EFFICIENT MEMBRANE TARGETING OF THE CHICK NICOTINIC ACETYLCHOLINE-RECEPTOR ALPHA-SUBUNIT IN STABLE INSECT-CELL LINES

We have synthesised the alpha-subunic of the chick nicotinic acetylcho line receptor (nAChR) in stable, continuous insect (Spodoptera frugipe rda) cell lines. A cDNA was integrated randomly into the insect cell g enome under control of a baculovius immediate early gene promoter. Tra nsformed cells were obtained by co-transfection of the insect cells wi th pIEK1.nAChR alpha, encoding the alpha-subunit cDNA, and pIEK1.neo, encoding the neomycin resistance gene. G-418-resistant clones were sel ected and expanded into continuous cell lines synthesising the chick n AChR a-subunit. Using fluorescence microscopy and ligand binding studi es we were able to demonstrate efficient membrane targeting of the rec eptor subunit in the insect cell plasma membrane. Stable insect cell l ines may thus have significant advantages over transient baculovirus v ectors for the synthesis and characterisation of heterologous receptor proteins.