Ectopically Expressed Human Olfactory Receptor OR51E1: a Search for Novel Molecular Probes

Abstract

Olfactory receptors comprise the largest family of G protein‐couples receptors (GPCRs) ‐ ~1400 functional genes in the mouse (Olfrs) and ~400 in the human (ORs). Most genes were assigned to it based solely on the sequence similarity. Recent studies demonstrated that many Olfrs are expressed outside of the nasal cavity and possibly involved in physiological processes such as muscle regeneration and controlling blood pressure. These ectopically expressed olfactory receptors might recognize cues other than odorants and be involved in processes independent of the sense of smell. Ligands were identified for fewer than 100 of ORs and Olfrs.Using transcriptomics analysis followed by RT‐PCR and in situ RNA hybridization we previously showed expression of several Olfrs and other olfactory signaling genes expressed in the mouse ocular tissues. We focused our attention on Olfr558 and demonstrated its expression in blood vessels of the eye. The human ortholog of Olfr558 (OR51E1 or PSGR2) was shown to be expressed in the prostate and dramatically up‐regulated in prostate cancer and is an established biomarker of malignancy. The human and mouse genes encoding this receptor are almost identical (94%), which is unique for GPCRs in general, and particularly for sensory chemoreceptors. This unusual evolutionary conservation points to the importance of its physiological function. While the function of this receptor or its native ligand is unknown, in vitro it responds to some short‐ and medium‐chain fatty acids (e.g., butyrate and valerate) by activating adenylyl cyclase. We found that growth of human prostate cancer cells that express OR51E1 is inhibited by valerate. The knockdown of OR51E1 gene with shRNA eliminated this effect of valerate.While butyrate and valerate can be used in such experiments, their low affinity and selectivity complicates interpretation of the data. Additionally, they are quickly metabolized in the body. Thus, there is a clear need for novel probes that can be used to dissect OR51E1 function. To address this need, we established a stable cell line overexpressing OR51E1 and demonstrated that in response to known OR51E1 agonists this cell line increases intracellular cAMP level. We then used this cell line for screening of a diverse chemical library of 9600 compounds, showing its suitability for high throughput screening. To demonstrate that our expression system can be used for expression of other ORs, we generated stable cell lines overexpressing two other receptors – OR51E2 and OR2AT4. We showed that both receptors are expressed as proteins of the expected molecular weight and localize on the cell surface. Since these ORs represents a large class of previously unexplored yet druggable targets, this project facilitates investigation of the entire family of ectopic olfactory receptors.