Limits...
Regulation of autophagy and the ubiquitin-proteasome system by the FoxO transcriptional network during muscle atrophy.

Milan G, Romanello V, Pescatore F, Armani A, Paik JH, Frasson L, Seydel A, Zhao J, Abraham R, Goldberg AL, Blaauw B, DePinho RA, Sandri M - Nat Commun (2015)

Bottom Line: Notably, in the setting of low nutrient signalling, we demonstrate that FoxOs are required for Akt activity but not for mTOR signalling.FoxOs control several stress-response pathways such as the unfolded protein response, ROS detoxification, DNA repair and translation.Finally, we identify FoxO-dependent ubiquitin ligases including MUSA1 and a previously uncharacterised ligase termed SMART (Specific of Muscle Atrophy and Regulated by Transcription).

View Article: PubMed Central - PubMed

Affiliation: Venetian Institute of Molecular Medicine, via Orus 2, 35129 Padova, Italy.

ABSTRACT
Stresses like low nutrients, systemic inflammation, cancer or infections provoke a catabolic state characterized by enhanced muscle proteolysis and amino acid release to sustain liver gluconeogenesis and tissue protein synthesis. These conditions activate the family of Forkhead Box (Fox) O transcription factors. Here we report that muscle-specific deletion of FoxO members protects from muscle loss as a result of the role of FoxOs in the induction of autophagy-lysosome and ubiquitin-proteasome systems. Notably, in the setting of low nutrient signalling, we demonstrate that FoxOs are required for Akt activity but not for mTOR signalling. FoxOs control several stress-response pathways such as the unfolded protein response, ROS detoxification, DNA repair and translation. Finally, we identify FoxO-dependent ubiquitin ligases including MUSA1 and a previously uncharacterised ligase termed SMART (Specific of Muscle Atrophy and Regulated by Transcription). Our findings underscore the central function of FoxOs in coordinating a variety of stress-response genes during catabolic conditions.

Show MeSH

Related in: MedlinePlus

FoxOs are required for expression of several atrogenes after denervation.Quantitative RT–PCR of the indicated atrogenes after 3 days from denervation. Data are normalized to GAPDH and expressed as fold increase of control innervated muscles. Values are mean±s.e.m. *P<0,05, **P<0.01 (Student's t-test).; cont, control; den, denervated.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4403316&req=5

f6: FoxOs are required for expression of several atrogenes after denervation.Quantitative RT–PCR of the indicated atrogenes after 3 days from denervation. Data are normalized to GAPDH and expressed as fold increase of control innervated muscles. Values are mean±s.e.m. *P<0,05, **P<0.01 (Student's t-test).; cont, control; den, denervated.

Mentions: To further determine whether the role of FoxO1,3,4 is critical in different catabolic conditions, we then used denervation as another model of muscle atrophy. Quantification of fibre size revealed that FoxO-deficient muscles were partially protected from atrophy (Fig. 5a, Supplementary Fig. 9). When we monitored muscle force, we confirmed the histological data. Soleus muscle of FoxO1,3,4 knockout mice generated higher strength than controls in the basal condition (Fig. 5b) and FoxO deletion was able to partially prevent weakness and maintain the same force of innervated FoxO1,3,4f/f (Fig. 5b,c). The decrease of force in denervated FoxO1,3,4-deficient soleus muscle is related to the presence of FoxO proteins in type I fibres that can promote protein breakdown in these myofibres. It is important to emphasize that the type I fibres express FoxOs because the promoter (MLC1f) driving the expression of Cre recombinase is active in type II fibres and not in type I fibres. However, comparison of the force/frequency curves of denervated wild-type versus denervated FoxO1,3,4 soleus muscles underlines the important protection conferred by the absence of FoxO family when nerve is damaged (Fig. 5c). Analyses of signalling confirmed the downregulation of P-AKT in knockout muscles. Denervation induced an increase of total and phospho-4EBP1 protein in FoxO1,3,4f/f but not in FoxO1,3,4−/− muscle (Fig. 5d). The changes in the phosphorylation of 4EBP1 were not ascribed to activation of the cellular energy sensor AMPK, revealed by checking its phosphorylation level or the downstream target ACC. In contrast to fasting, the autophagy system was mildly affected by the absence of FoxOs. Indeed, LC3 lipidation was slightly decreased in FoxO1,3,4−/− compared with controls after denervation (Fig. 5e). However, and consistent with fasting, the transcriptional upregulation of p62/SQSTM1 in denervated muscles was greatly attenuated (Figs 5e and 6). When we tested the expression of the different atrophy-related genes, we found only a partial suppression of these genes (Fig. 6). Interestingly, the lists of FoxO-dependent genes during denervation and fasting do not completely overlap (Supplementary Fig. 10). For instance, the ROS detoxifying factor, Nrf2/Nfe2l2, was not affected by FoxO deletion in fasting, while it was less activated in denervated FoxO1,3,4 muscles. Therefore, FoxO family members are necessary for muscle loss but their involvement in the atrophy programme depends on the catabolic condition.


