Trim32 reduces PI3K–Akt–FoxO signaling in muscle atrophy by promoting plakoglobin–PI3K dissociation

Activation of the PI3K–Akt–FoxO pathway induces cell growth, whereas its inhibition reduces cell survival and, in muscle, causes atrophy. Here, we report a novel mechanism that suppresses PI3K–Akt–FoxO signaling. Although skeletal muscle lacks desmosomes, it contains multiple desmosomal components, including plakoglobin. In normal muscle plakoglobin binds the insulin receptor and PI3K subunit p85 and promotes PI3K–Akt–FoxO signaling. During atrophy, however, its interaction with PI3K–p85 is reduced by the ubiquitin ligase Trim32 (tripartite motif containing protein 32). Inhibition of Trim32 enhanced plakoglobin binding to PI3K–p85 and promoted PI3K–Akt–FoxO signaling. Surprisingly, plakoglobin overexpression alone enhanced PI3K–Akt–FoxO signaling. Furthermore, Trim32 inhibition in normal muscle increased PI3K–Akt–FoxO signaling, enhanced glucose uptake, and induced fiber growth, whereas plakoglobin down-regulation reduced PI3K–Akt–FoxO signaling, decreased glucose uptake, and caused atrophy. Thus, by promoting plakoglobin–PI3K dissociation, Trim32 reduces PI3K–Akt–FoxO signaling in normal and atrophying muscle. This mechanism probably contributes to insulin resistance during fasting and catabolic diseases and perhaps to the myopathies and cardiomyopathies seen with Trim32 and plakoglobin mutations.     

FoxO1对下丘脑摄食中枢的影响研究进展

下丘脑是人体的摄食中枢,它通过抑制食欲的阿黑皮素原(POMC)神经元和促进食欲的神经肽相关蛋白(AgRP)神经元调节摄食及能量代谢。叉头转录因子O亚族1(FoxO1)是胰岛素信号通路和瘦素信号通路中重要的调节蛋白,FoxO1的生理作用是促进下丘脑Agrp基因表达、抑制Pomc基因表达,抑制瘦素信号通路的转录激活因子3(STAT3)蛋白对Pomc基因转录的促进作用,从而促进食欲。瘦素和胰岛素均可激活经典的IRS/PI(3)K/Akt信号通路,使FoxO1磷酸化失去活性,抑制食欲。此外,沉默信息调节因子Sirt1也可以通过去乙酰化,影响FoxO1的转录活性。本文综述了胰岛素、瘦素、Sirt1通过FoxO1调节下丘脑摄食中枢的作用机制。     

Hepatic FoxO1 Integrates Glucose Utilization and Lipid Synthesis through Regulation of Chrebp O-Glycosylation

In liver, glucose utilization and lipid synthesis are inextricably intertwined. When glucose availability exceeds its utilization, lipogenesis increases, leading to increased intrahepatic lipid content and lipoprotein secretion. Although the fate of three-carbon metabolites is largely determined by flux rate through the relevant enzymes, insulin plays a permissive role in this process. But the mechanism integrating insulin receptor signaling to glucose utilization with lipogenesis is unknown. Forkhead box O1 (FoxO1), a downstream effector of insulin signaling, plays a central role in hepatic glucose metabolism through the regulation of hepatic glucose production. In this study, we investigated the mechanism by which FoxO1 integrates hepatic glucose utilization with lipid synthesis. We show that FoxO1 overexpression in hepatocytes reduces activity of carbohydrate response element binding protein (Chrebp), a key regulator of lipogenesis, by suppressing O-linked glycosylation and reducing the protein stability. FoxO1 inhibits high glucose- or O-GlcNAc transferase (OGT)-induced liver-pyruvate kinase (L-PK) promoter activity by decreasing Chrebp recruitment to the L-PK promoter. Conversely, FoxO1 ablation in liver leads to the enhanced O-glycosylation and increased protein level of Chrebp owing to decreased its ubiquitination. We propose that FoxO1 regulation of Chrebp O-glycosylation is a mechanism linking hepatic glucose utilization with lipid synthesis.     

