A Novel Epac-specific cAMP Analogue Demonstrates Independent Regulation of Rap1 and ERK

Nat Cell Biol. 2002 Nov;4(11):901-6. doi: 10.1038/ncb874.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 8-Bromo Cyclic Adenosine Monophosphate / analogs & derivatives*
  • 8-Bromo Cyclic Adenosine Monophosphate / metabolism
  • Amino Acid Sequence
  • Animals
  • CHO Cells
  • Cell Line
  • Cell Line, Tumor
  • Cricetinae
  • Cyclic AMP / metabolism*
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Dose-Response Relationship, Drug
  • Enzyme Activation
  • Enzyme Inhibitors / pharmacology
  • Gene Expression Regulation*
  • Guanine Nucleotide Exchange Factors / metabolism*
  • Humans
  • In Vitro Techniques
  • Mice
  • Mitogen-Activated Protein Kinases / metabolism*
  • Molecular Sequence Data
  • NIH 3T3 Cells
  • Phosphorylation
  • Protein Structure, Tertiary
  • Sequence Homology, Amino Acid
  • Signal Transduction
  • Time Factors
  • Transfection
  • rap1 GTP-Binding Proteins / metabolism*

Substances

  • Enzyme Inhibitors
  • Guanine Nucleotide Exchange Factors
  • RAPGEF3 protein, human
  • 8-Bromo Cyclic Adenosine Monophosphate
  • 8-chloro-cyclic adenosine monophosphate
  • Cyclic AMP
  • Cyclic AMP-Dependent Protein Kinases
  • Mitogen-Activated Protein Kinases
  • rap1 GTP-Binding Proteins