Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Insulin regulates liver metabolism in vivo in the absence of hepatic Akt and Foxo1

Abstract

Considerable data support the idea that forkhead box O1 (Foxo1) drives the liver transcriptional program during fasting and is then inhibited by thymoma viral proto-oncogene 1 (Akt) after feeding. Here we show that mice with hepatic deletion of Akt1 and Akt2 were glucose intolerant, insulin resistant and defective in their transcriptional response to feeding in the liver. These defects were normalized with concomitant liver-specific deletion of Foxo1. Notably, in the absence of both Akt and Foxo1, mice adapted appropriately to both the fasted and fed state, and insulin suppressed hepatic glucose production normally. A gene expression analysis revealed that deletion of Akt in liver led to the constitutive activation of Foxo1-dependent gene expression, but again, concomitant ablation of Foxo1 restored postprandial regulation, preventing the inhibition of the metabolic response to nutrient intake caused by deletion of Akt. These results are inconsistent with the canonical model of hepatic metabolism in which Akt is an obligate intermediate for proper insulin signaling. Rather, they show that a major role of hepatic Akt is to restrain the activity of Foxo1 and that in the absence of Foxo1, Akt is largely dispensable for insulin- and nutrient-mediated hepatic metabolic regulation in vivo.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Liver-specific deletion of Akt1 in Akt2 whole-body knockout mice resulted in severe hyperglycemia and the disruption of Foxo1-regulated gene expression.
Figure 2: Aberrant insulin signaling and disrupted expression of Foxo1-regulated genes in DLKO but not 2LKO livers.
Figure 3: Deletion of both Akt1 and Akt2 in liver results in hyperglycemia, glucose intolerance and insulin resistance.
Figure 4: Deletion of Foxo1 normalizes hyperglycemia, glucose intolerance and hyperinsulinemia in DLKO livers despite defective insulin signaling.
Figure 5: Nutritional regulation of hepatic gene expression in the absence of Foxo1 alone and in the absence of both Akt and Foxo1.
Figure 6: Insulin-regulated expression of Foxo1 target genes was compromised in the Foxo1−/− and TLKO primary hepatocytes.

Similar content being viewed by others

References

  1. Saltiel, A.R. & Kahn, C.R. Insulin signalling and the regulation of glucose and lipid metabolism. Nature 414, 799–806 (2001).

    Article  CAS  PubMed  Google Scholar 

  2. White, M.F. Insulin signaling in health and disease. Science 302, 1710–1711 (2003).

    Article  CAS  PubMed  Google Scholar 

  3. Brown, M.S. & Goldstein, J.L. Selective versus total insulin resistance: a pathogenic paradox. Cell Metab. 7, 95–96 (2008).

    Article  CAS  PubMed  Google Scholar 

  4. Cho, H. et al. Insulin resistance and a diabetes mellitus-like syndrome in mice lacking the protein kinase Akt2 (PKB β). Science 292, 1728–1731 (2001).

    Article  CAS  PubMed  Google Scholar 

  5. Michael, M.D. et al. Loss of insulin signaling in hepatocytes leads to severe insulin resistance and progressive hepatic dysfunction. Mol. Cell 6, 87–97 (2000).

    Article  CAS  PubMed  Google Scholar 

  6. Dong, X.C. et al. Inactivation of hepatic Foxo1 by insulin signaling is required for adaptive nutrient homeostasis and endocrine growth regulation. Cell Metab. 8, 65–76 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Chen, W.S. et al. Growth retardation and increased apoptosis in mice with homozygous disruption of the Akt1 gene. Genes Dev. 15, 2203–2208 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Jacinto, E. et al. SIN1/MIP1 maintains rictor-mTOR complex integrity and regulates Akt phosphorylation and substrate specificity. Cell 127, 125–137 (2006).

    Article  CAS  PubMed  Google Scholar 

  9. Rintelen, F., Stocker, H., Thomas, G. & Hafen, E. PDK1 regulates growth through Akt and S6K in Drosophila. Proc. Natl. Acad. Sci. USA 98, 15020–15025 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Alessi, D.R. et al. Characterization of a 3-phosphoinositide–dependent protein kinase which phosphorylates and activates protein kinase Balpha. Curr. Biol. 7, 261–269 (1997).

    Article  CAS  PubMed  Google Scholar 

  11. Inoki, K., Li, Y., Zhu, T., Wu, J. & Guan, K.L. TSC2 is phosphorylated and inhibited by Akt and suppresses mTOR signalling. Nat. Cell Biol. 4, 648–657 (2002).

