Atrogin-1 inhibits Akt-dependent cardiac hypertrophy in mice via ubiquitin-dependent coactivation of Forkhead proteins
Hui-Hua Li ... David J. Glass, Cam Patterson
Hui-Hua Li ... David J. Glass, Cam Patterson
Published November 1, 2007
Citation Information: J Clin Invest. 2007;117(11):3211-3223. doi:10.1172/JCI31757.
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Categories: Research Article Cardiology

Atrogin-1 inhibits Akt-dependent cardiac hypertrophy in mice via ubiquitin-dependent coactivation of Forkhead proteins

Abstract

Cardiac hypertrophy is a major cause of human morbidity and mortality. Although much is known about the pathways that promote hypertrophic responses, mechanisms that antagonize these pathways have not been as clearly defined. Atrogin-1, also known as muscle atrophy F-box, is an F-box protein that inhibits pathologic cardiac hypertrophy by participating in a ubiquitin ligase complex that triggers degradation of calcineurin, a factor involved in promotion of pathologic hypertrophy. Here we demonstrated that atrogin-1 also disrupted Akt-dependent pathways responsible for physiologic cardiac hypertrophy. Our results indicate that atrogin-1 does not affect the activity of Akt itself, but serves as a coactivator for members of the Forkhead family of transcription factors that function downstream of Akt. This coactivator function of atrogin-1 was dependent on its ubiquitin ligase activity and the deposition of polyubiquitin chains on lysine 63 of Foxo1 and Foxo3a. Transgenic mice expressing atrogin-1 in the heart displayed increased Foxo1 ubiquitylation and upregulation of known Forkhead target genes concomitant with suppression of cardiac hypertrophy, while mice lacking atrogin-1 displayed the opposite physiologic phenotype. These experiments define a role for lysine 63–linked ubiquitin chains in transcriptional coactivation and demonstrate that atrogin-1 uses this mechanism to disrupt physiologic cardiac hypertrophic signaling through its effects on Forkhead transcription factors.

Authors

Hui-Hua Li, Monte S. Willis, Pamela Lockyer, Nathaniel Miller, Holly McDonough, David J. Glass, Cam Patterson

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Overexpression of atrogin-1 represses insulin- and IGF-1–induced cardiom...

Figure 1

Overexpression of atrogin-1 represses insulin- and IGF-1–induced cardiomyocyte hypertrophy and Foxo1 and Foxo3a dephosphorylation.

(A) Cardiomyocytes were infected with Ad-GFP or Ad–atrogin-1–GFP (MOI 10), and cells were stimulated with insulin (20 μg/ml–1), IGF-1 (20 ng/ml–1), or PE (100 μM) for 24 h and stained with α-actinin antibody (red). Quantitation of cell surface area was performed on 100 cells per group. *P < 0.001 vs. serum-free control and Ad-GFP. (B) Protein levels of endogenous Akt pathway members were determined in cardiomyocytes expressing atrogin-1 ectopically by IB with the indicated antibodies. (C) Foxo1 and Foxo3a expression and phosphorylation in cytoplasmic and nuclear fractions from cardiomyocytes were assessed by IB using anti-total or –phospho-Foxo1 and -Foxo3a antibodies after the indicated treatments. *P < 0.001 vs. Ad-GFP + insulin or IGF-1; #P < 0.01 vs. Ad-GFP + IGF-1. (D) Cardiomyocytes were infected with increasing MOIs of Ad–atrogin-1–GFP and Ad-GFP with IGF-1 (20 μg/ml–1) stimulation for 24 h. Endogenous Foxo1 and Foxo3a levels were determined by IB with the indicated antibodies. Maximal suppression of Foxo1 and Foxo3a occurred at MOI 30. *P < 0.001 vs. Ad-GFP + IGF-1 at MOI 30. (E) Cardiomyocytes were infected with Ad-siRNA–control or Ad-siRNA–atrogin-1 and treated with insulin (20 μg/ml–1) or PE (100 μM) for 24 h. (F) Cardiomyocytes treated as indicated were stained with α-actinin antibody. Quantitation of cell surface area was performed on 100 cells per group. Original magnification, ×200. *P < 0.01 vs. Ad-siRNA control + insulin or PE. Scale bars: 50 μm.