PGC-1 alpha and PGC-1 beta increase protein synthesis via ERR alpha in C2C12 myotubes

The transcriptional coactivators peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) and PGC-1β are positive regulators of skeletal muscle mass and energy metabolism; however, whether they influence muscle growth and metabolic adaptations via increased protein synthesis is not clear. This study revealed PGC-1α or PGC-1β overexpression in C2C12 myotubes increased protein synthesis and myotube diameter under basal conditions and attenuated the loss in protein synthesis following the treatment with the catabolic agent, dexamethasone. To investigate whether PGC-1α or PGC-1β signal through the Akt/mTOR pathway to increase protein synthesis, treatment with the PI3K and mTOR inhibitors, LY294002 and rapamycin, respectively, was undertaken but found unable to block PGC-1α or PGC-1β’s promotion of protein synthesis. Furthermore, PGC-1α and PGC-1β decreased phosphorylation of Akt and the Akt/mTOR substrate, p70S6K. In contrast to Akt/mTOR inhibition, the suppression of ERRα, a major effector of PGC-1α and PGC-1β activity, attenuated the increase in protein synthesis and myotube diameter in the presence of PGC-1α or PGC-1β overexpression. To characterize further the biological processes occurring, gene set enrichment analysis of genes commonly regulated by both PGC-1α and PGC-1β was performed following a microarray screen. Genes were found enriched in metabolic and mitochondrial oxidative processes, in addition to protein translation and muscle development categories. This suggests concurrent responses involving both increased metabolism and myotube protein synthesis. Finally, based on their known function or unbiased identification through statistical selection, two sets of genes were investigated in a human exercise model of stimulated protein synthesis to characterize further the genes influenced by PGC-1α and PGC-1β during physiological adaptive changes in skeletal muscle

Nuclear receptor/microRNA circuitry links muscle fiber type to energy metabolism

The mechanisms involved in the coordinate regulation of the metabolic and structural programs controlling muscle fitness and endurance are unknown. Recently, the nuclear receptor PPAR beta/delta was shown to activate muscle endurance programs in transgenic mice. In contrast, muscle-specific transgenic overexpression of the related nuclear receptor, PPAR alpha, results in reduced capacity for endurance exercise. We took advantage of the divergent actions of PPAR beta/delta and PPAR alpha to explore the downstream regulatory circuitry that orchestrates the programs linking muscle fiber type with energy metabolism. Our results indicate that, in addition to the well-established role in transcriptional control of muscle metabolic genes, PPAR beta/delta and PPAR alpha participate in programs that exert opposing actions upon the type I fiber program through a distinct muscle microRNA (miRNA) network, dependent on the actions of another nuclear receptor, estrogen-related receptor gamma (ERR gamma). Gain-of-function and loss-of-function strategies in mice, together with assessment of muscle biopsies from humans, demonstrated that type I muscle fiber proportion is increased via the stimulatory actions of ERR gamma on the expression of miR-499 and miR-208b. This nuclear receptor/miRNA regulatory circuit shows promise for the identification of therapeutic targets aimed at maintaining muscle fitness in a variety of chronic disease states, such as obesity, skeletal myopathies, and heart failure

ADRA1A-Gα_q signalling potentiates adipocyte thermogenesis through CKB and TNAP

Noradrenaline (NA) regulates cold-stimulated adipocyte thermogenesis(1). Aside from cAMP signalling downstream of β-adrenergic receptor activation, how NA promotes thermogenic output is still not fully understood. Here, we show that coordinated α(1)-adrenergic receptor (AR) and β(3)-AR signalling induces the expression of thermogenic genes of the futile creatine cycle(2,3), and that early B cell factors, oestrogen-related receptors and PGC1α are required for this response in vivo. NA triggers physical and functional coupling between the α(1)-AR subtype (ADRA1A) and Gα(q) to promote adipocyte thermogenesis in a manner that is dependent on the effector proteins of the futile creatine cycle, creatine kinase B and tissue-non-specific alkaline phosphatase. Combined Gα(q) and Gα(s) signalling selectively in adipocytes promotes a continual rise in whole-body energy expenditure, and creatine kinase B is required for this effect. Thus, the ADRA1A–Gα(q)–futile creatine cycle axis is a key regulator of facultative and adaptive thermogenesis

Negative regulation of the MHC class I enhancer in adenovirus type 12-transformed cells

The expression of major histocompatibility class I genes is repressed by the adenovirus 12 (Ad12) E1A gene product in Ad12-transformed cells. In contrast, the E1A gene product of adenovirus 5 (Ad5), does not repress class I expression. The ability of these viruses to reduce the levels of class I expression correlates with their ability to form tumors in syngeneic immunocompetent animals, Ad12 being highly oncogenic and Ad5 being non-oncogenic. The reduction of class I expression, previously attributed to decreased rates of transcription, is at least in part the result of an altered enhancer activity, as determined by transient transfection assays. Enhancer activity is reduced in Ad12, compared to Ad5 transformed cells, suggesting that Ad12 E1A may inactivate the enhancer. We show here that Ad12-transformed cells contain increased levels of a DNA binding activity to the R2 element of the enhancer. The increased binding is due to a 55-60 kDa protein whose binding specificity resembles that of the members of the nuclear hormone receptor superfamily, especially of the thyroid hormone/retinoic acid receptors. Constructs where R2 and a positive enhancer element R1 are cloned upstream of the class I H-2K\sp{\rm b} promoter, show that R2 can repress transcription in Ad12-transformed cells, by interfering with the ability of R1 to activate transcription. In contrast, R2 does not repress R1 activity in Ad5-transformed cells. Repression of R1 activity is released in Ad12-transformed cells by treatment with retinoic acid (RA), while the R2 element in Ad5-transformed cells does not respond to RA. These results are consistent with a model where a member of the retinoic acid receptor family binds to the R2 enhancer element and, in the absence of RA, represses class enhancer activity in Ad12-transformed cells. This retinoic acid receptor is not active in Ad5-transformed cells, since the R2 element in these cells cannot repress R1 activity, nor respond to retinoic acid

A Genetic Analysis of . . .

