Co-immunoprecipitation as a Strategy to Evaluate Receptor–receptor or Receptor–protein Interactions

Immunoprecipitation is a useful method for isolating proteins of interest from cellular extracts using specific antibodies. Following immunoprecipitation of a protein of interest, it can be determined via Western blot whether any other proteins have co-immunoprecipitated. This method has been routinely used over the past several decades to study protein–protein interactions and thereby elucidate cellular signaling pathways. The interactions of G protein-coupled receptors (GPCRs) with a variety of protein partners have proven tractable to analysis via co-immunoprecipitation. For example, it is in some cases possible to co-immunoprecipitate G proteins with GPCRs (Matesic et al., 1989; Law et al., 1991; Matesic et al., 1991; Law and Reisine, 1992; Okuma and Reisine, 1992; Georgoussi et al., 1995; Sidhu et al., 1998; Chalecka-Franaszek et al., 2000). This has been a useful method for helping to characterize the specificity of G protein coupling for certain receptors. The associations of arrestins with GPCRs have also been effectively studied via co-immunoprecipitation (Luttrell et al., 1999; Cheng et al., 2000; Cen et al., 2001; Chen et al., 2002; Conlan et al., 2002; Kishi et al., 2002; Perry et al., 2002), as have interactions between GPCRs and various other cytoplasmic proteins (Table 9.1). Finally, co-immunoprecipitation has been an effective method for studying GPCR dimerization (Table 9.2). Receptor–receptor interactions characterized via co-immunoprecipitation