Expression and intracellular localisation of odorant receptors in mammalian cell lines using Semliki Forest virus vectors
| Autor: | J R Pink, Kenneth Lundstrom, F Goepfert, K Monastyrskaia, R Hochstrasser, J Leighton, G Acuña |
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| Rok vydání: | 1999 |
| Předmět: |
Genetic Vectors
Gene Expression Olfaction Biology Semliki Forest virus Receptors Odorant Biochemistry Cell Line Mice Cricetinae Baby hamster kidney cell medicine Animals Receptor Caenorhabditis elegans Molecular Biology Neurons Endoplasmic reticulum Membrane Proteins Cell Biology Helminth Proteins biology.organism_classification Molecular biology Semliki forest virus Recombinant Proteins Rats medicine.anatomical_structure Membrane protein Cell culture Neuron Subcellular Fractions |
| Zdroj: | Journal of receptor and signal transduction research. 19(1-4) |
| ISSN: | 1079-9893 |
| Popis: | Odorant receptors are members of the G protein-coupled receptor superfamily. They are expressed on the surface of cilia of olfactory neurons, where they bind ligand (odorant). Studies of the molecular mechanisms of olfaction are complicated by the extremely large number of receptor genes, and difficulties in pairing a particular mammalian receptor to a specific odorant ligand 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 Semliki Forest virus (SFV) system. All receptors were epitope-tagged at the N- or 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) cells, were infected and tested. Immunofluorescence and confocal microscopy studies performed on permeabilised, non-permeabilised and native cells revealed 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 inserted into the plasma membrane. Similar results were obtained using primary neurons, indicating that like mature neurons, the immortalised OLF 442 cells are capable of providing for correct odorant receptor processing and targeting. |
| Databáze: | OpenAIRE |
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