    Article  CAS  PubMed  Google Scholar 

  12. Sengupta, S., Peterson, T.R. & Sabatini, D.M. Regulation of the mTOR complex 1 pathway by nutrients, growth factors, and stress. Mol. Cell 40, 310–322 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Düvel, K. et al. Activation of a metabolic gene regulatory network downstream of mTOR complex 1. Mol. Cell 39, 171–183 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  14. Porstmann, T. et al. SREBP activity is regulated by mTORC1 and contributes to Akt-dependent cell growth. Cell Metab. 8, 224–236 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Laplante, M. & Sabatini, D.M. mTORC1 activates SREBP-1c and uncouples lipogenesis from gluconeogenesis. Proc. Natl. Acad. Sci. USA 107, 3281–3282 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Li, S., Brown, M.S. & Goldstein, J.L. Bifurcation of insulin signaling pathway in rat liver: mTORC1 required for stimulation of lipogenesis, but not inhibition of gluconeogenesis. Proc. Natl. Acad. Sci. USA 107, 3441–3446 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Chakrabarti, P., English, T., Shi, J., Smas, C.M. & Kandror, K.V. Mammalian target of rapamycin complex 1 suppresses lipolysis, stimulates lipogenesis, and promotes fat storage. Diabetes 59, 775–781 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Kim, K.H. et al. Regulatory role of glycogen synthase kinase 3 for transcriptional activity of ADD1/SREBP1c. J. Biol. Chem. 279, 51999–52006 (2004).

    Article  CAS  PubMed  Google Scholar 

  19. Rena, G., Guo, S., Cichy, S.C., Unterman, T.G. & Cohen, P. Phosphorylation of the transcription factor forkhead family member FKHR by protein kinase B. J. Biol. Chem. 274, 17179–17183 (1999).

    Article  CAS  PubMed  Google Scholar 

  20. Nakae, J., Park, B.C. & Accili, D. Insulin stimulates phosphorylation of the forkhead transcription factor FKHR on serine 253 through a Wortmannin-sensitive pathway. J. Biol. Chem. 274, 15982–15985 (1999).

    Article  CAS  PubMed  Google Scholar 

  21. Brunet, A. et al. Akt promotes cell survival by phosphorylating and inhibiting a Forkhead transcription factor. Cell 96, 857–868 (1999).

    Article  CAS  PubMed  Google Scholar 

  22. Cheng, Z. & White, M.F. Targeting Forkhead box O1 from the concept to metabolic diseases: lessons from mouse models. Antioxid. Redox Signal. 14, 649–661 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Puigserver, P. et al. Insulin-regulated hepatic gluconeogenesis through FOXO1-PGC-1α interaction. Nature 423, 550–555 (2003).

    Article  CAS  PubMed  Google Scholar 

  24. Liu, Y. et al. A fasting inducible switch modulates gluconeogenesis via activator/coactivator exchange. Nature 456, 269–273 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Accili, D. & Arden, K.C. FoxOs at the crossroads of cellular metabolism, differentiation, and transformation. Cell 117, 421–426 (2004).

    Article  CAS  PubMed  Google Scholar 

  26. Zhang, W. et al. FoxO1 regulates multiple metabolic pathways in the liver: effects on gluconeogenic, glycolytic, and lipogenic gene expression. J. Biol. Chem. 281, 10105–10117 (2006).

    Article  CAS  PubMed  Google Scholar 

  27. Matsumoto, M., Han, S., Kitamura, T. & Accili, D. Dual role of transcription factor FoxO1 in controlling hepatic insulin sensitivity and lipid metabolism. J. Clin. Invest. 116, 2464–2472 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. O'Brien, R.M., Streeper, R.S., Ayala, J.E., Stadelmaier, B.T. & Hornbuckle, L.A. Insulin-regulated gene expression. Biochem. Soc. Trans. 29, 552–558 (2001).

    Article  CAS  PubMed  Google Scholar 

  29. Biggs, W.H. III, Meisenhelder, J., Hunter, T., Cavenee, W.K. & Arden, K.C. Protein kinase B/Akt-mediated phosphorylation promotes nuclear exclusion of the winged helix transcription factor FKHR1. Proc. Natl. Acad. Sci. USA 96, 7421–7426 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Matsumoto, M., Pocai, A., Rossetti, L., Depinho, R.A. & Accili, D. Impaired regulation of hepatic glucose production in mice lacking the forkhead transcription factor Foxo1 in liver. Cell Metab. 6, 208–216 (2007).

