Insulin disrupts β-adrenergic signalling to protein kinase A in adipocytes

Abstract

Hormones mobilize intracellular second messengers and initiate signalling cascades involving protein kinases and phosphatases, which are often spatially compartmentalized by anchoring proteins to increase signalling specificity1. These scaffold proteins may themselves be modulated by hormones2,3,4. In adipocytes, stimulation of β-adrenergic receptors increases cyclic AMP levels and activates protein kinase A (PKA)5, which stimulates lipolysis by phosphorylating hormone-sensitive lipase and perilipin6,7,8. Acute insulin treatment activates phosphodiesterase 3B, reduces cAMP levels and quenches β-adrenergic receptor signalling9. In contrast, chronic hyperinsulinaemic conditions (typical of type 2 diabetes) enhance β-adrenergic receptor-mediated cAMP production10. This amplification of cAMP signalling is paradoxical because it should enhance lipolysis, the opposite of the known short-term effect of hyperinsulinaemia. Here we show that in adipocytes, chronically high insulin levels inhibit β-adrenergic receptors (but not other cAMP-elevating stimuli) from activating PKA. We measured this using an improved fluorescent reporter and by phosphorylation of endogenous cAMP-response-element binding protein (CREB). Disruption of PKA scaffolding mimics the interference of insulin with β-adrenergic receptor signalling. Chronically high insulin levels may disrupt the close apposition of β-adrenergic receptors and PKA, identifying a new mechanism for crosstalk between heterologous signal transduction pathways.

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Figure 1: Re-engineering of AKAR2.
Figure 2: Effect of insulin pretreatment on PKA activity.
Figure 3: The insulin-induced delay in PKA activity is specific for a β-adrenergic-coupled pool of PKA.
Figure 4: β-adrenergic-coupled pool of PKA.

References

  1. 1

    Wong, W. & Scott, J. D. AKAP signalling complexes: focal points in space and time. Nature Rev. Mol. Cell Biol. 5, 959–970 (2004)

    CAS  Article  Google Scholar 

  2. 2

    Salvador, L. M. et al. Neuronal microtubule-associated protein 2D is a dual A-kinase anchoring protein expressed in rat ovarian granulosa cells. J. Biol. Chem. 279, 27621–27632 (2004)

    CAS  Article  Google Scholar 

  3. 3

    Carr, D. W., DeManno, D. A., Atwood, A., Hunzicker-Dunn, M. & Scott, J. D. Follicle-stimulating hormone regulation of A-kinase anchoring proteins in granulosa cells. J. Biol. Chem. 268, 20729–20732 (1993)

    CAS  PubMed  Google Scholar 

  4. 4

    Feliciello, A., Rubin, C. S., Avvedimento, E. V. & Gottesman, M. E. Expression of a kinase anchor protein 121 is regulated by hormones in thyroid and testicular germ cells. J. Biol. Chem. 273, 23361–23366 (1998)

    CAS  Article  Google Scholar 

  5. 5

    Taylor, S. S. et al. PKA: a portrait of protein kinase dynamics. Biochim. Biophys. Acta 1697, 259–269 (2004)

    CAS  Article  Google Scholar 

  6. 6

    Londos, C., Honnor, R. C. & Dhillon, G. S. cAMP-dependent protein kinase and lipolysis in rat adipocytes. III. Multiple modes of insulin regulation of lipolysis and regulation of insulin responses by adenylate cyclase regulators. J. Biol. Chem. 260, 15139–15145 (1985)

    CAS  PubMed  Google Scholar 

  7. 7

    Egan, J. J., Greenberg, A. S., Chang, M. K. & Londos, C. Control of endogenous phosphorylation of the major cAMP-dependent protein kinase substrate in adipocytes by insulin and beta-adrenergic stimulation. J. Biol. Chem. 265, 18769–18775 (1990)

    CAS  PubMed  Google Scholar 

  8. 8

    Greenberg, A. S. et al. Perilipin, a major hormonally regulated adipocyte-specific phosphoprotein associated with the periphery of lipid storage droplets. J. Biol. Chem. 266, 11341–11346 (1991)

    CAS  PubMed  Google Scholar 

  9. 9

    Elks, M. L. & Manganiello, V. C. Antilipolytic action of insulin: role of cAMP phosphodiesterase activation. Endocrinology 116, 2119–2121 (1985)

    CAS  Article  Google Scholar 

  10. 10

    Hupfeld, C. J., Dalle, S. & Olefsky, J. M. β-Arrestin 1 down-regulation after insulin treatment is associated with supersensitization of β2 adrenergic receptor Gαs signalling in 3T3–L1 adipocytes. Proc. Natl Acad. Sci. USA 100, 161–166 (2003)

    ADS  CAS  Article  Google Scholar 

  11. 11

    Zhang, J., Ma, Y., Taylor, S. S. & Tsien, R. Y. Genetically encoded reporters of protein kinase A activity reveal impact of substrate tethering. Proc. Natl Acad. Sci. USA 98, 14997–15002 (2001)

    ADS  CAS  Article  Google Scholar 

  12. 12

    Muslin, A. J., Tanner, J. W., Allen, P. M. & Shaw, A. S. Interaction of 14-3-3 with signalling proteins is mediated by the recognition of phosphoserine. Cell 84, 889–897 (1996)

