Programme Contents

Are the putative N-terminal α-helix and preceding residues important in GABA_A/C receptor function?

The GABA_A/C receptors are members of the pentameric Cys-loop superfamily of ligand-gated ion channels (pLGICs) and mediate inhibitory fast synaptic transmission in the nervous system. pLGICs comprise an N-terminal extracellular agonist-binding domain followed by a channel domain and intracellular domain. Available structural information shows the agonist binding domain comprises a β sandwich of ten β-strands, which form the agonist binding pocket, preceded by an N-terminal α-helix in eukaryotic structures, not present in prokaryotic structures. Sequence analysis of GABA_A/C receptors predicts an α-helix in a similar position. This putative α-helix in GABA_A/C receptors is, however, preceded by 10 to 46 additional residues not present in other pLGICs, which we term the N-terminal extension. The N-terminal α-helix has been shown to be functional essential in nicotinic acetylcholine receptors (Bar-Lev et al., 2011; Castillo et al., 2009). The role of the α-helix and N-terminal extension in GABA_A/C receptors has not been tested and is the subject of this study. We found in both homomeric ρ1 GABA_C and heteromeric α1β2γ2 GABA_A receptors that the putative α-helix is essential for receptor function but not the N-terminal extension. Partial or complete removal of the N-terminal extension in ρ1 GABA_C receptors did, however, affect cell surface expression and agonist sensitivity. Mutation of a putative furin protease cleavage motif within the N-terminal extension of ρ1 GABA_C receptors likewise impaired cell surface expression. Our results support the essential role of the N-terminal α-helix in eukaryotic pLGICs and provide evidence that the N-terminal extension has a subtle modulatory role on receptor function.

Castillo M, Mulet J, Aldea M, Gerber S, Sala S, Sala F, Criado M. (2009) Role of the N-terminal α-helix in biogenesis of α7 nicotinic receptors. Journal of Neurochemistry 108: 1399-1409.

Bar-Lev DD, Degani-Katzav N, Perelman A, Paas Y. (2011) Molecular dissection of a Cl⁻-selective Cys-loop receptor points to components that are dispensable or essential for channel activity. Journal of Biological Chemistry 286: 43830-41.