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FoxO1 induces apoptosis in skeletal myotubes in a DNA-binding-dependent manner.

ABSTRACT: Recent studies indicate that FoxO transcription factors play an important role in promoting muscle atrophy. To study mechanisms mediating effects of FoxO proteins on muscle wasting, FoxO1-estrogen receptor fusion proteins that are activated by treatment with 4-hydroxytamoxifen (4-OH-T) were stably transfected in C(2)C(12) skeletal myoblasts using the pBABE retroviral system and grown into multinucleated skeletal myotubes. Activation of FoxO1 resulted in significant muscle atrophy, which was accompanied by DNA fragmentation, evidenced by terminal deoxynucleotidyl transferase dUTP-mediated nick end labeling. Cells expressing a DNA-binding-deficient form of FoxO1 also exhibited significant atrophy on FoxO1 activation but no hallmark signs of apoptosis. FoxO1 activation resulted in a significant increase in muscle atrophy F-box (MAFbx)/atrogin-1, muscle-specific RING finger protein 1 (MuRF-1), and Bcl-2-interacting mediator of cell death (Bim) gene expression, with no significant increase in Bcl-2/adenovirus E1B 19-kDa-interacting protein 3 (BNip3) gene expression. Although the ability of FoxO1 to induce MuRF-1 gene expression appeared to be independent of DNA binding, expression of MAFbx/atrogin-1 and Bim was significantly blunted in cells expressing DNA-binding-deficient FoxO1. BNip3 gene expression was significantly elevated in DNA-binding-deficient mutant cells. These findings indicate that FoxO1 promotes skeletal muscle atrophy through induction of proteolytic and apoptotic machinery via DNA-binding-dependent and -independent mechanisms.

SUBMITTER: McLoughlin TJ

PROVIDER: S-EPMC2740395 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

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FoxO1 induces apoptosis in skeletal myotubes in a DNA-binding-dependent manner.

McLoughlin Thomas J TJ Smith Sierra M SM DeLong Alissa D AD Wang Hengbing H Unterman Terry G TG Esser Karyn A KA

American journal of physiology. Cell physiology 20090624 3

Recent studies indicate that FoxO transcription factors play an important role in promoting muscle atrophy. To study mechanisms mediating effects of FoxO proteins on muscle wasting, FoxO1-estrogen receptor fusion proteins that are activated by treatment with 4-hydroxytamoxifen (4-OH-T) were stably transfected in C(2)C(12) skeletal myoblasts using the pBABE retroviral system and grown into multinucleated skeletal myotubes. Activation of FoxO1 resulted in significant muscle atrophy, which was acco ...[more]

PMID: 19553561

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Project description:BackgroundSkeletal muscle wasting is an important comorbidity of Chronic Obstructive Pulmonary Disease (COPD) and is strongly correlated with morbidity and mortality. Patients who experience frequent acute exacerbations of COPD (AECOPD) have more severe muscle wasting and reduced recovery of muscle mass and function after each exacerbation. Serum levels of the pro-inflammatory acute phase protein Serum Amyloid A (SAA) can rise more than 1000-fold in AECOPD and are predictively correlated with exacerbation severity. The direct effects of SAA on skeletal muscle are poorly understood. Here we have examined SAA effects on pro-inflammatory cachectic cytokine expression (IL-6 and TNF?) and atrophy in C2C12 myotubes.ResultsSAA increased IL-6 (31-fold) and TNF? (6.5-fold) mRNA levels compared to control untreated cells after 3h of SAA treatment, and increased secreted IL-6 protein at 24h. OxPAPC, a dual TLR2 and TLR4 inhibitor, reduced the response to SAA by approximately 84% compared to SAA alone, and the TLR2 neutralising antibody T2.5 abolished SAA-induced expression of IL-6, indicating that SAA signalling in C2C12 myotubes is primarily via TLR2. SAA also reduced myotube width by 10-13% and induced a 2.5-fold increase in the expression of the muscle atrophy gene Atrogin-1, suggesting direct effects of SAA on muscle wasting. Blocking of TLR2 inhibited the SAA-induced decrease in myotube width and Atrogin-1 gene expression, indicating that SAA induces atrophy through TLR2.ConclusionsThese data demonstrate that SAA stimulates a robust pro-inflammatory response in skeletal muscle myotubes via the TLR2-dependent release of IL-6 and TNF?. Furthermore, the observed atrophy effects indicate that SAA could also be directly contributing to the wasting and poor recovery of muscle mass. Therapeutic strategies targeting this SAA-TLR2 axis may therefore ameliorate muscle wasting in AECOPD and a range of other inflammatory conditions associated with loss of muscle mass.

Dataset Information

FoxO1 induces apoptosis in skeletal myotubes in a DNA-binding-dependent manner.

Publications

FoxO1 induces apoptosis in skeletal myotubes in a DNA-binding-dependent manner.

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