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. 2008 Mar 25;105(12):4721-6.
doi: 10.1073/pnas.0800979105. Epub 2008 Mar 17.

Gene expression-based screening identifies microtubule inhibitors as inducers of PGC-1alpha and oxidative phosphorylation

Zoltan Arany  1 , Bridget K Wagner, Yanhong Ma, Jessica Chinsomboon, Dina Laznik, Bruce M Spiegelman
Affiliations

Gene expression-based screening identifies microtubule inhibitors as inducers of PGC-1alpha and oxidative phosphorylation

Zoltan Arany et al. Proc Natl Acad Sci U S A. .

Abstract

The transcriptional coactivator PGC-1alpha is a potent regulator of several metabolic pathways, including, in particular, the activation of oxidative phosphorylation and mitochondrial biogenesis. Recent evidence suggests that increasing PGC-1alpha activity may have beneficial effects in various conditions, including muscular dystrophy, diabetes, and neurodegenerative diseases. We describe here a high-throughput screen to identify small molecules that induce PGC-1alpha expression in skeletal muscle cells. A number of drug classes are identified, including glucocorticoids, microtubule inhibitors, and protein synthesis inhibitors. These drugs induce PGC-1alpha mRNA, and the expression of a number of genes known to be regulated by PGC-1alpha. No induction of these _target genes is seen in PGC-1alpha -/- cells, demonstrating that the drugs act through PGC-1alpha. These data demonstrate the feasibility of high-throughput screening for inducers of PGC-1alpha. Moreover, the data identify microtubule inhibitors and protein synthesis inhibitors as modulators of PGC-1alpha and oxidative phosphorylation.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Screen for small molecules that induce PGC-1α. (A) Schema of the screen. (B Upper) Myotubes were treated with the indicated doses of AICAR for 24 h, and levels of PGC-1α mRNA were measured. (B Lower) PGC-1α mRNA levels in myotubes 48 h after infection with adenovirus encoding for PGC-1α. (C) Representative graph of qPCR amplification curves detecting PGC-1α mRNA levels from one 384-well qPCR plate of the screen. (Inset) Shown are the complete curves. (D) Bin diagram of fold induction of PGC-1α in DMSO-control wells and in wells containing a synthetic glucocorticoid.
Fig. 2.
Fig. 2.
Protein synthesis inhibitors induce PGC-1α in primary skeletal muscle cells. (A) Myotubes were treated with the indicated doses of anisomycin, emetine, or cycloheximide, and levels of PGC-1α mRNA were evaluated after 16 h. (B) Myotubes were treated with 200 ng/ml or 1 μg/ml of anisomycin or emetine for the indicated times, and PGC-1α mRNA was measured. (C) PGC-1α mRNA levels in myotubes treated with anisomycin or emetine after pretreatment for 30 min with actinomycin D. (D) Myotubes were treated with 30 ng/ml of anisomycin for 1–3 days, as indicated, and the mRNA levels of cycs, esrra, and cox5b were measured by qPCR.
Fig. 3.
Fig. 3.
Tubulin inhibitors induce PGC-1α in primary skeletal muscle cells and synergize with glucocorticoids. (A) Myotubes were treated with the indicated doses of colchicine or taxol, and levels of PGC-1α mRNA were evaluated after 16 h. (B) PGC-1α mRNA levels after treatment for 16 h with the indicated doses of taxol or colchicine with and without 1 μM dexamethasone. (C) Myotubes were treated with 30 ng/ml colchicine + 1 μM dexamethasone for 24 or 48 h, and PGC-1α protein levels were measured by Western blotting. (D) PGC-1α mRNA levels in myotubes treated with colchicine + 1 μM dexamethasone after pretreatment for 30 min with the indicated inhibitors. AC, adenylase cyclase inhibitor SQ 22536, p38 MAPK inhibitors SB1, sb203580, and SB2, sb202190.
Fig. 4.
Fig. 4.
Tubulin inhibitors + glucocorticoids induce oxidative phosphorylation genes and activity in a PGC-1α-dependent fashion. (A) Myotubes were treated with the indicated drugs for 1–3 days, as indicated, and the mRNA levels of cycs, esrra, cox5b, and glut4 were measured by qPCR. (B) Primary skeletal muscle cells were prepared from PGC-1α +/+ and −/− animals, differentiated into myotubes, and treated with 30 ng/ml colchicine + 1 μM dexamethasone. After 48 or 72 h, PGC-1α mRNA levels were measured. (C) Total and uncoupled respiration in PGC-1α +/+ and −/− myotubes treated for 72 h with 30 ng/ml colchicine + 1 μM dexamethasone.
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