FOXO1 activates glutamine synthetase gene in mouse skeletal muscles through a region downstream of 3′-UTR: possible contribution to ammonia detoxification

Abstract

Skeletal muscle is a reservoir of energy in the form of protein, which is degraded under catabolic conditions, resulting in the formation of amino acids and ammonia as a byproduct. The expression of FOXO1, a forkhead-type transcription factor, increases during starvation and exercise. In agreement, transgenic FOXO1-Tg mice that overexpress FOXO1 in skeletal muscle exhibit muscle atrophy. The aim of this study was to examine the role of FOXO1 in amino acid metabolism. The mRNA and protein expressions of glutamine synthetase (GS) were increased in skeletal muscle of FOXO1-Tg mice. Fasting induced FOXO1 and GS expression in wild-type mice but hardly increased GS expression in muscle-specific FOXO1 knockout (FOXO1-KO) mice. Activation of FOXO1 also increased GS mRNA and protein expression in C2C12 myoblasts. Using a transient transfection reporter assay, we observed that FOXO1 activated the GS reporter construct. Mutation of a putative FOXO1-binding consensus sequence in the downstream genomic region of GS decreased basal and FOXO1-dependent reporter activity significantly. A chromatin immunoprecipitation assay showed that FOXO1 was recruited to the 3′ region of GS in C2C12 myoblasts. These results suggest that FOXO1 directly upregulates GS expression. GS is considered to mediate ammonia clearance in skeletal muscle. In agreement, an intravenous ammonia challenge increased blood ammonia concentrations to a twofold higher level in FOXO1-KO than in wild-type mice, demonstrating that the capacity for ammonia disposal correlated inversely with the expression of GS in muscle. These data indicate that FOXO1 plays a role in amino acid metabolism during protein degradation in skeletal muscle.

Restricting Glutamine or Glutamine-Dependent Purine and Pyrimidine Syntheses Promotes Human T Cells with High FOXP3 Expression and Regulatory Properties

“…Although entirely speculative at present, these might include FOXO transcription factors previously shown to support GS expression in skeletal muscle and cell lines, as well as FOXP3 expression in T cells (36)(37)(38). The extensive regulation of FOXOs includes feedback loops to control their own transcription and activation as well as FOXP3 within the PTEN/PI3K/Akt/mTOR pathway (38,39).…”

Vitamin D Attenuates FOXO1-Target Atrophy Gene Expression in C2C12 Muscle Cells

“…In addition, we showed that FOXO1 activates the genes involved in protein degradation (6,7). FOXO1 activates the expression of atrophy-related genes such as atrogin 1 and cathepsin L in various muscle atrophyrelated conditions, including a glucocorticoid treatment (6)(7)(8). These findings suggest that FOXO1 in skeletal muscles plays an important role in muscle atrophy.…”

“…Specifically, based on data that FOXO1 gene expression is induced by energy deprivation (4), we developed transgenic mice that overexpressed FOXO1 (FOXO1 Tg mice) and showed muscle atrophy (5). In addition, we showed that FOXO1 activates the genes involved in protein degradation (6,7). FOXO1 activates the expression of atrophy-related genes such as atrogin 1 and cathepsin L in various muscle atrophyrelated conditions, including a glucocorticoid treatment (6)(7)(8).…”

Assessment of a western blot signal for the Bcnt/Cfdp1, a tentative component of Srcap chromatin remodeling complex; trial to overcome off-target problems

“…Bloom-deficient ES cells, selected for populations of homozygous mutant cells 4 0 0 following mitotic recombination [18], and was listed as a cell line of mouse Bcnt/Cfdp1 knockout cells [21], but the expected cells were not obtained so far (W. Kobayashi, 4 1 3 personal communication). Finally, we were able to assign the signal of endogenous 4 1 4 mBcnt/Cfdp1 detected by two Abs raised against unrelated immunogens.…”