GABAA receptors are pentameric ligand-gated ion channels (pLGIC) that mediate inhibitory fast synaptic transmission in the central nervous system. Consistent with recent pLGIC structures, sequence analysis predicts an α-helix near the N-terminus of each GABAA receptor subunit. Preceding each α-helix are 8-36 additional residues, which we term the Nterminal extension. In homomeric GABAC receptors and nicotinic acetylcholine receptors (nAChR), the N-terminal α-helix is functionally essential. Here we determined the role of the N-terminal extension and putative α-helix in heteromeric α1β2γ2 GABAA receptors. This role was most prominent in the α1 subunit, with deletion of the N-terminal extension or further deletion of the putative α-helix both dramatically reduced the number of functional receptors at the cell surface. Conversely, deletion of the β2 or γ2 N-terminal extension had little effect on the number of functional cell-surface receptors. Additional deletion of the putative α-helix in the β2 or γ2 subunits did, however, decrease both functional cell surface receptors and incorporation of the γ2 subunit into mature receptors. In the β2 subunit only, α-helix deletions affected GABA sensitivity and desensitization. Our findings demonstrate that N-terminal extensions and α-helices make key subunit-specific contributions to assembly, consistent with both regions being involved in inter-subunit interactions.
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