To find novel components in the glucocorticoid signal transduction pathway, we performed a yeast genetic screen to identify ligand-effect modulators (LEMs), proteins that modulate the cellular response to hormone. We isolated several mutants that conferred increased glucocorticoid receptor (GR) activity in response to dexamethasone and analyzed two of them in detail. These studies identify two genes, LEM3 and LEM4, which correspond to YNL323w and ERG6, respectively. LEM3 is a putative transmembrane protein of unknown function, and ERG6 is a methyltransferase in the ergosterol biosynthetic pathway. Analysi

Peroxisome proliferator-activated receptor γ coactivator 1 (PGC-1)- and estrogen-related receptor (ERR)-induced regulator in muscle 1 (Perm1) is a tissue-specific regulator of oxidative capacity in skeletal muscle cells

Mitochondrial oxidative metabolism and energy transduction pathways are critical for skeletal and cardiac muscle function. The expression of genes important for mitochondrial biogenesis and oxidative metabolism are under the control of members of the peroxisome proliferator-activated receptor γ coactivator 1 (PGC-1) family of transcriptional coactivators and the estrogen-related receptor (ERR) subfamily of nuclear receptors. Perturbations in PGC-1 and/or ERR activities have been associated with alterations in capacity for endurance exercise, rates of muscle atrophy, and cardiac function. The mechanism(s) by which PGC-1 and ERR proteins regulate muscle-specific transcriptional programs is not fully understood. We show here that PGC-1α and ERRs induce the expression of a so far uncharacterized muscle-specific protein, PGC-1- and ERR-induced regulator in muscle 1 (Perm1), which regulates the expression of selective PGC-1/ERR target genes. Perm1 is required for the basal as well as PGC-1α-enhanced expression of genes with roles in glucose and lipid metabolism, energy transfer, and contractile function. Silencing of Perm1 in cultured myotubes compromises respiratory capacity and diminishes PGC-1α-induced mitochondrial biogenesis. Our findings support a role for Perm1 acting downstream of PGC-1α and ERRs to regulate muscle-specific pathways important for energy metabolism and contractile function. Elucidating the function of Perm1 may enable novel approaches for the treatment of disorders with compromised skeletal muscle bioenergetics, such as mitochondrial myopathies and age-related/disease-associated muscle atrophies

Genetic Separation of FK506 Susceptibility and Drug Transport in the Yeast Pdr5 ATP-binding Cassette Multidrug Resistance Transporter

Overexpression of the yeast Pdr5 ATP-binding cassette transporter leads to pleiotropic drug resistance to a variety of structurally unrelated cytotoxic compounds. To identify Pdr5 residues involved in substrate recognition and/or drug transport, we used a combination of random in vitro mutagenesis and phenotypic screening to isolate novel mutant Pdr5 transporters with altered substrate specificity. A plasmid library containing randomly mutagenized PDR5 genes was transformed into appropriate drug-sensitive yeast cells followed by phenotypic selection of Pdr5 mutants. Selected mutant Pdr5 transporters were analyzed with respect to their expression levels, subcellular localization, drug resistance profiles to cycloheximide, rhodamines, antifungal azoles, steroids, and sensitivity to the inhibitor FK506. DNA sequencing of six PDR5 mutant genes identified amino acids important for substrate recognition, drug transport, and specific inhibition of the Pdr5 transporter. Mutations were found in each nucleotide-binding domain, the transmembrane domain 10, and, most surprisingly, even in predicted extracellular hydrophilic loops. At least some point mutations identified appear to influence folding of Pdr5, suggesting that the folded structure is a major substrate specificity determinant. Surprisingly, a S1360F exchange in transmembrane domain 10 not only caused limited substrate specificity, but also abolished Pdr5 susceptibility to inhibition by the immunosuppressant FK506. This is the first report of a mutation in a yeast ATP-binding cassette transporter that allows for the functional separation of substrate transport and inhibitor susceptibility

Activation of nuclear receptor coactivator PGC-1α by arginine methylation

Peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), a tissue-specific and inducible transcriptional coactivator for several nuclear receptors, plays a key role in energy metabolism. We report here that PGC-1α coactivator activity is potentiated by arginine methylation by protein arginine methyltransferase 1 (PRMT1), another nuclear receptor coactivator. Mutation of three substrate arginines in the C-terminal region of PGC-1α abolished the cooperative coactivator function of PGC-1α and PRMT1, and compromised the ability of PGC-1α to induce endogenous target genes. Finally, endogenous PRMT1 contributes to PGC-1α coactivator activity, and to the induction of genes important for mitochondrial biogenesis