Expression and intracellular localisation of odorant receptors in mammalian cell lines using Semliki Forest virus vectors

Odorant receptors are members of the G protein-coupled receptor superfamily . They are expressed on the surface of cilia of olfactory neurons, where th ey bind ligand (odorant). Studies of the molecular mechanisms of olfaction are complicated by the extremely large number of receptor genes, and diffic ulties in pairing a particular mammalian receptor to a specific odorant lig and in vivo. Here we report expression and localisation studies of two rat odorant receptor genes (17 and OR5), and C. elegans odr-10, using the Semli ki Forest virus (SFV) system. All receptors were epitope-tagged at the N- o r C-terminus in order to facilitate their detection in infected cells, and determine the localisation and membrane-orientation of recombinant proteins . The immortalised mouse olfactory neuronal cell line OLF 442, rat cortical and striatal primary neuron cultures, and the baby hamster kidney (BHK) ce lls, were infected and tested. Immunofluorescence and confocal microscopy s tudies performed on permeabilised, non-permeabilised and native cells revea led that in BHK cells the rat receptors 17 and OR5 were not targeted to the plasma membrane and remained in the endoplasmic reticulum. In contrast, in the mouse olfactory cell line OLF 442 both rat receptors were correctly in serted into the plasma membrane. Similar results were obtained using primar y neurons, indicating that like mature neurons, the immortalised OLF 442 ce lls are capable of providing for correct odorant receptor processing and ta rgeting.