    CAS  Article  Google Scholar 

  13. 13

    Durocher, D. et al. The molecular basis of FHA domain:phosphopeptide binding specificity and implications for phospho-dependent signalling mechanisms. Mol. Cell 6, 1169–1182 (2000)

    CAS  Article  Google Scholar 

  14. 14

    Nerbonne, J. M., Richard, S., Nargeot, J. & Lester, H. A. New photoactivatable cyclic nucleotides produce intracellular jumps in cyclic AMP and cyclic GMP concentrations. Nature 310, 74–76 (1984)

    ADS  CAS  Article  Google Scholar 

  15. 15

    Kosaki, A., Yamada, K. & Kuzuya, H. Reduced expression of the leptin gene (ob) by catecholamine through a Gs protein-coupled pathway in 3T3–L1 adipocytes. Diabetes 45, 1744–1749 (1996)

    CAS  Article  Google Scholar 

  16. 16

    Fasshauer, M., Klein, J., Neumann, S., Eszlinger, M. & Paschke, R. Adiponectin gene expression is inhibited by β-adrenergic stimulation via protein kinase A in 3T3–L1 adipocytes. FEBS Lett. 507, 142–146 (2001)

    CAS  Article  Google Scholar 

  17. 17

    Fasshauer, M., Klein, J., Neumann, S., Eszlinger, M. & Paschke, R. Isoproterenol inhibits resistin gene expression through a Gs-protein-coupled pathway in 3T3–L1 adipocytes. FEBS Lett. 500, 60–63 (2001)

    CAS  Article  Google Scholar 

  18. 18

    Mayr, B. & Montminy, M. Transcriptional regulation by the phosphorylation-dependent factor CREB. Nature Rev. Mol. Cell Biol. 2, 599–609 (2001)

    CAS  Article  Google Scholar 

  19. 19

    Scott, J. D. & Faux, M. C. Use of synthetic peptides in the dissection of protein-targeting interactions. Methods Mol. Biol. 88, 161–185 (1998)

    CAS  PubMed  Google Scholar 

  20. 20

    Fraser, I. D. et al. Assembly of an A kinase-anchoring protein–β2-adrenergic receptor complex facilitates receptor phosphorylation and signalling. Curr. Biol. 10, 409–412 (2000)

    CAS  Article  Google Scholar 

  21. 21

    Lin, F., Wang, H. & Malbon, C. C. Gravin-mediated formation of signalling complexes in β2-adrenergic receptor desensitization and resensitization. J. Biol. Chem. 275, 19025–19034 (2000)

    CAS  Article  Google Scholar 

  22. 22

    Jurevicius, J. & Fischmeister, R. cAMP compartmentation is responsible for a local activation of cardiac Ca2+ channels by β-adrenergic agonists. Proc. Natl Acad. Sci. USA 93, 295–299 (1996)

    ADS  CAS  Article  Google Scholar 

  23. 23

    Zaccolo, M. & Pozzan, T. Discrete microdomains with high concentration of cAMP in stimulated rat neonatal cardiac myocytes. Science 295, 1711–1715 (2002)

    ADS  CAS  Article  Google Scholar 

  24. 24

    Tao, J., Wang, H. Y. & Malbon, C. C. Protein kinase A regulates AKAP250 (gravin) scaffold binding to the β2-adrenergic receptor. EMBO J. 22, 6419–6429 (2003)

    CAS  Article  Google Scholar 

  25. 25

    Griesbeck, O., Baird, G. S., Campbell, R. E., Zacharias, D. A. & Tsien, R. Y. Reducing the environmental sensitivity of yellow fluorescent protein. Mechanism and applications. J. Biol. Chem. 276, 29188–29194 (2001)

    CAS  Article  Google Scholar 

  26. 26

    Dalle, S. et al. Insulin induces heterologous desensitization of G-protein-coupled receptor and insulin-like growth factor I signalling by downregulating β-arrestin-1. Mol. Cell. Biol. 22, 6272–6285 (2002)

    CAS  Article  Google Scholar 

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Acknowledgements

We thank Y. Ma for technical assistance and discussions and M. B. Yaffe for providing FHA cDNA. This work was supported by the Howard Hughes Medical Institute, the Alliance for Cell Signaling (R.Y.T.), the NIH (R.Y.T, C.J.H. and S.S.T) and the Johns Hopkins University School of Medicine and W. M. Keck Foundation (J.Z.). Author Contributions J.Z. and C.J.H. performed all the experiments; all authors contributed to planning the experiments, interpreting the data and writing the paper.

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Correspondence to Roger Y. Tsien.

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Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Figures

Supplementary Figures S1-S4, showing effect of insulin treatment on βAR-associated cAMP generation, effect of insulin treatment on PKA activity in the nucleus, effect of the functionally inactive Ht31p, and a model for insulin modulated compartmentation in adipocytes. (PDF 93 kb)

Supplementary Methods

Additional information about the imaging methods used in this study. (DOC 24 kb)

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Zhang, J., Hupfeld, C., Taylor, S. et al. Insulin disrupts β-adrenergic signalling to protein kinase A in adipocytes. Nature 437, 569–573 (2005). https://doi.org/10.1038/nature04140

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