Conversion of the ion selectivity of the 5-HT(3a) receptor from cationic to anionic reveals a conserved feature of the ligand-gated ion channel superfamily.
The 5-hydroxytryptamine(3) (5-HT(3)) receptor is a member of a superfamily of ligand-gated ion channels, which includes nicotinic acetylcholine, gamma-aminobutyric acid, and glycine receptors. The receptors are either cation or anion selective, leading to their distinctive involvement in either excitatory or inhibitory neurotransmission. Using a combination of site-directed mutagenesis and electrophysiological characterization of homomeric 5-HT(3A) receptors expressed in HEK293 cells, we have identified a set of mutations that convert the ion selectivity of the 5-HT(3A) receptor from cationic to anionic; these were substitution of V13'T in M2 together with neutralization of glutamate residues (E-1'A) and the adjacent insertion of a proline residue (P-1') in the M1-M2 loop. Mutant receptors showed significant chloride permeability (P(Cl)/P(Na) = 12.3, P(Na)/P(Cl) = 0.08), whereas WT receptors are predominantly permeable to sodium (P(Na)/P(Cl) > 20, P(Cl)/P(Na) < 0.05). Since the equivalent mutations have previously been shown to convert alpha7 nicotinic acetylcholine receptors from cationic to anionic (Galzi J.-L., Devillers-Thiery, A, Hussy, N., Bertrand, S. Changeux, J. P., and Bertrand, D. (1992) Nature 359, 500-505) and, recently, the converse mutations have allowed the construction of a cation selective glycine receptor (Keramidas, A., Moorhouse, A. J., French, C. R., Schofield, P. R., and Barry, P. H. (2000) Biophys. J. 78, 247-259), it appears that the determinants of ion selectivity represent a conserved feature of the ligand-gated ion channel superfamily.
Gunthorpe
M J
MJ
Division of Neurobiology, Medical Research Council Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, United Kingdom. s.lummis@mole.bio.ca.ac.uk
Lummis
S C
SC
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000838:Anions; D002412:Cations; D002460:Cell Line; D002712:Chlorides / Q000378:metabolism; D004563:Electrochemistry; D006801:Humans; D015640:Ion Channel Gating / Q000502:physiology*; D008024:Ligands; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018408:Patch-Clamp Techniques; D017434:Protein Structure, Tertiary; D011985:Receptors, Serotonin / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D044406:Receptors, Serotonin, 5-HT3; D011994:Recombinant Proteins; D016415:Sequence Alignment; D012964:Sodium / Q000378:metabolism
D000838:Anions; D002412:Cations; D002712:Chlorides; D008024:Ligands; D011985:Receptors, Serotonin; D044406:Receptors, Serotonin, 5-HT3; D011994:Recombinant Proteins; D012964:Sodium