Modulation of FoxO1 phosphorylation/acetylation by baicalin during aging

Baicalin is a flavonoid known to modify various redox-related biological activities. Included is its ability to suppress reactive species (RS) producing activity and modulate nuclear factor-κB through cellular redox regulation with enhanced thiol ability. FoxO regulates various genes that are known to be involved in cellular metabolism related to cell death and the oxidative stress response. One such case is the prevention of FoxO1 expression by activated insulin-induced phosphatidylinositol 3-kinase (PI3K)/Akt, which leads to increased oxidative stress and aging processes. In the present study, we attempted to elucidate the molecular modulation of antioxidant baicalin on the insulin-induced FoxO1 inactivation. We used HEK293T cultured cells and kidney tissue isolated from 24-month-old rats treated with baicalin at a dose of 10 or 20 mg/kg/day for 10 days. We found that baicalin enhanced catalase and suppressed RS production in cell system and in isolated kidney tissue in contrast to the nontreated aged rats. Results also showed activation of insulin signaling (PI3K/Akt), FoxO1 phosphorylation/acetylation and the down-regulation of catalase and manganese superoxide dismutase, both of which are FoxO1-targeting genes. Furthermore, baicalin-treated rats showed a decreased FoxO1 phosphorylation via PI3K/Akt cascade and FoxO1 acetylation by the cAMP-response element-binding protein binding protein (CBP). These results strongly suggest that treatment with baicalin influenced phosphorylation/acetylation of FoxO1 by up-regulating PI3K/Akt signaling through insulin in aged rats. Our results further reveal that baicalin regulated FoxO1 phosphorylation via PI3K/Akt by insulin and FoxO1 acetylation by the interaction of CBP and SIRT1, leading to changes in catalase gene expression during aging.     

Hepatic insulin resistance in ob/ob mice involves increases in ceramide,aPKC activity,and selective impairment of Akt-dependent FoxO1 phosphorylation

Pathogenesis of insulin resistance in leptin-deficient ob/ob mice is obscure. In another form of diet-dependent obesity, high-fat-fed mice, hepatic insulin resistance involves ceramide-induced activation of atypical protein kinase C (aPKC), which selectively impairs protein kinase B (Akt)-dependent forkhead box O1 protein (FoxO1) phosphorylation on scaffolding protein, 40 kDa WD(tryp-x-x-asp)-repeat propeller/FYVE protein (WD40/ProF), thereby increasing gluconeogenesis. Resultant hyperinsulinemia activates hepatic Akt and mammalian target of rapamycin C1, and further activates aPKC; consequently, lipogenic enzyme expression increases, and insulin signaling in muscle is secondarily impaired. Here, in obese minimally-diabetic ob/ob mice, hepatic ceramide and aPKC activity and its association with WD40/ProF were increased. Hepatic Akt activity was also increased, but Akt associated with WD40/ProF was diminished and accounted for reduced FoxO1 phosphorylation and increased gluconeogenic enzyme expression. Most importantly, liver-selective inhibition of aPKC decreased aPKC and increased Akt association with WD40/ProF, thereby restoring FoxO1 phosphorylation and reducing gluconeogenic enzyme expression. Additionally, lipogenic enzyme expression diminished, and insulin signaling in muscle, glucose tolerance, obesity, hepatosteatosis, and hyperlipidemia improved. In conclusion, hepatic ceramide accumulates in response to CNS-dependent dietary excess irrespective of fat content; hepatic insulin resistance is prominent in ob/ob mice and involves aPKC-dependent displacement of Akt fromWD40/ProF and subsequent impairment of FoxO1 phosphorylation and increased expression of hepatic gluconeogenic and lipogenic enzymes; and hepatic alterations diminish insulin signaling in muscle.     