    Article  CAS  PubMed  Google Scholar 

  31. Haeusler, R.A., Kaestner, K.H. & Accili, D. FoxOs function synergistically to promote glucose production. J. Biol. Chem. 285, 35245–35248 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Altomonte, J. et al. Inhibition of Foxo1 function is associated with improved fasting glycemia in diabetic mice. Am. J. Physiol. Endocrinol. Metab. 285, E718–E728 (2003).

    Article  CAS  PubMed  Google Scholar 

  33. Samuel, V.T. et al. Targeting foxo1 in mice using antisense oligonucleotide improves hepatic and peripheral insulin action. Diabetes 55, 2042–2050 (2006).

    Article  CAS  PubMed  Google Scholar 

  34. Easton, R.M. et al. Role for Akt3/protein kinase Bγ in attainment of normal brain size. Mol. Cell. Biol. 25, 1869–1878 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Zhang, L., Rubins, N.E., Ahima, R.S., Greenbaum, L.E. & Kaestner, K.H. Foxa2 integrates the transcriptional response of the hepatocyte to fasting. Cell Metab. 2, 141–148 (2005).

    Article  PubMed  Google Scholar 

  36. Sakoda, H. et al. Differing roles of Akt and serum- and glucocorticoid-regulated kinase in glucose metabolism, DNA synthesis, and oncogenic activity. J. Biol. Chem. 278, 25802–25807 (2003).

    Article  CAS  PubMed  Google Scholar 

  37. Daitoku, H., Yamagata, K., Matsuzaki, H., Hatta, M. & Fukamizu, A. Regulation of PGC-1 promoter activity by protein kinase B and the forkhead transcription factor FKHR. Diabetes 52, 642–649 (2003).

    Article  CAS  PubMed  Google Scholar 

  38. Perrot, V. & Rechler, M.M. The coactivator p300 directly acetylates the forkhead transcription factor Foxo1 and stimulates Foxo1-induced transcription. Mol. Endocrinol. 19, 2283–2298 (2005).

    Article  CAS  PubMed  Google Scholar 

  39. van der Vos, K.E. & Coffer, P.J. The extending network of FOXO transcriptional target genes. Antioxid. Redox Signal. 14, 579–592 (2011).

    Article  CAS  PubMed  Google Scholar 

  40. Hirota, K. et al. A combination of HNF-4 and Foxo1 is required for reciprocal transcriptional regulation of glucokinase and glucose-6-phosphatase genes in response to fasting and feeding. J. Biol. Chem. 283, 32432–32441 (2008).

    Article  CAS  PubMed  Google Scholar 

  41. Inoki, K., Li, Y., Zhu, T., Wu, J. & Guan, K.L. TSC2 is phosphorylated and inhibited by Akt and suppresses mTOR signalling. Nat. Cell Biol. 4, 648–657 (2002).

    Article  CAS  PubMed  Google Scholar 

  42. Kovacina, K.S. et al. Identification of a proline-rich Akt substrate as a 14–3-3 binding partner. J. Biol. Chem. 278, 10189–10194 (2003).

    Article  CAS  PubMed  Google Scholar 

  43. Taniguchi, C.M., Emanuelli, B. & Kahn, C.R. Critical nodes in signalling pathways: insights into insulin action. Nat. Rev. Mol. Cell Biol. 7, 85–96 (2006).

    Article  CAS  PubMed  Google Scholar 

  44. Leavens, K.F., Easton, R.M., Shulman, G.I., Previs, S.F. & Birnbaum, M.J. Akt2 is required for hepatic lipid accumulation in models of insulin resistance. Cell Metab. 10, 405–418 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Slack, C., Giannakou, M.E., Foley, A., Goss, M. & Partridge, L. dFOXO-independent effects of reduced insulin-like signaling in Drosophila. Aging Cell 10, 735–748 (2011).

    Article  CAS  PubMed  Google Scholar 

  46. van der Horst, A. & Burgering, B.M. Stressing the role of FoxO proteins in lifespan and disease. Nat. Rev. Mol. Cell Biol. 8, 440–450 (2007).

    Article  CAS  PubMed  Google Scholar 

  47. Partridge, L. & Bruning, J.C. Forkhead transcription factors and ageing. Oncogene 27, 2351–2363 (2008).

    Article  CAS  PubMed  Google Scholar 

  48. Yamagata, K. et al. Arginine methylation of FOXO transcription factors inhibits their phosphorylation by Akt. Mol. Cell 32, 221–231 (2008).

