin clusters and mean miniature inhibitory postsynaptic current amplitudes, whereas a dominant negative TC10 variant has opposite effects. The enhancement of Cb-induced gephyrin clustering by GTP-TC10 does not depend on the guanine nucleotide exchange activity of Cb but involves an interaction that resembles reported interactions of other small GTPases with their effectors. Our data indicate that GTP-TC10 activates the major src homology 3 domain-containing Cb variants by relieving autoinhibition and thus define an alternative GTPase-driven signaling pathway in the genesis of inhibitory synapses.RhoQ | Cdc42 | postsynaptic scaffold | synaptogenesis | neuroligin

Gephyrin: a central GABAergic synapse organizer

“…Neuroligin-2 (NL-2) and the Rho-like GTPase, TC10, have been shown to have this role by interacting with the collybistin SH3 domain. 76,77 Furthermore, a recent structural study demonstrated that the collybistins can adopt open and closed conformations that act as switchable adaptors, and that NL-2 favors the open conformation by competing with a closed-conformation-favoring intramolecular interaction in collybistin. 78 The proposed functions of collybistin were demonstrated in vivo: collybistin-deficient mice displayed region-specific reductions in synaptic gephyrin clustering, reduced clustering of the GABA A receptor γ2-subunit, decreased GABAergic synaptic transmission, impaired hippocampal synaptic plasticity (i.e., increased long-term potentiation and decreased long-term depression) and impaired spatial learning.…”

Protein Kinase C-Dependent Growth-Associated Protein 43 Phosphorylation Regulates Gephyrin Aggregation at Developing GABAergic Synapses

“…Although gephyrin forms submembranous clusters in neurons, it is also known to form large aggregates/ clumps in nonneuronal cells (13,43). We examined how GAP43 and its two S41 mutants could differentially affect gephyrin folding and aggregation in HEK293T cells.…”

Activity-dependent inhibitory synapse remodeling through gephyrin phosphorylation

“…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)(8)(9)(10)(11)(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)(14)(15).…”