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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.

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FoxO members are redundant and control a new set of ubiquitin ligases.(a–c) Frequency histograms showing the distribution of cross-sectional areas (μm2) of gastrocnemius muscles from muscle-specific (a) FoxO1−/−, (b) FoxO3−/− and (c) FoxO4−/− mice. Data are shown as mean±s.e.m. of four muscles each group. *P<0.05, **P<0.01. (d,e) qRT–PCR of the novel ubiquitin ligases MUSA1, Fbxo21/SMART, Fbxo31, Itch from 24 starved (d) or denervated (e) FoxO1,3,4f/f and FoxO1,3,4−/− mice. Data are normalized to GAPDH and expressed as fold increase of fed control mice. n=4 muscles for each group. (f) ChIP qPCR of FoxO3 and (g) FoxO1 on the promoters of MUSA1, Fbxo21/SMART and Itch. IgG was used as the reference. n=3. (h–j) qRT–PCR of MUSA1, Fbxo21/SMART, Fbxo31, Itch in (h) FoxO1, (i) FoxO3 and (j) FoxO4 knockout mice after 3 days of denervation. Values are normalized to GAPDH and expressed as fold increase of control mice. n=4 muscles for each group. Data are shown as mean±s.e.m. Error bars indicate s.e.m. *P<0.05, **P<0.01 (Student's t-test). con control; den, denervated. MUSA1 S1, FoxO-binding site1; MUSA1 S2, FoxO-binding site2; MUSA1 S3, FoxO-binding site3.
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f7: FoxO members are redundant and control a new set of ubiquitin ligases.(a–c) Frequency histograms showing the distribution of cross-sectional areas (μm2) of gastrocnemius muscles from muscle-specific (a) FoxO1−/−, (b) FoxO3−/− and (c) FoxO4−/− mice. Data are shown as mean±s.e.m. of four muscles each group. *P<0.05, **P<0.01. (d,e) qRT–PCR of the novel ubiquitin ligases MUSA1, Fbxo21/SMART, Fbxo31, Itch from 24 starved (d) or denervated (e) FoxO1,3,4f/f and FoxO1,3,4−/− mice. Data are normalized to GAPDH and expressed as fold increase of fed control mice. n=4 muscles for each group. (f) ChIP qPCR of FoxO3 and (g) FoxO1 on the promoters of MUSA1, Fbxo21/SMART and Itch. IgG was used as the reference. n=3. (h–j) qRT–PCR of MUSA1, Fbxo21/SMART, Fbxo31, Itch in (h) FoxO1, (i) FoxO3 and (j) FoxO4 knockout mice after 3 days of denervation. Values are normalized to GAPDH and expressed as fold increase of control mice. n=4 muscles for each group. Data are shown as mean±s.e.m. Error bars indicate s.e.m. *P<0.05, **P<0.01 (Student's t-test). con control; den, denervated. MUSA1 S1, FoxO-binding site1; MUSA1 S2, FoxO-binding site2; MUSA1 S3, FoxO-binding site3.

Mentions: Since all FoxO members are expressed in muscles and are under AKT regulation, we investigated whether they play synergistic roles or have specific functions. To address this point, we generated muscle-specific individual FoxO knockout animals. We used denervation as a model of muscle atrophy. Deletion of FoxO1 did not protect from muscle atrophy (Fig. 7a, Supplementary Fig. 11). However the presence of FoxO1 is required for the optimal induction of several atrophy-related genes including MuRF1, Cathepsin L, Gabarap L, GADD45a and TGIF (Supplementary Fig. 12).


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)

FoxO members are redundant and control a new set of ubiquitin ligases.(a–c) Frequency histograms showing the distribution of cross-sectional areas (μm2) of gastrocnemius muscles from muscle-specific (a) FoxO1−/−, (b) FoxO3−/− and (c) FoxO4−/− mice. Data are shown as mean±s.e.m. of four muscles each group. *P<0.05, **P<0.01. (d,e) qRT–PCR of the novel ubiquitin ligases MUSA1, Fbxo21/SMART, Fbxo31, Itch from 24 starved (d) or denervated (e) FoxO1,3,4f/f and FoxO1,3,4−/− mice. Data are normalized to GAPDH and expressed as fold increase of fed control mice. n=4 muscles for each group. (f) ChIP qPCR of FoxO3 and (g) FoxO1 on the promoters of MUSA1, Fbxo21/SMART and Itch. IgG was used as the reference. n=3. (h–j) qRT–PCR of MUSA1, Fbxo21/SMART, Fbxo31, Itch in (h) FoxO1, (i) FoxO3 and (j) FoxO4 knockout mice after 3 days of denervation. Values are normalized to GAPDH and expressed as fold increase of control mice. n=4 muscles for each group. Data are shown as mean±s.e.m. Error bars indicate s.e.m. *P<0.05, **P<0.01 (Student's t-test). con control; den, denervated. MUSA1 S1, FoxO-binding site1; MUSA1 S2, FoxO-binding site2; MUSA1 S3, FoxO-binding site3.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4403316&req=5

f7: FoxO members are redundant and control a new set of ubiquitin ligases.(a–c) Frequency histograms showing the distribution of cross-sectional areas (μm2) of gastrocnemius muscles from muscle-specific (a) FoxO1−/−, (b) FoxO3−/− and (c) FoxO4−/− mice. Data are shown as mean±s.e.m. of four muscles each group. *P<0.05, **P<0.01. (d,e) qRT–PCR of the novel ubiquitin ligases MUSA1, Fbxo21/SMART, Fbxo31, Itch from 24 starved (d) or denervated (e) FoxO1,3,4f/f and FoxO1,3,4−/− mice. Data are normalized to GAPDH and expressed as fold increase of fed control mice. n=4 muscles for each group. (f) ChIP qPCR of FoxO3 and (g) FoxO1 on the promoters of MUSA1, Fbxo21/SMART and Itch. IgG was used as the reference. n=3. (h–j) qRT–PCR of MUSA1, Fbxo21/SMART, Fbxo31, Itch in (h) FoxO1, (i) FoxO3 and (j) FoxO4 knockout mice after 3 days of denervation. Values are normalized to GAPDH and expressed as fold increase of control mice. n=4 muscles for each group. Data are shown as mean±s.e.m. Error bars indicate s.e.m. *P<0.05, **P<0.01 (Student's t-test). con control; den, denervated. MUSA1 S1, FoxO-binding site1; MUSA1 S2, FoxO-binding site2; MUSA1 S3, FoxO-binding site3.
Mentions: Since all FoxO members are expressed in muscles and are under AKT regulation, we investigated whether they play synergistic roles or have specific functions. To address this point, we generated muscle-specific individual FoxO knockout animals. We used denervation as a model of muscle atrophy. Deletion of FoxO1 did not protect from muscle atrophy (Fig. 7a, Supplementary Fig. 11). However the presence of FoxO1 is required for the optimal induction of several atrophy-related genes including MuRF1, Cathepsin L, Gabarap L, GADD45a and TGIF (Supplementary Fig. 12).

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