Myogenic Basic Helix-Loop-Helix Proteins Regulate the Expression of Peroxisomal Proliferator Activated Receptor-γ Coactivator-1α
Endocrinology
Myogenic Basic Helix-Loop-Helix Proteins Regulate the Expression of Peroxisomal Proliferator Activated Receptor- Coactivator-1
Ju Hui Chang 0
Kwang Huei Lin 0
Chung Hsuan Shih 0
Yu Jung Chang 0
Hsiang Chung Chi 0
Shen Liang Chen 0
0 Department of Life Sciences, National Central University (J.H.C. , C.H.S., Y.J.C., H.C.C., S.L.C.), Jhongli 32054 , Taiwan; and Department of Biochemistry, Chang Gung University (K.H.L.) , Kweisan 333 , Taiwan
Peroxisomal proliferator-activated receptor- coactivator-1 (PGC-1 ), a transcriptional coactivator, is selectively expressed in slow-twitch fibers in skeletal muscle. Ectopic expression of the PGC-1 gene in either a cell or an animal has been shown to promote fast to slow fiber-type switch. The expression of PGC-1 in muscle is regulated by myocyte enhancer factor 2 and Forkhead in rhabdomyosarcoma, two transcription factors implicated in terminal muscle differentiation. In this study we found that PGC-1 expression was activated during terminal muscle differentiation in both C2C12 and Sol8 myoblasts. Using retrovirus-mediated MyoD overexpression in C3H10T1/2 cells, we also demonstrated that MyoD, the master regulator of terminal differentiation, could activate PGC-1 expression in vivo. Our transient transfection results also show that myogenic basic helix-loop-helix
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P
EROXISOMAL PROLIFERATOR-activated
receptor(PPAR ) coactivator-1 (PGC1 ) is a transcriptional
coactivator identified in the screening for proteins
associating with PPAR . It is highly expressed in tissues requiring
high-energy metabolism, such as heart, skeletal muscle, and
brown fat (
1, 2
). Its expression in brown fat is highly induced
when animals are exposed to a cold environment, implying
its role in metabolic thermogenesis adaptation. Later studies
have proven its function in the biogenesis of mitochondria,
gluconeogenesis, and fatty acid oxidation (
3, 4
). Since its
identification, PGC-1 has been found to coactivate the
function of many nuclear hormone receptors, including thyroid
hormone, estrogen, and glucocorticoid receptors, and many
more (
1, 5– 8
). The wide variety of its partners suggests that
PGC-1 is not only implicated in the energy metabolism, but
is also involved in many other physiological processes.
First Published Online March 9, 2006
Abbreviations: bHLH, Basic helix-loop-helix; CMB, confluent; Ct,
maximum cycle threshold; DTT, dithiothreitol; FKHR, Forkhead in
rhabdomyosarcoma; GAPDH, glyceraldehyde-3-phosphate dehydrogenase,
GFP, green fluorescence protein; GST, glutathione-S-transferase; h,
human; MEF2, myocyte enhancer factor 2; MRF, myogenic regulatory
factor; MT, myotube; Mt, mutant; Pfu, DNA polymerase-derived from
Pyrococcus furiosus; PGC-1, peroxisome proliferation activation
receptor- coactivator 1; PGC1 , peroxisomal proliferator activated
receptor- coactivator 1 ; PMB, proliferating; PNK, polynucleotide kinase;
SDS, sodium dodecyl sulfate.
Endocrinology is published monthly by The Endocrine Society (http://
www.endo-society.org), the foremost professional society serving the
endocrine community.
(bHLH) proteins, especially MyoD, can activate PGC-1
expression by targeting its promoter. Myogenic bHLH protein
target sites on PGC-1 promoter were localized to a short
region ( 49 to 2) adjacent to the transcription start site,
which contains two putative E boxes. Mutation of either site
significantly reduced MyoD-mediated transactivation in the
cells, suggesting that both sites are required for myogenic
bHLH protein-mediated activation. However, only one site,
the E2 box, was directly bound by
glutathione-S-transferaseMyoD protein in EMSAs. Our results indicate that myogenic
bHLH proteins not only are involved in lineage determination
and terminal differentiation, but also are directly implicated
in activation of the key fiber-type and metabolic switch gene,
PGC-1 . (Endocrinology 147: 3093–3106, 2006)
In muscle, PGC-1 is preferentially expressed in
slowtwitch fibers, which are much higher in mitochondria content
and are more dependent on oxidative metabolism than
fasttwitch fibers (
9
). In transgenic mice, as the expression of
PGC-1 is driven by muscle creatine kinase promoter, the
putative fast-twitch fibers are converted into slow-twitch
fibers, which are characterized by the expression of troponin
I slow, myoglobin, and genes of mitochondria oxidative
metabolism (
9
). The activation of slow-twitch muscle-specific
genes by PGC-1 is mediated through MEF2 transcription
factors binding to the upstream enhancer sites. MEF2
proteins also activate the transcription of PGC-1 gene and thus
establish a positive feedback loop (
10
). Hormones both
activating, such as thyroid hormone, and inhibiting, such as
insulin, oxidation metabolism can also regulate the
expression of PGC-1 either positively or negatively (
11–13
). The
signals of insulin in the nucleus are mediated by Forkhead
in rhabdomyosarcoma (FKHR), a forkhead transcription
factor that b (...truncated)