title: cAMP regulates DEP domain-mediated binding of the guanine nucleotide exchange factor Epac1 to phosphatidic acid at the plasma membrane.
abstract: Epac1 is a cAMP-regulated guanine nucleotide exchange factor for the small G protein Rap. Upon cAMP binding, Epac1 undergoes a conformational change that results in its release from autoinhibition. In addition, cAMP induces the translocation of Epac1 from the cytosol to the plasma membrane. This relocalization of Epac1 is required for efficient activation of plasma membrane-located Rap and for cAMP-induced cell adhesion. This translocation requires the Dishevelled, Egl-10, Pleckstrin (DEP) domain, but the molecular entity that serves as the plasma membrane anchor and the possible mechanism of regulated binding remains elusive. Here we show that Epac1 binds directly to phosphatidic acid. Similar to the cAMP-induced Epac1 translocation, this binding is regulated by cAMP and requires the DEP domain. Furthermore, depletion of phosphatidic acid by inhibition of phospholipase D1 prevents cAMP-induced translocation of Epac1 as well as the subsequent activation of Rap at the plasma membrane. Finally, mutation of a single basic residue within a polybasic stretch of the DEP domain, which abolishes translocation, also prevents binding to phosphatidic acid. From these results we conclude that cAMP induces a conformational change in Epac1 that enables DEP domain-mediated binding to phosphatidic acid, resulting in the tethering of Epac1 at the plasma membrane and subsequent activation of Rap.
authors.affiliation: Molecular Cancer Research, University Medical Center Utrecht, 3584 CG, Utrecht, The Netherlands.
authors.forename: Sarah V
authors.initials: SV
authors.lastname: Consonni
authors.forename: Martijn
authors.initials: M
authors.lastname: Gloerich
authors.forename: Emma
authors.initials: E
authors.lastname: Spanjaard
authors.forename: Johannes L
authors.initials: JL
authors.lastname: Bos
mesh_terms: D002462:Cell Membrane / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008055:Lipids / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry; D010712:Phosphatidic Acids / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011994:Recombinant Proteins / Q000737:chemistry; D020827:rap1 GTP-Binding Proteins / Q000737:chemistry
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D008055:Lipids; D008081:Liposomes; D010712:Phosphatidic Acids; C430803:RAP1A protein, human; C116122:RAPGEF3 protein, human; D011994:Recombinant Proteins; D000242:Cyclic AMP; D020827:rap1 GTP-Binding Proteins