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The protein kinase A anchoring protein mAKAP coordinates two integrated cAMP effector pathways

Abstract

Cyclic adenosine 3′, 5′-monophosphate (cAMP) is a ubiquitous mediator of intracellular signalling events. It acts principally through stimulation of cAMP-dependent protein kinases (PKAs)1,2 but also activates certain ion channels and guanine nucleotide exchange factors (Epacs)3. Metabolism of cAMP is catalysed by phosphodiesterases (PDEs)4,5. Here we identify a cAMP-responsive signalling complex maintained by the muscle-specific A-kinase anchoring protein (mAKAP) that includes PKA, PDE4D3 and Epac1. These intermolecular interactions facilitate the dissemination of distinct cAMP signals through each effector protein. Anchored PKA stimulates PDE4D3 to reduce local cAMP concentrations, whereas an mAKAP-associated ERK5 kinase module suppresses PDE4D3. PDE4D3 also functions as an adaptor protein that recruits Epac1, an exchange factor for the small GTPase Rap1, to enable cAMP-dependent attenuation of ERK5. Pharmacological and molecular manipulations of the mAKAP complex show that anchored ERK5 can induce cardiomyocyte hypertrophy. Thus, two coupled cAMP-dependent feedback loops are coordinated within the context of the mAKAP complex, suggesting that local control of cAMP signalling by AKAP proteins is more intricate than previously appreciated.

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Figure 1: Bidirectional control of the mAKAP-associated PDE4D3 activity.
Figure 2: Epac1 suppresses mAKAP-associated ERK5 activity.
Figure 3: The mAKAP complex facilitates cytokine-induced cardiac hypertrophy.

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Acknowledgements

This work was supported by grants from the National Institutes of Health (to J.D.S. and M.S.K.) and the American Heart Association (to K.L.D.-K.). The authors wish to thank N. Mayer, D. Bleckinger and R. Mouton for technical assistance, and R. Tsien, M. Houslay, J. E. Dixon, J. L. Bos and P. Stork for reagents.

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Correspondence to John D. Scott.

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Supplementary information

Supplementary Notes

This file contains the Supplementary Figure Legends and the Supplementary Video Legends. (DOC 32 kb)

Supplementary Video S1

This representative movie shows HeLa cells transfected with the AKAR-PKA reporter and treated with forskolin. (MOV 996 kb)

Supplementary Video S2

This representative movie shows HeLa cells transfected with the AKAR-PKA-PDE reporter and treated with forskolin. (MOV 817 kb)

Supplementary Video S3

This representative movie shows HeLa cells transfected with the AKAR-PKA-PDE reporter, pre-treated with either milrinone or rolipram, and then treated with forskolin. (MOV 1365 kb)

Supplementary Figures S1 and S2

Supplementary Figure S1: mAKAP and ERK5 co-localize at the nuclear membrane of hypertrophic rat neonatal ventriculocytes (RNV). Supplementary Figure S2: mAKAP scaffolds PDE4D3 and ERK5 to form a ternary complex (PDF 126 kb)

Supplementary Figures S3 and S4

Supplementary Figure S3: mapping of the ERK5 binding fragment on mAKAP. Supplementary Figure S4: PDE4D3 links ERK5 to mAKAP. (PDF 99 kb)

Supplementary Figures S5 and S6

Supplementary Figure S5. mAKAP-anchored PDE activity is negatively regulated by ERK. Supplementary Figure S6. PDE4D3 serves as an adapter protein to tether Epac1 to the mAKAP complex. (PDF 94 kb)

Supplementary Figures S7 and S8

Supplementary Figure S7: PDE4D3 tethers Epac1 to the mAKAP complex. Supplementary Figure S8: PDE4D3 binds directly to both Epac1 and ERK5. (PDF 84 kb)

Supplementary Figures S9 and S10

Supplementary Figure S9: mAKAP silenced RNV exhibit reduced hypertrophy in response to LIF stimulation. Supplementary Figure S10: expression of the mAKAP 585-1286 fragment displaces endogenous mAKAP and ablates LIF induced hypertrophy in RNV. (PDF 140 kb)

Supplementary Figures S11 and S12

Supplementary Figure S11: mAKAP plays a role in LIF-induced cardiac hypertrophy. Supplementary Figure S12: displacement of mAKAP inhibits LIF-induced expression of ANF, an indicator of hypertrophy. (PDF 143 kb)

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Dodge-Kafka, K., Soughayer, J., Pare, G. et al. The protein kinase A anchoring protein mAKAP coordinates two integrated cAMP effector pathways. Nature 437, 574–578 (2005). https://doi.org/10.1038/nature03966

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