Regulation of autophagy and the ubiquitin-proteasome system by the FoxO transcriptional network during muscle atrophy.

Milan G, Romanello V, Pescatore F, Armani A, Paik JH, Frasson L, Seydel A, Zhao J, Abraham R, Goldberg AL, Blaauw B, DePinho RA, Sandri M - Nat Commun (2015)

FoxOs are required for expression of several atrogenes after denervation.Quantitative RT–PCR of the indicated atrogenes after 3 days from denervation. Data are normalized to GAPDH and expressed as fold increase of control innervated muscles. Values are mean±s.e.m. *P<0,05, **P<0.01 (Student's t-test).; cont, control; den, denervated.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4403316&req=5

f6: FoxOs are required for expression of several atrogenes after denervation.Quantitative RT–PCR of the indicated atrogenes after 3 days from denervation. Data are normalized to GAPDH and expressed as fold increase of control innervated muscles. Values are mean±s.e.m. *P<0,05, **P<0.01 (Student's t-test).; cont, control; den, denervated.
Mentions: To further determine whether the role of FoxO1,3,4 is critical in different catabolic conditions, we then used denervation as another model of muscle atrophy. Quantification of fibre size revealed that FoxO-deficient muscles were partially protected from atrophy (Fig. 5a, Supplementary Fig. 9). When we monitored muscle force, we confirmed the histological data. Soleus muscle of FoxO1,3,4 knockout mice generated higher strength than controls in the basal condition (Fig. 5b) and FoxO deletion was able to partially prevent weakness and maintain the same force of innervated FoxO1,3,4f/f (Fig. 5b,c). The decrease of force in denervated FoxO1,3,4-deficient soleus muscle is related to the presence of FoxO proteins in type I fibres that can promote protein breakdown in these myofibres. It is important to emphasize that the type I fibres express FoxOs because the promoter (MLC1f) driving the expression of Cre recombinase is active in type II fibres and not in type I fibres. However, comparison of the force/frequency curves of denervated wild-type versus denervated FoxO1,3,4 soleus muscles underlines the important protection conferred by the absence of FoxO family when nerve is damaged (Fig. 5c). Analyses of signalling confirmed the downregulation of P-AKT in knockout muscles. Denervation induced an increase of total and phospho-4EBP1 protein in FoxO1,3,4f/f but not in FoxO1,3,4−/− muscle (Fig. 5d). The changes in the phosphorylation of 4EBP1 were not ascribed to activation of the cellular energy sensor AMPK, revealed by checking its phosphorylation level or the downstream target ACC. In contrast to fasting, the autophagy system was mildly affected by the absence of FoxOs. Indeed, LC3 lipidation was slightly decreased in FoxO1,3,4−/− compared with controls after denervation (Fig. 5e). However, and consistent with fasting, the transcriptional upregulation of p62/SQSTM1 in denervated muscles was greatly attenuated (Figs 5e and 6). When we tested the expression of the different atrophy-related genes, we found only a partial suppression of these genes (Fig. 6). Interestingly, the lists of FoxO-dependent genes during denervation and fasting do not completely overlap (Supplementary Fig. 10). For instance, the ROS detoxifying factor, Nrf2/Nfe2l2, was not affected by FoxO deletion in fasting, while it was less activated in denervated FoxO1,3,4 muscles. Therefore, FoxO family members are necessary for muscle loss but their involvement in the atrophy programme depends on the catabolic condition.

Bottom Line: Notably, in the setting of low nutrient signalling, we demonstrate that FoxOs are required for Akt activity but not for mTOR signalling.FoxOs control several stress-response pathways such as the unfolded protein response, ROS detoxification, DNA repair and translation.Finally, we identify FoxO-dependent ubiquitin ligases including MUSA1 and a previously uncharacterised ligase termed SMART (Specific of Muscle Atrophy and Regulated by Transcription).

View Article: PubMed Central - PubMed

Affiliation: Venetian Institute of Molecular Medicine, via Orus 2, 35129 Padova, Italy.

ABSTRACT
Stresses like low nutrients, systemic inflammation, cancer or infections provoke a catabolic state characterized by enhanced muscle proteolysis and amino acid release to sustain liver gluconeogenesis and tissue protein synthesis. These conditions activate the family of Forkhead Box (Fox) O transcription factors. Here we report that muscle-specific deletion of FoxO members protects from muscle loss as a result of the role of FoxOs in the induction of autophagy-lysosome and ubiquitin-proteasome systems. Notably, in the setting of low nutrient signalling, we demonstrate that FoxOs are required for Akt activity but not for mTOR signalling. FoxOs control several stress-response pathways such as the unfolded protein response, ROS detoxification, DNA repair and translation. Finally, we identify FoxO-dependent ubiquitin ligases including MUSA1 and a previously uncharacterised ligase termed SMART (Specific of Muscle Atrophy and Regulated by Transcription). Our findings underscore the central function of FoxOs in coordinating a variety of stress-response genes during catabolic conditions.

Show MeSH
Related in: MedlinePlus