title: A novel Epac-specific cAMP analogue demonstrates independent regulation of Rap1 and ERK.
abstract: cAMP is involved in a wide variety of cellular processes that were thought to be mediated by protein kinase A (PKA). However, cAMP also directly regulates Epac1 and Epac2, guanine nucleotide-exchange factors (GEFs) for the small GTPases Rap1 and Rap2 (refs 2,3). Unfortunately, there is an absence of tools to discriminate between PKA- and Epac-mediated effects. Therefore, through rational drug design we have developed a novel cAMP analogue, 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate (8CPT-2Me-cAMP), which activates Epac, but not PKA, both in vitro and in vivo. Using this analogue, we tested the widespread model that Rap1 mediates cAMP-induced regulation of the extracellular signal-regulated kinase (ERK). However, both in cell lines in which cAMP inhibits growth-factor-induced ERK activation and in which cAMP activates ERK, 8CPT-2Me-cAMP did not affect ERK activity. Moreover, in cell lines in which cAMP activates ERK, inhibition of PKA and Ras, but not Rap1, abolished cAMP-mediated ERK activation. We conclude that cAMP-induced regulation of ERK and activation of Rap1 are independent processes.
authors.affiliation: Department of Physiological Chemistry and Centre for Biomedical Genetics, University Medical Center Utrecht, Universiteitsweg 100, 3584CG Utrecht, The Netherlands.
authors.forename: Jorrit M
authors.initials: JM
authors.lastname: Enserink
authors.forename: Anne E
authors.initials: AE
authors.lastname: Christensen
authors.forename: Johan
authors.initials: J
authors.lastname: de Rooij
authors.forename: Miranda
authors.initials: M
authors.lastname: van Triest
authors.forename: Frank
authors.initials: F
authors.lastname: Schwede
authors.forename: Hans Gottfried
authors.initials: HG
authors.lastname: Genieser
authors.forename: Stein O
authors.initials: SO
authors.lastname: Døskeland
authors.forename: Jonathan L
authors.initials: JL
authors.lastname: Blank
authors.forename: Johannes L
authors.initials: JL
authors.lastname: Bos
mesh_terms: D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000031:analogs & derivatives* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D045744:Cell Line, Tumor; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D020827:rap1 GTP-Binding Proteins / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D020662:Guanine Nucleotide Exchange Factors; C116122:RAPGEF3 protein, human; D015124:8-Bromo Cyclic Adenosine Monophosphate; C044258:8-chloro-cyclic adenosine monophosphate; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D020827:rap1 GTP-Binding Proteins