Histone deacetylases 1 and 2 regulate autophagy flux and skeletal muscle homeostasis in mice - PubMed (original) (raw)

Histone deacetylases 1 and 2 regulate autophagy flux and skeletal muscle homeostasis in mice

Viviana Moresi et al. Proc Natl Acad Sci U S A. 2012.

Abstract

Maintenance of skeletal muscle structure and function requires efficient and precise metabolic control. Autophagy plays a key role in metabolic homeostasis of diverse tissues by recycling cellular constituents, particularly under conditions of caloric restriction, thereby normalizing cellular metabolism. Here we show that histone deacetylases (HDACs) 1 and 2 control skeletal muscle homeostasis and autophagy flux in mice. Skeletal muscle-specific deletion of both HDAC1 and HDAC2 results in perinatal lethality of a subset of mice, accompanied by mitochondrial abnormalities and sarcomere degeneration. Mutant mice that survive the first day of life develop a progressive myopathy characterized by muscle degeneration and regeneration, and abnormal metabolism resulting from a blockade to autophagy. HDAC1 and HDAC2 regulate skeletal muscle autophagy by mediating the induction of autophagic gene expression and the formation of autophagosomes, such that myofibers of mice lacking these HDACs accumulate toxic autophagic intermediates. Strikingly, feeding HDAC1/2 mutant mice a high-fat diet from the weaning age releases the block in autophagy and prevents myopathy in adult mice. These findings reveal an unprecedented and essential role for HDAC1 and HDAC2 in maintenance of skeletal muscle structure and function and show that, at least in some pathological conditions, myopathy may be mitigated by dietary modifications.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

dKO mice die perinatally or develop a progressive myopathy. (A) H&E sections of various muscles of dKO mice and control mice, a few hours after birth. (Scale bars, 40 μm.) (B) Picture of dKO and control mice a few hours after birth. (C) Electron microscopic pictures of diaphragm isolated from 1-d-old control and dKO mice, showing ultrastructural abnormalities, including swollen and cup-shaped mitochondria (asterisk). (Scale bars, 0.5 μm.) (D) H&E sections of TA muscle from control and dKO mice at indicated ages. (Scale bars, 40 μm.)

Fig. 2.

HDAC1 and HDAC2 regulate autophagy flux in skeletal muscle. (A) Western blot showing autophagy markers (LC3 I-II and p62), phosphorylated AMPK (P-AMPK), and total AMPK in skeletal muscle of neonatal control and dKO mice. Tubulin was used as loading control. (B) Densitometric analysis of Western blot showing p62 and P-AMPK expression in dKO relative to control (Ctrl) mice. Values were normalized to tubulin. n = 4. *P < 0.05; **P < 0.005. (C) Western blot showing LC3 I-II and p62 in skeletal muscle of 4-wk-old control and dKO mice, fed or fasted for 24 h. Tubulin was used as loading control. (D) Densitometric analysis of Western blot showing LC3 II and p62 expression in dKO relative to Ctrl mice. Values were normalized to tubulin. n = 6. **P < 0.005. (E) Immunostaining of histological sections of TA muscle for p62 (green) and nuclei (blue) at 7 wk of age. (Scale bars, 40 μm.)

Fig. 3.

HDAC1 and HDAC2 regulate autophagosome formation in skeletal muscle. (A) Representative fields of GFP-LC3-electroporated TA muscle of adult control and dKO mice. Eight days after the electroporation, mice were fasted for 24 h. (Scale bars, 40 μm.) (B) Counts of GFP-LC3 punctae per myofiber in control (Ctrl) or dKO mice, fed or fasted for 24 h. n = 4–7 (≈400 fibers each). Values are mean ± SEM. **P < 0.005. (C) Real-time RT-PCR analysis, at 4 wk of age, for atg5, atg7, gaparapl1, LC3, and p62 mRNA of control (Ctrl) and dKO mice, fed or fasted for 48 h. Data are expressed as mean ± SEM. n = 4. *P < 0.05. **P < 0.005. (D) Representative fields of TA muscles 1 wk after coelectroporation of GFP-LC3 and either control or a combination of HDAC1 and HDAC2 expression plasmids. (Scale bars, 40 μm.) (E) Counts of GFP-LC3 punctae per myofiber in muscles coelectroporated with GFP-LC3 and either control or a combination of HDAC1 and HDAC2 expression plasmids. n = 6–8 (≈400 fibers each). Values are mean ± SEM. **P < 0.005. (F) Western blot analysis for autophagy markers LC3 I-II and p62, HDAC1, and HDAC2 in TA muscles 8 d after electroporation of either a control plasmid or a combination of HDAC1 and HDAC2 expressing plasmids.

Fig. 4.

Altered skeletal muscle energy expenditure in dKO mice. (A) Histological sections of TA isolated from 7-wk-old control and dKO mice were analyzed by metachromatic ATPase, SDH, and NADH staining. (Scale bars, 100 μm for metachromatic ATPase, 40 μm for SDH and NADH staining.) (B) Real-time RT-PCR analysis of different Myh genes, Ppargc1a, Ppargc1b, Acadm, and Ucp2 in control (Ctrl) and dKO mice, at 7 wk of age. Data are expressed as mean ± SEM. n = 4. *P < 0.05. dKO mice show significantly higher (C) O2 consumption, (D) CO2 production, and (E) heat production than control mice, between 5 and 7 wk of age. *P < 0.05. Values are mean ± SEM. n = 7.

Fig. 5.

HFD prevents the block of autophagy flux, myopathy, and perinatal mortality in dKO mice. (A) H&E sections of TA muscle of 3-mo-old control and dKO mice, fed with either normal chow or HFD for 8 wk. (Scale bars, 40 μm.) (B) Western blot analysis for autophagy markers LC3 I-II and p62 in control and dKO mice, fed with normal chow or HFD for 8 wk. Tubulin was used as loading control.

Fig. 6.

Model for the regulation of skeletal muscle homeostasis by HDAC1 and HDAC2. HDAC1 and HDAC2 promote autophagosome induction and formation. In the absence of HDAC1 and HDAC2 in skeletal muscle, autophagy flux is impaired, causing accumulation of protein aggregates and damaged mitochondria and eventually muscle degeneration.

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Histone deacetylases 1 and 2 regulate autophagy flux and skeletal muscle homeostasis in mice - PubMed (original) (raw)