Integrated expression analysis of muscle hypertrophy identifies Asb2 as a negative regulator of muscle mass
Jonathan R. Davey, Kevin I. Watt, Benjamin L. Parker, Rima Chaudhuri, James G. Ryall, Louise Cunningham, Hongwei Qian, Vittorio Sartorelli, Marco Sandri, Jeffrey Chamberlain, David E. James, Paul Gregorevic
Jonathan R. Davey, Kevin I. Watt, Benjamin L. Parker, Rima Chaudhuri, James G. Ryall, Louise Cunningham, Hongwei Qian, Vittorio Sartorelli, Marco Sandri, Jeffrey Chamberlain, David E. James, Paul Gregorevic
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Research Article Aging Muscle biology

Integrated expression analysis of muscle hypertrophy identifies Asb2 as a negative regulator of muscle mass

Abstract

The transforming growth factor-β (TGF-β) signaling network is a critical regulator of skeletal muscle mass and function and, thus, is an attractive therapeutic target for combating muscle disease, but the underlying mechanisms of action remain undetermined. We report that follistatin-based interventions (which modulate TGF-β network activity) can promote muscle hypertrophy that ameliorates aging-associated muscle wasting. However, the muscles of old sarcopenic mice demonstrate reduced response to follistatin compared with healthy young-adult musculature. Quantitative proteomic and transcriptomic analyses of young-adult muscles identified a transcription/translation signature elicited by follistatin exposure, which included repression of ankyrin repeat and SOCS box protein 2 (Asb2). Increasing expression of ASB2 reduced muscle mass, thereby demonstrating that Asb2 is a TGF-β network–responsive negative regulator of muscle mass. In contrast to young-adult muscles, sarcopenic muscles do not exhibit reduced ASB2 abundance with follistatin exposure. Moreover, preventing repression of ASB2 in young-adult muscles diminished follistatin-induced muscle hypertrophy. These findings provide insight into the program of transcription and translation events governing follistatin-mediated adaptation of skeletal muscle attributes and identify Asb2 as a regulator of muscle mass implicated in the potential mechanistic dysfunction between follistatin-mediated muscle growth in young and old muscles.

Authors

Jonathan R. Davey, Kevin I. Watt, Benjamin L. Parker, Rima Chaudhuri, James G. Ryall, Louise Cunningham, Hongwei Qian, Vittorio Sartorelli, Marco Sandri, Jeffrey Chamberlain, David E. James, Paul Gregorevic

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Figure 4

Asb2 is an atrogene that is downregulated during follistatin treatment.

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Asb2 is an atrogene that is downregulated during follistatin treatment....
(A) C57Bl/6 mice were injected with rAAV:MCS in the left TA and rAAV:indFST in the right TA. Immunoblot analysis of ASB2 expression in response to indicated follistatin treatments. (B) Quantification of ASB2 expression presented in A (ratio-paired t -test; P < 0.05; no DOX, n = 6; 2 days DOX, n = 7; 28 days DOX, n = 8; ± SEM). (C) Quantitative PCR analysis of Asb2 mRNA in response to indicated treatments (ratio-paired t-test; P < 0.05; no DOX, n = 6; 2 days DOX, n = 5; 28 days DOX, n = 4; ± SEM). (D) The mass of TA muscles of C57Bl/6 mice after injection of rAAV:MCS in the left TA and rAAV:SMAD7 in the right TA. Mass values were compared between contralateral TA muscles from individual mice (ratio-paired t-test, P < 0.05, n = 5, ± SEM). (E) Immunoblot analysis of mice from D for ASB2 expression and SMAD signaling. Quantification of immunoblots from E for (F) ASB2 expression (ratio-paired t test, P < 0.05, n = 5, ± SEM) and (G) ratio of phosphorylated to total SMAD3 and ratio of phosphorylated to total SMAD2 (ratio-paired t test, P < 0.05, n = 5, ± SEM). (H) C57Bl/6 mice were injected with rAAV:MCS in the left TA and rAAV:Asb2 in the right TA. The mass of TA muscles from rAAV:Asb2-injected muscles were compared with contralateral control muscles (ratio-paired t test; P < 0.05; 14 days, n = 5; 28 days, n = 4; ± SEM). (I) Change in muscle mass between rAAV:Asb2- and control-treated TA muscles from H at indicated time points (unpaired t test; P < 0.05; 14 days, n = 5; 28 days, n = 4; ± SEM). Recombinant adeno-associated virus, rAAV; ankyrin repeat-containing protein with a suppressor of cytokine signaling box 2, Asb2; control construct, MCS; follistatin, FST; tibialis anterior muscle, TA; doxycycline, DOX; tetracycline responsive follistatin, indFST.