Analysis of EGF receptor interactions with the alpha subunit of the stimulatory GTP binding protein of adenylyl cyclase, Gs.

Besides stimulating the mitogen-activated protein kinase, phospholipase Cgamma, and phosphatidylinositol 3-kinase cascades, in certain tissues and cells such as the heart, partotid gland, and luteal cells, activation of the epidermal growth factor (EGF) receptor also stimulates second-messenger systems that involve the heterotrimeric G proteins. For instance, in the heart EGF increases contractility and heart rate by elevating cellular cyclic adenosine monophosphate (cAMP) levels. This is the result of EGF-elicited activation of adenylyl cyclase via the stimulatory guanosine 5'-triphosphate (GTP)-binding protein Gs. In this context, the single transmembrane EGF receptor acts like a heptahelical G protein-coupled receptor. Here we have described the methods used to study interactions between the EGF receptor and heterotrimeric G proteins. Moreover, we have also described how the stoichiometry of EGF receptor association with the alpha subunit of Gs can be monitored in vitro. Several other single transmembrane receptors and proteins can also activate heterotrimeric G proteins, and, therefore, the methodologies described in this chapter can be adapted to other systems.