Characterization of the 5-HT1A receptor of the honeybee (Apis mellifera) and involvement of serotonin in phototactic behavior

Serotonin plays a key role in modulating various physiological and behavioral processes in both protostomes and deuterostomes. The vast majority of serotonin receptors belong to the superfamily of G-protein-coupled receptors. We report the cloning of a cDNA from the honeybee (Am5-ht1A) sharing high similarity with members of the 5-HT₁ receptor class. Activation of Am5-HT_1A by serotonin inhibited the production of cAMP in a dose-dependent manner (EC₅₀ = 16.9 nM). Am5-HT_1A was highly expressed in brain regions known to be involved in visual information processing. Using in vivo pharmacology, we could demonstrate that Am5-HT_1A receptor ligands had a strong impact on the phototactic behavior of individual bees. The data presented here mark the first comprehensive study—from gene to behavior—of a 5-HT_1A receptor in the honeybee, paving the way for the eventual elucidation of additional roles of this receptor subtype in the physiology and behavior of this social insect.     

X-ray structure of a prokaryotic pentameric ligand-gated ion channel

Pentameric ligand-gated ion channels (pLGICs) are key players in the early events of electrical signal transduction at chemical synapses. The family codes for a structurally conserved scaffold of channel proteins that open in response to the binding of neurotransmitter molecules. All proteins share a pentameric organization of identical or related subunits that consist of an extracellular ligand-binding domain followed by a transmembrane channel domain. The nicotinic acetylcholine receptor (nAChR) is the most thoroughly studied member of the pLGIC family (for recent reviews see refs 1-3). Two sources of structural information provided an architectural framework for the family. The structure of the soluble acetylcholine-binding protein (AChBP) defined the organization of the extracellular domain and revealed the chemical basis of ligand interaction. Electron microscopy studies of the nAChR from Torpedo electric ray have yielded a picture of the full-length protein and have recently led to the interpretation of an electron density map at 4.0 A resolution. Despite the wealth of experimental information, high-resolution structures of any family member have so far not been available. Until recently, the pLGICs were believed to be only expressed in multicellular eukaryotic organisms. The abundance of prokaryotic genome sequences, however, allowed the identification of several homologous proteins in bacterial sources. Here we present the X-ray structure of a prokaryotic pLGIC from the bacterium Erwinia chrysanthemi (ELIC) at 3.3 A resolution. Our study reveals the first structure of a pLGIC at high resolution and provides an important model system for the investigation of the general mechanisms of ion permeation and gating within the family.     

Structure of a potentially open state of a proton-activated pentameric ligand-gated ion channel

The X-ray structure of a pentameric ligand-gated ion channel from Erwinia chrysanthemi (ELIC) has recently provided structural insight into this family of ion channels at high resolution. The structure shows a homo-pentameric protein with a barrel-stave architecture that defines an ion-conduction pore located on the fivefold axis of symmetry. In this structure, the wide aqueous vestibule that is encircled by the extracellular ligand-binding domains of the five subunits narrows to a discontinuous pore that spans the lipid bilayer. The pore is constricted by bulky hydrophobic residues towards the extracellular side, which probably serve as barriers that prevent the diffusion of ions. This interrupted pore architecture in ELIC thus depicts a non-conducting conformation of a pentameric ligand-gated ion channel, the thermodynamically stable state in the absence of bound ligand. As ligand binding promotes pore opening in these ion channels and the specific ligand for ELIC has not yet been identified, we have turned our attention towards a homologous protein from the cyanobacterium Gloebacter violaceus (GLIC). GLIC was shown to form proton-gated channels that are activated by a pH decrease on the extracellular side and that do not desensitize after activation. Both prokaryotic proteins, ELIC and GLIC form ion channels that are selective for cations over anions with poor discrimination among monovalent cations, characteristics that resemble the conduction properties of the cation-selective branch of the family that includes acetylcholine and serotonin receptors. Here we present the X-ray structure of GLIC at 3.1 A resolution. The structure reveals a conformation of the channel that is distinct from ELIC and that probably resembles the open state. In combination, both structures suggest a novel gating mechanism for pentameric ligand-gated ion channels where channel opening proceeds by a change in the tilt of the pore-forming helices.