Ectopically Expressed Olfactory GPCRs OR51E1/2: Ligand Specificity, Signal Transduction, and Cytostatic and Cytotoxic Activity

Abstract

Olfactory receptors (ORs) are the largest family of G protein-coupled receptors: there are ~380 ORs in humans and ~1,000 in mice. This family was originally cloned from the nasal epithelium, but several years ago, expression of many OR genes was detected in the airways, gut, blood vessels, brain, and other organs. These GPCRs are referred to as ectopic ORs (eORs). Understanding eOR function has lagged behind that of other GPCRs, and the majority of ORs remain to be orphan receptors. One reason for this delay is that the expression of functionally active ORs has not been technically possible for many years. In contrast to the most other Class A GPCRs, which can be produced using standard methods such as transient transfection, stability and trafficking ORs requires special chaperones and/or modification with unique signal peptides. Difficulties with detection and functional analysis of both endogenous and overexpressed eORs caused many controversies, and overall, this vast and potentially interesting family of GPCRs has been grossly understudied. Here, we investigated two eORs, OR51E1 and OR51E2, which due to their marked upregulation in prostate cancer are also known as “prostate-specific G protein-coupled receptors” (PSGR2 and PSGR, respectively). To investigate their signaling and effects on proliferation and survival we developed an inducible expression system in a prostate cancer cell line LNCaP. The robust expression and trafficking of ORs allowed us to analyze their function using cAMP, Ca2+, protein phosphorylation and other assays. Consistent with previous studies, OR51E1 stimulated adenylyl cyclase in response to treatment by short- to medium-chain organic acids (C3-C9) an some branched aliphatic acids. OR51E2 responded to acetate and propionate, but not to the longer chain organic acids. Other putative ligands mentioned in the literature did not have activity. Stimulation of LNCaP cells with butyrate inhibited their growth, and the knockdown of the endogenous OR51E1 negated this cytostatic effect. Interestingly, induced overexpression of OR51E1 or OR51E2 suppressed LNCaP cell proliferation. Overexpression of OR2AT4, beta2-adrenergic receptor or treatment of cells with forskolin did not suppress cell proliferation, indicating that rise in cAMP is not sufficient to induce cytostasis. These findings indicate that like other GPCRs, eORs can signal via more than one G protein pathway. Overexpression of OR51E1 caused upregulation of cytostatic and cell death markers including p27, p21, and p53, strongly increased annexin V staining and stimulated ERK1/2. OR51E1 had no effect on proliferation of HEK293 cells, indicating that cytotoxicity of OR51E1/2 is specific for LNCaP cells. Elucidating this mechanism will contribute to understanding etiology of prostate cancer and may illuminated these two and possibly other eORs as future therapeutic targets.