Glycine Receptors Caught between Genome and Proteome - Functional Implications of RNA Editing and Splicing

Abstract

Information processing in the brain requires a delicate balance between excitation and inhibition. Glycine receptors (GlyR) are involved in inhibitory mechanisms mainly at a synaptic level, but potential novel roles for these receptors recently emerged due to the discovery of posttranscriptional processing. GLR transcripts are edited through enzymatic modification of a single nucleotide leading to amino acid substitution within the neurotransmitter binding domain. RNA editing produces gain-of-function receptors well suited for generation and maintenance of tonic inhibition of neuronal excitability. As neuronal activity deprivation in early stages of development or in epileptic tissue is detrimental to neurons and because RNA editing of GlyR is up-regulated in temporal lobe epilepsy patients with a severe course of disease a pathophysiological role of these receptors emerges. This review contains a state-of-the-art discussion of (patho)physiological implications of GlyR RNA editing.

Keywords: GABA; RNA editing; RNA splicing; epilepsy; hippocampus.

Figure 1

Apparent agonist affinities of GlyR α1^185P and α1^185L. Dose-response curves were obtained from transfected HEK293 cells. (A,C) Example traces of Cl⁻ currents obtained at indicated agonist concentrations. At −70 mV holding potential, maximal current responses (α1^185P, taurine: 3.3 ± 0.3 nA, glycine: 2.5 ± 0.3 nA; α1^185L, taurine: 1.7 ± 0.3 nA, glycine: 1.6 ± 0.3 nA) were obtained with 2 mM glycine and 10 mM taurine. Taurine (B) and glycine (D) dose-response curves of HEK293 cells expressing GlyR α1^185P or α1^185L are shown. Taurine and glycine current amplitudes were normalized to maximal current amplitudes. Each data point (B,D) represents the average current amplitude of 13–21 sampled cells. Hill coefficients: α1^185P, taurine: 1.06 ± 0.09, glycine: 1.88 ± 0.17; α1^185L, taurine: 0.84 ± 0.07, glycine: 1.08 ± 0.09. Values represent mean ± SD.

Figure 2

Apparent agonist affinities of GlyR α3L^185P and α3L^185L. Dose-response curves were obtained from transfected HEK293 cells. (A) Example traces of Cl⁻ currents obtained at indicated agonist concentrations. (B) Glycine dose-response curves of HEK293 cells expressing GlyR α3L^185P or α3L^185L are shown. Glycine current amplitudes were normalized to maximal current amplitudes. Each data point represents the average current amplitude of 13–21 sampled cells. At −70 mV holding potential, the apparent affinities for glycine (EC_50(Glycine) [μM]) were 70.9 ± 16.1 (α3L^185P) and 7.4 ± 0.9 (α3L^185L). Hill coefficients were 0.87 ± 0.17 (α3L^185P) and 0.70 ± 0.06 (α3L^185L). Thus, the long splice variant of RNA-edited GlyR α3L^185L is a high affinity receptor as well. Values represent mean ± SD.

Figure 3

Tertiary structure of AChBP and projection of GlyR α subunit associated structural determinants. Positions of the glycine (yellow) binding pocket, formed by amino acids GY and KYT (C-loop), of the disulfide bridge between β-sheets 6’ and 7 as well as of proline residues at positions 185 (α1 and α3) and 192 (α2) are shown. N- and C-termini and the location of the chloride channel pore are indicated. β-sheets, α-helices and coils are color-coded (green, red and black, respectively). GlyR sequence was projected on the structure view of AChBP downloaded from www.pdb.org. The position of β-sheets served as reference points.

Figure 4

Activation of GlyR α3K^185P and GlyR α3K^185L by GABA applications. (A1) Example traces of outside-out currents evoked by activation of GlyR α3K^185P in response to the application of glycine (0.1 mM) and various concentrations of GABA (Vh = −50 mV; Filter 1 kHz). (A2) Example traces of outside-out currents evoked by activation of GlyR α3K^185L in response to the application of glycine (0.1 mM) and various concentrations of GABA (Vh = −50 mV; Filter 1 kHz). (B) Concentration-response curves for GABA-evoked currents on outside-out patches containing GlyR α3K^185P (○) or GlyR α3K^185L (●). Measurements were performed on averaged traces. The amplitude of GABA-evoked currents was normalized to the peak amplitude of the current evoked by the application of 0.1 mM glycine (relative current). Numbers above data points indicate the number of measurements per point (mean ± SD).