Distinct γ2 subunit domains mediate clustering and synaptic function of postsynaptic GABA_A receptors and gephyrin

Modulation of the concentration of postsynaptic GABA_A receptors contributes to functional plasticity of inhibitory synapses. The γ2 subunit of GABA_A receptor is specifically required for clustering of these receptors, for recruitment of the submembrane scaffold protein gephyrin to postsynaptic sites, and for postsynaptic function of GABAergic inhibitory synapses. To elucidate this mechanism, we here have mapped the γ2 subunit domains required for restoration of postsynaptic clustering and function of GABA_A receptors in γ2 subunit mutant neurons. Transfection of γ2^-/- neurons with the γ2 subunit but not the α2 subunit rescues postsynaptic clustering of GABA_A receptors, results in recruitment of gephyrin to postsynaptic sites, and restores the amplitude and frequency of miniature inhibitory postsynaptic currents to wild-type levels. Analogous analyses of chimeric γ2/α2 subunit constructs indicate, unexpectedly, that the fourth transmembrane domain of the γ2 subunit is required and sufficient for postsynaptic clustering of GABA_A receptors, whereas cytoplasmic γ2 subunit domains are dispensable. In contrast, both the major cytoplasmic loop and the fourth transmembrane domain of the γ2 subunit contribute to efficient recruitment of gephyrin to postsynaptic receptor clusters and are essential for restoration of miniature IPSCs. Our study points to a novel mechanism involved in targeting of GABA _A receptors and gephyrin to inhibitory synapses.