Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2006 Sep;116(9):2464-72.
doi: 10.1172/JCI27047. Epub 2006 Aug 10.

Dual role of transcription factor FoxO1 in controlling hepatic insulin sensitivity and lipid metabolism

Michihiro Matsumoto  1 , Seongah Han, Tadahiro Kitamura, Domenico Accili
Affiliations

Dual role of transcription factor FoxO1 in controlling hepatic insulin sensitivity and lipid metabolism

Michihiro Matsumoto et al. J Clin Invest. 2006 Sep.

Abstract

Hepatic insulin resistance affects both carbohydrate and lipid metabolism. It has been proposed that insulin controls these 2 metabolic branches through distinct signaling pathways. FoxO transcription factors are considered effectors of the pathway regulating hepatic glucose production. Here we show that adenoviral delivery of constitutively nuclear forkhead box O1 (FoxO1) to mouse liver results in steatosis arising from increased triglyceride accumulation and decreased fatty acid oxidation. FoxO1 gain of function paradoxically increased insulin sensitivity by promoting Akt phosphorylation, while FoxO1 inhibition via siRNA decreased it. We show that FoxO1 regulation of Akt phosphorylation does not require DNA binding and is associated with repression of the pseudokinase tribble 3 (Trb3), a modulator of Akt activity. This unexpected dual role of FoxO1 in promoting insulin sensitivity and lipid synthesis in addition to glucose production has the potential to explain the peculiar admixture of insulin resistance and sensitivity that is commonly observed in the metabolic syndrome.

