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

Abstract

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.

Design and Synthesis of High‐Affinity Dimeric Inhibitors Targeting the Interactions between Gephyrin and Inhibitory Neurotransmitter Receptors

“…[1a,b, 2b, 5,6,7] Truncation of 1 a to a 14-residue core binding peptide (1 b), which could be resolved in a X-ray crystal structure, [5] resulted in a roughly threefold weaker affinity (K D = (6 AE 2) mm).…”

“…[2e, 4] Gephyrin interacts with GlyRs and GABA A Rs via a universal binding site [2b] located within the gephyrin E domain (GephE), [5] although distinct GlyR and GABA A R interactions have very recently been described. [6] The affinity of the gephyrin-receptor interaction depends on the oligomeric state of gephyrin and the number of gephyrin-binding receptor subunits in the functional receptor. [7] Consequently, the oligomerization of gephyrin, GlyRs, and GABA A Rs are interdependent.…”

Shared function and moonlighting proteins in molybdenum cofactor biosynthesis

“…In addition to its interaction with the glycine receptor, gephyrin also interacts with the intracellular loop of the GABA A receptor. Gephyrin thereby is a central player at inhibitory synapses, as it is the structural receptor scaffold and acts at a platform for facilitating protein-protein interactions, bringing receptors, cytoskeletal elements and signaling proteins into close proximity (Maric et al, 2014). Gephyrin thus regulates clustering and diffusion of these receptors of fast inhibitory transmission.…”

“…Gephyrin thus regulates clustering and diffusion of these receptors of fast inhibitory transmission. The gephyrin protein network in particular also tethers the GABA A receptors to the cytoskeleton by direct interaction with profilin and other proteins like Mena/VASP (Maric et al, 2014) (Figure 5). Gephyrin in total is comprized of three domains, the N-terminal G-domain, the C-terminal E-domain and a central domain referred to as linker.…”

Physiological properties of Glycinergic synapses with defined subunit compositions

“…Pentameric GlyRs can be formed as homomers consist of α subunits only, or as heteromers comprising α and β subunits in the form of 3α:2β or 2α:3β . The β subunit is important for synaptic localization as it interacts directly with gephyrin, an intracellular protein essential for clustering GlyRs at synapses (Sola et al, 2004;Maric et al, 2014). The maturation of glycinergic synapses is associated with changes in the composition and functional properties of GlyRs.…”