Retrovirus-mediated expression of the GalR1 galanin receptor: implication for efficient stable expression of functional G protein-coupled receptors.
The rat GalR1 galanin receptor was used as a prototypic G protein-coupled receptor to test the feasibility of heterologous expression in a retrovirus-based system. The system utilizes an independent retroviral vector pMX, a virus-packaging cell line BOSC23 and a pre-B cell line BA/F3 as the host for expression. A polyclonal cell population that expresses high ligand affinity (KD = 0.18 nM) and high level (7 pmol/mg) of GalR1 was generated within days with no drug sensitivity-based selection. The expression represented a 20-fold increase over the expression level of GalR1 achieved in CHO cells. The affinity of galanin for the expressed receptor was decreased by 19-fold in the presence of GTP-gamma-S, suggesting that the expression system can produce active galanin receptor functionally coupled to G proteins. The fast and efficient method to generate stable cell lines and to prepared large quantities of receptors may provide a general application for expression of other G protein-coupled receptors.
Wang
S
S
Department of CNS/CV Biological Research, Schering-Plough Research Institute, Kenilworth, NJ 07033, USA.
Clemmons
A
A
Bayne
M
M
Graziano
M P
MP
D000818:Animals; D002460:Cell Line; D019204:GTP-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D019004:Galanin / Q000378:metabolism; D005822:Genetic Vectors; D051381:Rats; D044088:Receptors, Galanin; D018013:Receptors, Neuropeptide / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis*; D012190:Retroviridae / Q000235:genetics*
D044088:Receptors, Galanin; D018013:Receptors, Neuropeptide; D011994:Recombinant Proteins; D019004:Galanin; D019204:GTP-Binding Proteins