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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)1. 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)4,5. 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.

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Figure 1: Identification of an Epac-specific cAMP analogue.
Figure 2: ERK is inhibited by 8-Br-cAMP, but not by 8CPT-2Me-cAMP.
Figure 3: cAMP-induced activation of ERK and Rap1 are distinct processes.
Figure 4: cAMP-induced ERK activation is mediated by PKA and Ras.

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Acknowledgements

We thank R. A. J. Challiss for kindly providing the CHO-β2M3 cell line, and W. Bokkers and A.G.M. van Gorp for technical support. We also thank members of our laboratories for continuous support, discussions and critical reading of the manuscript. This research was supported by a grant from the Council of Earth and Life Sciences of The Netherlands Organisation for Scientific Research (NWO-ALW).

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Correspondence to Johannes L. Bos.

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H.G. Genieser owns BIOLOG Life Science Institute, which will sell 8-pCPT-2′OMe-cAMP for research purposes.

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Enserink, J., Christensen, A., de Rooij, J. et al. A novel Epac-specific cAMP analogue demonstrates independent regulation of Rap1 and ERK. Nat Cell Biol 4, 901–906 (2002). https://doi.org/10.1038/ncb874

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