Direct Binding of the β1 Adrenergic Receptor to the Cyclic AMP-Dependent Guanine Nucleotide Exchange Factor CNrasGEF Leads to Ras Activation

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Abstract
G-protein-coupled receptors (GPCRs) can indirectly activate Ras primarily through the βγ subunits of G proteins, which recruit c-Src, phosphatidylinositol 3-kinase, and Grb2-SOS. However, a direct interaction between a Ras activator (guanine nucleotide exchange factor [GEF]) and GPCRs that leads to Ras activation has never been demonstrated. We report here a novel mechanism for a direct GPCR-mediated Ras activation. The β1 adrenergic receptor (β1-AR) binds to the PDZ domain of the cyclic AMP (cAMP)-dependent Ras exchange factor, CNrasGEF, via its C-terminal SkV motif. In cells heterologously expressing β1-AR and CNrasGEF, Ras is activated by the β1-AR agonist isoproterenol, and this activation is abolished in β1-AR mutants that cannot bind CNrasGEF or in CNrasGEF mutants lacking the catalytic CDC25 domain or cAMP-binding domain. Moreover, the activation is transduced via Gsα and not via Gβγ. In contrast to β1-AR, the β2-AR neither binds CNrasGEF nor activates Ras via CNrasGEF after agonist stimulation. These results suggest a model whereby the physical interaction between the β1-AR and CNrasGEF facilitates the transduction of Gsα-induced cAMP signal into the activation of Ras. The present study provides the first demonstration of direct physical association between a Ras activator and a GPCR, leading to agonist-induced Ras activation