Approaches to the Stable Transfection of G Protein-Coupled Receptors

Heterologous expression in mammalian cell lines provides an invaluable tool with which to explore determinants of G protein-coupled receptor (GPCR) function. The use of stably transfected cell lines enables a receptor to be studied in the absence of other related receptor subtypes. With this technique, molecular determinants of ligand binding can be identified; signal transduction pathways of a particular receptor can be delineated; interactions between a transfected receptor and other expressed receptors, both endogenous and exogenous, can be established; interactions between a GPCR and different G proteins can be characterized; and biochemical modifications of GPCRs, such as phosphorylation, can be studied. Many different features of GPCR function can be explored through the use of stably transfected cell lines. One of the major advantages of stably transfected cell lines is that a known, constant quantity of the transfected receptor is expressed for many generations. Both stably transfected pools and clonal cell lines can be studied. Stably transfected pools, or clonal cell lines, that express high levels of a transfected receptor facilitate ligand binding studies; cell lines expressing physiologically relevant levels of a transfected receptor can be used to examine second messenger-couplmg pathways. In planning to establish a stably transfected cell line expressing a GPCR of interest, there are a number of different factors that must be taken into consideration.