Autophagy-regulating TP53INP2 mediates muscle wasting and is repressed in diabetes
David Sala, … , Antonio L. Serrano, Antonio Zorzano
David Sala, … , Antonio L. Serrano, Antonio Zorzano
Published April 8, 2014
Citation Information: J Clin Invest. 2014;124(5):1914-1927. https://doi.org/10.1172/JCI72327.
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Research Article Endocrinology

Autophagy-regulating TP53INP2 mediates muscle wasting and is repressed in diabetes

Abstract

A precise balance between protein degradation and synthesis is essential to preserve skeletal muscle mass. Here, we found that TP53INP2, a homolog of the Drosophila melanogaster DOR protein that regulates autophagy in cellular models, has a direct impact on skeletal muscle mass in vivo. Using different transgenic mouse models, we demonstrated that muscle-specific overexpression of Tp53inp2 reduced muscle mass, while deletion of Tp53inp2 resulted in muscle hypertrophy. TP53INP2 activated basal autophagy in skeletal muscle and sustained p62-independent autophagic degradation of ubiquitinated proteins. Animals with muscle-specific overexpression of Tp53inp2 exhibited enhanced muscle wasting in streptozotocin-induced diabetes that was dependent on autophagy; however, TP53INP2 ablation mitigated experimental diabetes-associated muscle loss. The overexpression or absence of TP53INP2 did not affect muscle wasting in response to denervation, a condition in which autophagy is blocked, further indicating that TP53INP2 alters muscle mass by activating autophagy. Moreover, TP53INP2 expression was markedly repressed in muscle from patients with type 2 diabetes and in murine models of diabetes. Our results indicate that TP53INP2 negatively regulates skeletal muscle mass through activation of autophagy. Furthermore, we propose that TP53INP2 repression is part of an adaptive mechanism aimed at preserving muscle mass under conditions in which insulin action is deficient.

Authors

David Sala, Saška Ivanova, Natàlia Plana, Vicent Ribas, Jordi Duran, Daniel Bach, Saadet Turkseven, Martine Laville, Hubert Vidal, Monika Karczewska-Kupczewska, Irina Kowalska, Marek Straczkowski, Xavier Testar, Manuel Palacín, Marco Sandri, Antonio L. Serrano, Antonio Zorzano

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Figure 4

TP53INP2 promotes degradation of ubiquitinated proteins in skeletal muscle.

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TP53INP2 promotes degradation of ubiquitinated proteins in skeletal musc...
(A) p62 and NBR1 protein levels in total homogenates of gastrocnemius muscles from WT and SKM-Tg mice. Representative images are shown. (B) Western blot analysis of ubiquitinated protein content in total homogenates of gastrocnemius muscles from WT and SKM-Tg mice with or without chloroquine treatment. Representative images are shown. (C) Western blot analysis of p62, NBR1, TP53INP2, and ubiquitinated protein (FK2) content in protein extracts from day 5 C2C12 myotubes overexpressing TP53INP2 or LacZ (control). Cells were treated with bafilomycin (Baf.) as indicated. Thin black lines indicate that lanes were run on the same gel but were noncontiguous. Representative images are shown. (D) Immunofluorescence against p62 and LC3 in control (LacZ) or TP53INP2-overexpressing C2C12 day 5 myotubes. Cells were treated with bafilomycin at 200 nM during 3 hours as indicated. p62 is stained in red and LC3 is stained in green. Scale bar: 20 μm. (E) Immunofluorescence against ubiquitin (FK2) and TP53INP2 in C2C12 day 5 myotubes. Cells were treated with puromycin at 50 μg/ml during 4 hours as indicated. Ub (FK2) is stained in red and TP53INP2 is stained in green. Scale bar: 20 μm. (F) HEK293T cells were transfected with plasmids coding for GFP-Ub and/or mouse TP53INP2 or mouse TP53INP2 3KR mutant. Pull-down was performed using GFP-Trap beads and inputs and pellets were probed in Western blot assays with anti-GFP and anti-TP53INP2 antibodies.