Activation of G-protein-coupled receptors (GPCRs) mobilizes compartmentalized pulses of cyclic AMP. The main cellular effector of cAMP is protein kinase A (PKA), which is assembled as an inactive holoenzyme consisting of two regulatory (R) and two catalytic (PKAc) subunits. cAMP binding to R subunits dissociates the holoenzyme and releases the catalytic moiety, which phosphorylates a wide array of cellular proteins. Reassociation of PKAc and R components terminates the signal. Here we report that the RING ligase praja2 controls the stability of mammalian R subunits. Praja2 forms a stable complex with, and is phosphorylated by, PKA. Rising cAMP levels promote praja2-mediated ubiquitylation and subsequent proteolysis of compartmentalized R subunits, leading to sustained substrate phosphorylation by the activated kinase. Praja2 is required for efficient nuclear cAMP signalling and for PKA-mediated long-term memory. Thus, praja2 regulates the total concentration of R subunits, tuning the strength and duration of PKA signal output in response to cAMP.
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This work was supported by Grants from Associazione Italiana per la Ricerca sul Cancro (AIRC) and the Italian Ministry of University and Research (MIUR) 2008–2009 (2007KS47FW_002). E.S. was supported from Austrian Science Fund (FWF) Grant P22608 and Junior Researcher Support 2009 (University of Innsbruck). Special thanks to M. Gottesman, E. Avvedimento and K. Bister for discussion and critical reading of the manuscript, D. Viaggiano for help in immunohistochemistry and confocal microscopy analysis, R. D. Donne for help in carrying out experiments and S. McKnight and M. Ginsberg for providing the vectors for epitope-tagged R subunits. This manuscript is dedicated to the memory of M. Graziano.
The authors declare no competing financial interests.
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Lignitto, L., Carlucci, A., Sepe, M. et al. Control of PKA stability and signalling by the RING ligase praja2. Nat Cell Biol 13, 412–422 (2011). https://doi.org/10.1038/ncb2209
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