Journal of Molecular Neuroscience

, Volume 18, Issue 3, pp 169–178 | Cite as

Immunological characterization of 5-HT3 receptor transmembrane topology

Article

Abstract

The 5-hydroxytryptamine3 (5-HT3) receptor is a member of the Cys-loop family of ligand-gated ion channels. These receptors are pentamers with the greatest homology to nicotinic acetylcholine (nACh) receptors. The proposed topological organization of a 5-HT3 receptor subunit is based largely on hydropathy profiles and by homology to nACh receptors, and indicates a large N-terminal extracellular domain and four transmembrane regions. There is, however, little direct evidence for this model. We therefore investigated the topology of the 5-HT3A receptor subunit using a panel of 5-HT3 receptor-specific antisera that interact with defined regions of the receptor. An antiserum generated against a short peptide from the N-terminal domain of the 5-HT3A receptor subunit, pAb120, was shown to bind to 5-HT3 receptor-expressing cells with intact cell membranes, indicating that the N-terminal end of the subunit is extracellular. Two antisera generated against regions of the loop between predicted transmembrane regions three and four did not bind to cells with intact membranes. However on membrane permeabilization these antibodies both bound to the receptor in intracellular areas, thus indicating that the loop between transmembrane domains three and four is intracellular. These data therefore provide direct evidence for an extracellular N-terminal domain and an intracellular loop between the third and fourth transmembrane domains, thus supporting the conventional ligand-gated ion channel subunit topological model.

Index Entries

Serotonin 5-hydroxytryptamine ligand-gated ion channel 5-HT3 receptor immunochemistry 

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Copyright information

© Humana Press Inc 2002

Authors and Affiliations

  1. 1.Division of Neurobiology, Laboratory of Molecular BiologyMRC CentreCambridgeUK
  2. 2.Department of BiochemistryUniversity of CambridgeCambridgeUK

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