Direct binding of the beta1 adrenergic receptor to the cyclic AMP-dependent guanine nucleotide exchange factor CNrasGEF leads to Ras activation.
G-protein-coupled receptors (GPCRs) can indirectly activate Ras primarily through the betagamma 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 beta1 adrenergic receptor (beta1-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 beta1-AR and CNrasGEF, Ras is activated by the beta1-AR agonist isoproterenol, and this activation is abolished in beta1-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 Gsalpha and not via Gbetagamma. In contrast to beta1-AR, the beta2-AR neither binds CNrasGEF nor activates Ras via CNrasGEF after agonist stimulation. These results suggest a model whereby the physical interaction between the beta1-AR and CNrasGEF facilitates the transduction of Gsalpha-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
Pak
Youngshil
Y
Program in Cell Biology, The Hospital for Sick Children, and Biochemistry Department, University of Toronto, Toronto, M5G 1X8 Ontario, Canada.
Pham
Nam
N
Rotin
Daniela
D
D000818:Animals; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000378:metabolism; D049452:Green Fluorescent Proteins; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006801:Humans; D008024:Ligands; D008164:Luminescent Proteins / Q000378:metabolism; D008954:Models, Biological; D009419:Nerve Tissue Proteins; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D051381:Rats; D018342:Receptors, Adrenergic, beta-1 / Q000235:genetics / Q000378:metabolism*; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013552:Swine; D018631:ras Proteins / Q000378:metabolism*
D020662:Guanine Nucleotide Exchange Factors; D008024:Ligands; D008164:Luminescent Proteins; D009419:Nerve Tissue Proteins; D021122:Protein Subunits; C403525:RAPGEF2 protein, human; D018342:Receptors, Adrenergic, beta-1; D018343:Receptors, Adrenergic, beta-2; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D000242:Cyclic AMP; D019205:GTP-Binding Protein alpha Subunits, Gs; D018631:ras Proteins