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Expression of the alpha 1b-adrenergic receptor and G protein subunits in mammalian cell lines using the Semliki Forest virus expression system.
Journal of Receptor and Signal Transduction Research
Date de publication
Semliki Forest virus (SFV) vectors have been efficiently used for rapid high level expression of several G protein-coupled receptors. Here we describe the use of SFV vectors to express the alpha 1b-adrenergic receptor (AR) alone or in the presence of the G protein alpha q and/or beta 2 and gamma 2 subunits. Infection of baby hamster kidney (BHK) cells with recombinant SFV-alpha 1b-AR particles resulted in high specific binding activity of the alpha 1b-AR (24 pmol receptor/mg protein). Time-course studies indicated that the highest level of receptor expression was obtained 30 hours post-infection. The stimulation of BHK cells, with epinephrine led to a 5-fold increase in inositol phosphate (IP) accumulation, confirming the functional coupling of the receptor to G protein-mediated activation of phospholipase C. The SFV expression system represents a rapid and reproducible system to study the pharmacological properties and interactions of G protein coupled receptors and of G protein subunits.
Animals, CHO Cells, COS Cells, Cell Line, Cricetinae, GTP-Binding Proteins/biosynthesis, GTP-Binding Proteins/chemistry, Gene Expression, Genetic Vectors, Inositol Phosphates/metabolism, Kinetics, Protein Conformation, Receptors, Adrenergic, alpha-1/biosynthesis, Receptors, Adrenergic, alpha-1/chemistry, Recombinant Proteins/biosynthesis, Recombinant Proteins/chemistry, Semliki forest virus/genetics
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