    Article  CAS  PubMed  Google Scholar 

  49. Housley, M.P. et al. O-GlcNAc regulates FoxO activation in response to glucose. J. Biol. Chem. 283, 16283–16292 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Huang, H. et al. Skp2 inhibits FOXO1 in tumor suppression through ubiquitin-mediated degradation. Proc. Natl. Acad. Sci. USA 102, 1649–1654 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. van der Vos, K.E. & Coffer, P.J. FOXO-binding partners: it takes two to tango. Oncogene 27, 2289–2299 (2008).

    Article  CAS  PubMed  Google Scholar 

  52. Okamoto, H., Obici, S., Accili, D. & Rossetti, L. Restoration of liver insulin signaling in Insr knockout mice fails to normalize hepatic insulin action. J. Clin. Invest. 115, 1314–1322 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Gelling, R.W. et al. Insulin action in the brain contributes to glucose lowering during insulin treatment of diabetes. Cell Metab. 3, 67–73 (2006).

    Article  CAS  PubMed  Google Scholar 

  54. Obici, S., Zhang, B.B., Karkanias, G. & Rossetti, L. Hypothalamic insulin signaling is required for inhibition of glucose production. Nat. Med. 8, 1376–1382 (2002).

    Article  CAS  PubMed  Google Scholar 

  55. Hill, J.W. et al. Direct insulin and leptin action on pro-opiomelanocortin neurons is required for normal glucose homeostasis and fertility. Cell Metab. 11, 286–297 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Inoue, H. et al. Role of hepatic STAT3 in brain-insulin action on hepatic glucose production. Cell Metab. 3, 267–275 (2006).

    Article  CAS  PubMed  Google Scholar 

  57. Zhou, Y. et al. Regulation of glucose homeostasis through a XBP-1-FoxO1 interaction. Nat. Med. 17, 356–365 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Wan, M. et al. Loss of Akt1 in mice increases energy expenditure and protects against diet-induced obesity. Mol. Cell. Biol. 32, 96–106 (2011).

    Article  PubMed  Google Scholar 

  59. Summers, S.A., Whiteman, E.L., Cho, H., Lipfert, L. & Birnbaum, M.J. Differentiation-dependent suppression of platelet-derived growth factor signaling in cultured adipocytes. J. Biol. Chem. 274, 23858–23867 (1999).

    Article  CAS  PubMed  Google Scholar 

  60. Miller, R.A. et al. Adiponectin suppresses gluconeogenic gene expression in mouse hepatocytes independent of LKB1-AMPK signaling. J. Clin. Invest. 121, 2518–2528 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

The authors would like to thank M. Magnuson (Vanderbilt University) for providing the antibody to Gck and D. Accili (Columbia University) for sharing the Foxo1loxP/loxP mice. We are grateful to J. Schug, who helped with the microarray data analysis and generated the heat map and the density plot. The Functional Genomics Core and the Transgenic, Knockout, Mouse Phenotyping and Biomarker Cores of the University of Pennsylvania Diabetes and Endocrinology Research Center (NIH grant P30 DK19525) were instrumental in this work. This work was supported by the US National Institutes of Health grant RO1 DK56886 (M.J.B.) and the diabetes training grant T32 DK007314 (M.L.).

Author information

Authors and Affiliations

Authors

Contributions

M.L. conceived of the hypothesis, designed and performed the experiments and analyzed the data. M.W. and K.F.L. performed experiments. Q.C., B.R.M. and S.F. provided technical assistance. The R.S.A. lab performed the hyperinsulinemic-euglycemic clamps experiments. K.U. and C.R.K. generated the Akt1loxP/loxP mice. M.J.B. conceived the hypothesis and directed the project. M.L. and M.J.B. prepared the manuscript.

Corresponding author

Correspondence to Morris J Birnbaum.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–11 (PDF 3499 kb)

Supplementary Table 1

Metabolic responsive genes in the control livers (XLS 423 kb)

Supplementary Table 2

Metabolic responsive genes that respond to the loss of Akt in the fed livers (XLS 220 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Lu, M., Wan, M., Leavens, K. et al. Insulin regulates liver metabolism in vivo in the absence of hepatic Akt and Foxo1. Nat Med 18, 388–395 (2012). https://doi.org/10.1038/nm.2686

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nm.2686

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing