Up-regulation of GABAB Receptor Signaling by Constitutive Assembly with the K+ Channel Tetramerization Domain-containing Protein 12 (KCTD12)

GABAB receptors are the G-protein coupled receptors (GPCRs) for GABA, the main inhibitory neurotransmitter in the central nervous system. Native GABAB receptors comprise principle and auxiliary subunits that regulate receptor properties in distinct ways. The principle subunits GABAB1a, GABAB1b, and GABAB2 form fully functional heteromeric GABAB(1a,2) and GABAB(1b,2) receptors. Principal subunits regulate forward trafficking of the receptors from the endoplasmic reticulum to the plasma membrane and control receptor distribution to axons and dendrites. The auxiliary subunits KCTD8, -12, -12b, and -16 are cytosolic proteins that influence agonist potency and G-protein signaling of GABAB(1a,2) and GABAB(1b,2) receptors. Here, we used transfected cells to study assembly, surface trafficking, and internalization of GABAB receptors in the presence of the KCTD12 subunit. Using bimolecular fluorescence complementation and metabolic labeling, we show that GABAB receptors associate with KCTD12 while they reside in the endoplasmic reticulum. Glycosylation experiments support that association with KCTD12 does not influence maturation of the receptor complex. Immunoprecipitation and bioluminescence resonance energy transfer experiments demonstrate that KCTD12 remains associated with the receptor during receptor activity and receptor internalization from the cell surface. We further show that KCTD12 reduces constitutive receptor internalization and thereby increases the magnitude of receptor signaling at the cell surface. Accordingly, knock-out or knockdown of KCTD12 in cultured hippocampal neurons reduces the magnitude of the GABAB receptor-mediated K+ current response. In summary, our experiments support that the up-regulation of functional GABAB receptors at the neuronal plasma membrane is an additional physiological role of the auxiliary subunit KCTD12. Background: KCTD12 regulates agonist potency and kinetics of GABAB receptor signaling. Results: KCTD12 stably associates with GABAB receptors as early as in the endoplasmic reticulum and reduces constitutive receptor internalization from the cell surface. Conclusion: KCTD12 increases GABAB receptor surface expression and the magnitude of receptor signaling. Significance: KCTD12 not only regulates agonist potency and kinetics but also the magnitude of GABAB receptor signaling.