Abstract
Cyclic AMP controls several signalling cascades within cells, and changes in the amounts of this second messenger have an essential role in many cellular events. Here we describe a new methodology for monitoring the fluctuations of cAMP in living cells. By tagging the cAMP effector protein kinase A with two suitable green fluorescent protein mutants, we have generated a probe in which the fluorescence resonance energy transfer between the two fluorescent moieties is dependent on the levels of cAMP. This new methodology opens the way to the elucidation of the biochemistry of cAMP in vivo.
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Acknowledgements
We thank G. Ronconi and M. Santato for technical assistance. This work was supported by grants to T.P. from the Italian Association for Cancer Research (AIRC; grant number 98), from Telethon (grant 1226), from the CNR target project Biotechnology (grant 049), from the EU Programs Biotechnology (grants BIO4CT960382 and TMR FMRXCT960382), and from the Armenise Harvard Foundation, and to C.Y.C. and R.Y.T. from the Howard Hughes Medical Institute.
Correspondence and requests for materials should be addressed to T.P.
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Zaccolo, M., De Giorgi, F., Cho, C. et al. A genetically encoded, fluorescent indicator for cyclic AMP in living cells. Nat Cell Biol 2, 25–29 (2000). https://doi.org/10.1038/71345
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DOI: https://doi.org/10.1038/71345