Activation of Pheromone-Sensitive Neurons Is Mediated by Conformational Activation of Pheromone-Binding Protein

Abstract

Detection of volatile odorants by olfactory neurons is thought to result from direct activation of seven-transmembrane odorant receptors by odor molecules. Here, we show that detection of the Drosophila pheromone, 11-cis vaccenyl acetate (cVA), is instead mediated by pheromone-induced conformational shifts in the extracellular pheromone-binding protein, LUSH. We show that LUSH undergoes a pheromone-specific conformational change that triggers the firing of pheromone-sensitive neurons. Amino acid substitutions in LUSH that are predicted to reduce or enhance the conformational shift alter sensitivity to cVA as predicted in vivo. One substitution, LUSH(D118A), produces a dominant-active LUSH protein that stimulates T1 neurons through the neuronal receptor components Or67d and SNMP in the complete absence of pheromone. Structural analysis of LUSH(D118A) reveals that it closely resembles cVA-bound LUSH. Therefore, the pheromone-binding protein is an inactive, extracellular ligand converted by pheromone molecules into an activator of pheromone-sensitive neurons and reveals a distinct paradigm for detection of odorants.

New Insights into the Mechanism of Odorant Detection by the Malaria-transmitting Mosquito Anopheles gambiae

“…The effect of indole on the structure of OBP4 correlates well with our previous findings for the Drosophila OBP LUSH, where binding of ligand induces a significant conformational ordering that is required to regulate its interactions with other components of the olfactory signaling cascade including sensory neuron membrane protein (26,53,57,58). Conformational heterogeneity has been observed in several other OBPs (14,17,59,60) and is most often associated with residues that regulate access to the ligand binding pocket.…”

“…In contrast, we and others have proposed that in some cases a specific complex between the OBP and odorant is required for odorant receptor activation (20 -25). In support of this, we demonstrated that a key amino acid substitution in a critical loop of the Drosophila melanogaster OBP LUSH could activate pheromone receptors in the complete absence of pheromone (23,26,27), providing direct evidence that the OBP is the ligand for the receptor complex. These differences in function most likely reflect the level of control, with responses to general odorants being less tightly regulated than other sexual or species-specific responses.…”

“…1E). In LUSH substitution of Asp-118 with alanine in this loop generated a constitutively active protein (26). In OBP4 (and OBP1) the homologous residue is a proline, and this loop adopts the same conformation seen in OBP1.…”

“…However, medium length OBPs like AgOBP4, LUSH (22,26,37), or honey bee PBP Asp-1 (56) do not undergo any pH-dependent change in the C-terminal tail that affects ligand binding. In this study we saw no change in the structure of apoOBP4 at low pH values, and further indole could still bind and induce the structural ordering of OBP4 even at pH 4.6.…”

Queen mandibular pheromone: questions that remain to be resolved

“…Here, the pheromone binds to carrier proteins that help transport the pheromone to olfactory receptors (ORs) located in the ORN membrane (Laughlin et al 2008;Vogt and Riddiford 1981). The candidate carrier protein in the drone antenna is ASP1 which contains a hydrophobic domain that is able to bind the apolar components of 9ODA (Pesenti et al 2008).…”