Gα13 ablation reprograms myofibers to oxidative phenotype and enhances whole-body metabolism
Ja Hyun Koo, Tae Hyun Kim, Shi-Young Park, Min Sung Joo, Chang Yeob Han, Cheol Soo Choi, Sang Geon Kim
Ja Hyun Koo, Tae Hyun Kim, Shi-Young Park, Min Sung Joo, Chang Yeob Han, Cheol Soo Choi, Sang Geon Kim
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Research Article Metabolism Muscle biology

Gα13 ablation reprograms myofibers to oxidative phenotype and enhances whole-body metabolism

Abstract

Skeletal muscle is a key organ in energy homeostasis owing to its high requirement for nutrients. Heterotrimeric G proteins converge signals from cell-surface receptors to potentiate or blunt responses against environmental changes. Here, we show that muscle-specific ablation of Gα13 in mice promotes reprogramming of myofibers to the oxidative type, with resultant increases in mitochondrial biogenesis and cellular respiration. Mechanistically, Gα13 and its downstream effector RhoA suppressed nuclear factor of activated T cells 1 (NFATc1), a chief regulator of myofiber conversion, by increasing Rho-associated kinase 2–mediated (Rock2-mediated) phosphorylation at Ser243. Ser243 phosphorylation of NFATc1 was reduced after exercise, but was higher in obese animals. Consequently, Gα13 ablation in muscles enhanced whole-body energy metabolism and increased insulin sensitivity, thus affording protection from diet-induced obesity and hepatic steatosis. Our results define Gα13 as a switch regulator of myofiber reprogramming, implying that modulations of Gα13 and its downstream effectors in skeletal muscle are a potential therapeutic approach to treating metabolic diseases.

Authors

Ja Hyun Koo, Tae Hyun Kim, Shi-Young Park, Min Sung Joo, Chang Yeob Han, Cheol Soo Choi, Sang Geon Kim

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Figure 7

Rock2 phosphorylates NFATc1 at Ser243.

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Rock2 phosphorylates NFATc1 at Ser243. (A) In vitro kinase assay. Recomb...
(A) In vitro kinase assay. Recombinant proteins and [γ-32P]-ATP were incubated with or without Y-27632 (10 μM), and phosphorylation was visualized by autoradiography. (B) LC-MS/MS analysis of human NFATc1 after incubation with human Rock25-554. (C) Schematic illustration of NFATc1 protein domains and sequence comparisons of positive hits among different species. (D) NFATc1 transcription activity assay. NFATc1-KO C2C12 myotubes were transfected with an NFATc1 reporter plasmid in combination with a WT or serine-to-alanine mutant NFATc1 plasmid (n = 3 each). Data represent the mean ± SEM. *P < 0.05 and **P < 0.01, by Student’s t test. (E) In vitro kinase assay. An 8-mer peptide (hNFATc1236-243) surrounding the Ser241 residue was added to the reaction mixture where indicated. Kinase activity was determined by incorporation of the [γ-32P]-ATP label. (F and G) Immunoblots for phosphorylated NFATc1 (p-NFATc1) (Ser243). (F) NFATc1-KO C2C12 myotubes were transfected with a plasmid encoding WT NFATc1 or its S243A mutant. Adenoviral infection of Gα13QL was done 12 hours after the transfection, and samples were prepared 24 hours later. (G) C2C12 myotubes were transfected with siRNA against Rock2 or with control siRNA. After 12 hours, Gα13QL was transfected via adenovirus, and the samples were prepared 12 hours later.