PubMed Disclaimer

Figures

Figure 1
Figure 1. FoxO1 overexpression in liver.
Macroscopic view of livers (A) and oil red O staining of liver sections (B) from representative mice injected with control empty adenovirus or adenovirus expressing FoxO1ADA and collected after an overnight fast. (C) Liver TG content in fasted and ad libitum–fed FoxO1ADA or control mice (n = 6 for each group). (D) Gene expression analysis by real-time RT-PCR in livers from ad libitum-fed control (n = 6; white bars) or FoxO1ADA mice (n = 6; black bars). Each PCR was carried out in triplicate. *P < 0.05 and **P < 0.01. (E) Western blot analysis of liver extracts from adenovirus-injected mice. (F) Effect of FoxO1 inhibition by siRNA on insulin-induced Akt phosphorylation. We transduced cells with adenovirus encoding control siRNA or FoxO1siRNA and immunoblotted with the indicated antibodies. (G) Effect of FoxO1 DNA binding–deficient transduction on insulin-induced Akt phosphorylation. All data are representative of at least 3 independent experiments.
Figure 2
Figure 2. FoxO1ADA promotes Akt activation independent of increased Irs2 expression.
(A and B) Effects of FoxO1ADA in SV40-transformed hepatocytes transduced with FoxO1ADA at different MOIs. We incubated cells with insulin for 10 minutes and subjected cell lysates to immunoblot analysis with the indicated antibodies. All data are representative of at least 3 independent experiments. Fao hepatoma cells transduced with FoxO1ADA (C) or Irs2 (D) adenovirus were preincubated for 24 hours in the absence or presence of insulin. For immunoprecipitation, cells were treated for 2 minutes with insulin, and lysates were immunoprecipitated with antibodies against Irs2 and immunoblotted with either anti-phosphotyrosine or anti-Irs2 antibodies. For direct immunoblot analysis, the immunoprecipitation step was omitted. All data are representative of at least 2 independent experiments. (E) C2C12 myotubes transduced with FoxO1ADA adenovirus were incubated with insulin for 10 minutes and analyzed by direct immunoblotting with the indicated antibodies or immunoprecipitated and immunoblotted with antibodies to either Irs1 or Irs2.
Figure 3
Figure 3. FoxO1ADA does not affect expression of Akt pathway components in hepatocytes.
(A) SV40 hepatocytes from WT (Insr+/+) or insulin receptor–deficient (Insr–/–) mice transduced with FoxO1ADA adenovirus were incubated with insulin, lysed, and subjected to immunoblot analysis with the indicated antibodies. (B) Fao cells transduced with FoxO1ADA adenovirus were incubated in the absence or presence of insulin for 2 minutes and then lysed. Cell lysates were either immunoblotted with the indicated antibodies or immunoprecipitated with antibody against anti-phosphotyrosine (p-Y IP) and immunoblotted with antibody against the PI3K p85 subunit. (C) Hepatocytes transduced with FoxO1ADA adenovirus were preincubated with cycloheximide (Cx) for 16 hours. Thereafter, cells were treated with insulin, lysed, and analyzed by immunoblot with the indicated antibodies.
Figure 4
Figure 4. FoxO1 suppresses Trb3 expression in an insulin-dependent manner.
Northern blot analyses of Fao cells transduced with FoxO1ADA adenovirus and incubated with FSK-DEX and/or insulin for 20 hours (A); FoxO1ADA and Myr-p110 and treated with insulin in the absence or presence of LY294002 (LY) (B); FoxO1ADA and Myr-Akt and treated with or without insulin (C). 36b4 is a housekeeping gene used as a control for gel loading. (D) Hepatocytes transduced with FoxO1ADA (upper panel) or Irs2 (lower panel) adenoviruses were preincubated with insulin. Thereafter, cells were treated for 10 minutes with fresh insulin-containing medium, lysed, and immunoblotted with the indicated antibodies. All data are representative of at least 2 independent experiments. (E) Immunoblot analysis of the effect of cycloheximide on Trb3 expression. (F) Gene expression analysis by real-time RT-PCR in livers of control or FoxO1ADA mice (n = 6 for each). Each PCR was carried out in triplicate. *P < 0.05. (G) Western blot analysis of Trb3 expression in liver from control or FoxO1ADA mice. Northern blot and immunoblot data are representative of 3 independent experiments. (H) Northern blot analyses of Fao cells transduced with FoxO1ADA and Pgc1α.
Figure 5
Figure 5. FoxO1 regulation ofTrb3 promoter activity.
(A) SV40 hepatocytes were transiently cotransfected with reporter plasmids and incubated with FSK-DEX and/or insulin or LY294002. (B) Eight hours after cotransfection, cells were incubated with insulin for 16 hours. (C) Cells were transduced with either empty, Myr-p110, or Myr-Akt adenovirus. After 24 hours, cells were transiently cotransfected with the indicated plasmids and adenoviruses. (D) Deletion analysis of the Trb3 promoter in SV40 hepatocytes cultured in the presence of FSK-DEX (white bars), FoxO1ADA (black bars), or LY294002 (gray bars). *P < 0.05 vs. vector-transfected cells; **P< 0.01; and #P < 0.05 vs. β-gal–transduced cells. (E) ChIP in SV40 hepatocytes. We immunoprecipitated the following with anti-FoxO1 (upper panel) or control antiserum (middle panel): lane 1, untransduced cells; lane 2, cells transduced with FoxO1ADA; lanes 3–4, cells transduced with WT FoxO1. Bottom panel: total input DNA. (F) Mutation analysis. Control (WT) or mutant Trb3 promoter with mutations of the Foxo sites (Mutant) were assayed in basal conditions (white bars) and in the presence of LY294002 (black bars) or FoxO1ADA (gray bars). **P < 0.01. (G) Effect of FoxO1 knockdown on Trb3 promoter activity. SV40 hepatocytes were transiently cotransfected with either FoxO1siRNA or control siRNA with Trb3 promoter reporter construct at final concentration of 30 nM. **P < 0.01.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Accili D. Lilly lecture 2003: the struggle for mastery in insulin action: from triumvirate to republic. Diabetes. 2004;53:1633–1642. - PubMed
    1. Cherrington A.D. Banting lecture 1997. Control of glucose uptake and release by the liver in vivo. Diabetes. 1999;48:1198–1214. - PubMed
    1. Schwarz J.M., Linfoot P., Dare D., Aghajanian K. Hepatic de novo lipogenesis in normoinsulinemic and hyperinsulinemic subjects consuming high-fat, low-carbohydrate and low-fat, high-carbohydrate isoenergetic diets. Am. J. Clin. Nutr. 2003;77:43–50. - PubMed
    1. Iozzo P., et al. Defective liver disposal of free fatty acids in patients with impaired glucose tolerance. J. Clin. Endocrinol. Metab. 2004;89:3496–3502. - PubMed
    1. Ginsberg H.N., Zhang Y.L., Hernandez-Ono A. Regulation of plasma triglycerides in insulin resistance and diabetes. Arch. Med. Res. 2005;36:232–240. - PubMed

Publication types

MeSH terms