The E3 Ligase MuRF1 degrades myosin heavy chain protein in dexamethasone-treated skeletal muscle.

Skeletal muscle atrophy occurs as a side effect of treatment with synthetic glucocorticoids such as dexamethasone (DEX) and is a hallmark of cachectic syndromes associated with increased cortisol levels. The E3 ubiquitin ligase MuRF1 (muscle RING finger protein 1) is transcriptionally upregulated by DEX treatment. Differentiated myotubes treated with DEX undergo depletion of myosin heavy chain protein (MYH), which physically associates with MuRF1. This loss of MYH can be blocked by inhibition of MuRF1 expression. When wild-type and MuRF1(-/-) mice are treated with DEX, the MuRF1(-/-) animals exhibit a relative sparing of MYH. In vitro, MuRF1 is shown to function as an E3 ubiquitin ligase for MYH. These data identify the mechanism by which MYH is depleted under atrophy conditions and demonstrate that inhibition of a single E3 ligase, MuRF1, is sufficient to maintain this important sarcomeric protein.

Genetic deletion of Trb3, the mammalian Drosophila tribbles homolog, displays normal hepatic insulin signaling and glucose homeostasis.

Trb3, a mammalian homolog of Drosophila tribbles, was proposed as a suppressor of Akt activity, predominantly in conditions of fasting and diabetes. Given these prior studies, we sought to determine whether Trb3 plays a major role in modulating hepatic insulin sensitivity. To answer this question, we produced mice in which a lacZ reporter was knocked into the locus containing the gene Trib3, resulting in a Trib3 null animal. Trib3 expression analyses demonstrated that the Trib3 is expressed in liver, adipose tissues, heart, kidney, lung, skin, small intestine, stomach, and denervated, but not normal, skeletal muscle. Trib3(-/-) mice are essentially identical to their wild-type littermates in overall appearance and body composition. Phenotypic analysis of Trib3(-/-) mice did not detect any alteration in serum glucose, insulin, or lipid levels; glucose or insulin tolerance; or energy metabolism. Studies in Trib3(-/-) hepatocytes revealed normal Akt and glycogen synthase kinase- 3beta phosphorylation patterns, glycogen levels, and expressions of key regulatory gluconeogenic and glycolytic genes. These data demonstrate that deletion of Trib3 has minimal effect on insulin-induced Akt activation in hepatic tissue, and, as such, they question any nonredundant role for Trb3 in the maintenance of glucose and energy homeostasis in mice.

Absence of the lipid phosphatase SHIP2 confers resistance to dietary obesity.

Genetic ablation of Inppl1, which encodes SHIP2 (SH2-domain containing inositol 5-phosphatase 2), was previously reported to induce severe insulin sensitivity, leading to early postnatal death. In the previous study, the targeting construct left the first eighteen exons encoding Inppl1 intact, generating a Inppl1(EX19-28-/-) mouse, and apparently also deleted a second gene, Phox2a. We report a new SHIP2 knockout (Inppl1(-/-)) targeted to the translation-initiating ATG, which is null for Inppl1 mRNA and protein. Inppl1(-/-) mice are viable, have normal glucose and insulin levels, and normal insulin and glucose tolerances. The Inppl1(-/-) mice are, however, highly resistant to weight gain when placed on a high-fat diet. These results suggest that inhibition of SHIP2 would be useful in the effort to ameliorate diet-induced obesity, but call into question a dominant role of SHIP2 in modulating glucose homeostasis.

Conditional activation of akt in adult skeletal muscle induces rapid hypertrophy.

Skeletal muscle atrophy is a severe morbidity caused by a variety of conditions, including cachexia, cancer, AIDS, prolonged bedrest, and diabetes. One strategy in the treatment of atrophy is to induce the pathways normally leading to skeletal muscle hypertrophy. The pathways that are sufficient to induce hypertrophy in skeletal muscle have been the subject of some controversy. We describe here the use of a novel method to produce a transgenic mouse in which a constitutively active form of Akt can be inducibly expressed in adult skeletal muscle and thereby demonstrate that acute activation of Akt is sufficient to induce rapid and significant skeletal muscle hypertrophy in vivo, accompanied by activation of the downstream Akt/p70S6 kinase protein synthesis pathway. Upon induction of Akt in skeletal muscle, there was also a significant decrease in adipose tissue. These findings suggest that pharmacologic approaches directed toward activating Akt will be useful in inducing skeletal muscle hypertrophy and that an increase in lean muscle mass is sufficient to decrease fat storage.

The IGF-1/PI3K/Akt pathway prevents expression of muscle atrophy-induced ubiquitin ligases by inhibiting FOXO transcription factors.

Skeletal muscle size depends upon a dynamic balance between anabolic (or hypertrophic) and catabolic (or atrophic) processes. Previously, no link between the molecular mediators of atrophy and hypertrophy had been reported. We demonstrate a hierarchy between the signals which mediate hypertrophy and those which mediate atrophy: the IGF-1/PI3K/Akt pathway, which has been shown to induce hypertrophy, prevents induction of requisite atrophy mediators, namely the muscle-specific ubiquitin ligases MAFbx and MuRF1. Moreover, the mechanism for this inhibition involves Akt-mediated inhibition of the FoxO family of transcription factors; a mutant form of FOXO1, which prevents Akt phosphorylation, thereby prevents Akt-mediated inhibition of MuRF1 and MAFbx upregulation. Our study thus defines a previously uncharacterized function for Akt, which has important therapeutic relevance: Akt is not only capable of activating prosynthetic pathways, as previously demonstrated, but is simultaneously and dominantly able to suppress catabolic pathways, allowing it to prevent glucocorticoid and denervation-induced muscle atrophy.

The growth arrest-specific gene product Gas6 promotes the survival of human oligodendrocytes via a phosphatidylinositol 3-kinase-dependent pathway.

Microarray analysis revealed that transcripts for the Axl and Mer receptor tyrosine kinases are expressed at high levels in O4+-immunopanned oligodendrocytes isolated from second trimester human fetal spinal cord. In humans the sole known ligand for the Axl/Rse/Mer kinases is growth arrest-specific gene 6 (Gas6), which in the CNS is secreted by neurons and endothelial cells. We hypothesized that Gas6 is a survival factor for oligodendrocytes and receptor activation signals downstream to the phosphatidylinositol 3 (PI3)-kinase/Akt pathway to increase cell survival in the absence of cell proliferation. To test this hypothesis, we grew enriched human oligodendrocytes for 6 d on a monolayer of NIH3T3 cells stably expressing Gas6. CNP+ oligodendrocytes on Gas6-secreting 3T3 cells had more primary processes and arborizations than those plated solely on 3T3 cells. Also, a twofold increase in CNP+ and MBP+ oligodendrocytes was observed when they were plated on the Gas6-secreting cells. The effect was abolished in the presence of Axl-Fc but remained unchanged in the presence of the irrelevant receptor fusion molecule TrkA-Fc. A significant decrease in CNP+/TUNEL+ oligodendrocytes was observed when recombinant human Gas6 (rhGas6) was administered to oligodendrocytes plated on poly-L-lysine, supporting a role for Gas6 signaling in oligodendrocyte survival during a period of active myelination in human fetal spinal cord development. PI3-kinase inhibitors blocked the anti-apoptotic effect of rhGas6, whereas a MEK/ERK inhibitor had no effect. Thus Gas6 sustains human fetal oligodendrocyte viability by receptor activation and downstream signaling via the PI3-kinase/Akt pathway.