FoxO转录因子

FoxO家族是转录调节因子 ,也是INS IGF 1信号通路中的关键分子。FoxO基因在进化上高度保守 ,其氨基酸序列中含有 3个高度保守PKB磷酸化基序。FoxO受PI3K PKB磷酸化级联通路的调节 ,其活性与磷酸化状态直接相关。FoxO对细胞增殖、细胞凋亡等生理过程有重要调节作用 ,并可能在免疫系统发育中对免疫细胞的凋亡及亚群间的平衡起一定调节作用。     

Liquid fructose down-regulates liver insulin receptor substrate 2 and gluconeogenic enzymes by modifying nutrient sensing factors in rats

High consumption of fructose-sweetened beverages has been linked to a high prevalence of chronic metabolic diseases. We have previously shown that a short course of fructose supplementation as a liquid solution induces glucose intolerance in female rats. In the present work, we characterized the fructose-driven changes in the liver and the molecular pathways involved. To this end, female rats were supplemented or not with liquid fructose (10%, w/v) for 7 or 14 days. Glucose and pyruvate tolerance tests were performed, and the expression of genes related to insulin signaling, gluconeogenesis and nutrient sensing pathways was evaluated.Fructose-supplemented rats showed increased plasma glucose excursions in glucose and pyruvate tolerance tests and reduced hepatic expression of several genes related to insulin signaling, including insulin receptor substrate 2 (IRS-2). However, the expression of key gluconeogenic enzymes, glucose-6-phosphatase and phosphoenolpyruvate carboxykinase, was reduced. These effects were caused by an inactivation of hepatic forkhead box O1 (FoxO1) due to an increase in its acetylation state driven by a reduced expression and activity of sirtuin 1 (SIRT1). Further contributing to FoxO1 inactivation, fructose consumption elevated liver expression of the spliced form of X-box-binding-protein-1 as a consequence of an increase in the activity of the mammalian target of rapamycin 1 and protein 38-mitogen activated protein kinase (p38-MAPK). Liquid fructose affects both insulin signaling (IRS-2 and FoxO1) and nutrient sensing pathways (p38-MAPK, mTOR and SIRT1), thus disrupting hepatic insulin signaling without increasing the expression of key gluconeogenic enzymes.     

FoxO1 target Gpr17 activates AgRP neurons to regulate food intake

Hypothalamic neurons expressing Agouti-related peptide (AgRP) are critical for initiating food intake, but druggable biochemical pathways that control this response remain elusive. Thus, genetic ablation of insulin or leptin signaling in AgRP neurons is predicted to reduce satiety but fails to do so. FoxO1 is a shared mediator of both pathways, and its inhibition is required to induce satiety. Accordingly, FoxO1 ablation in AgRP neurons of mice results in reduced food intake, leanness, improved glucose homeostasis, and increased sensitivity to insulin and leptin. Expression profiling of flow-sorted FoxO1-deficient AgRP neurons identifies G-protein-coupled receptor Gpr17 as a FoxO1 target whose expression is regulated by nutritional status. Intracerebroventricular injection of Gpr17 agonists induces food intake, whereas Gpr17 antagonist cangrelor curtails it. These effects are absent in Agrp-Foxo1 knockouts, suggesting that pharmacological modulation of this pathway has therapeutic potential to treat obesity.     

Forkhead transcription factor FoxO1 inhibits insulin- and transforming growth factor-β-stimulated plasminogen activator inhibitor-1 expression

Elevated levels of plasminogen activator inhibitor-1 (PAI-1) are considered a risk factor for chronic liver disease in patients with hyperinsulinemia. Insulin increases the expression of PAI-1, and inactivates the forkhead box-containing protein FoxO1. We were interested in whether the inactivation of FoxO1 is involved in the activation of PAI-1 expression under conditions of insulin stimulation. Here, we examined whether adenoviral-mediated expression of a constitutively active form of FoxO1 (Ad-CA-FoxO1) inhibited insulin-stimulated PAI-1 expression in human HepG2 hepatocellular liver carcinoma cells and mouse AML12 hepatocytes. Treatment of cells with insulin increased PAI-1 gene expression, and this effect was abolished by Ad-CA-FoxO1. Insulin also increased the transforming growth factor (TGF)-β-induced expression of PAI-1 mRNA, and Ad-CA-FoxO1 inhibited this effect. Transient transfection assays using a reporter gene under the control of the PAI-1 promoter revealed that CA-FoxO1 inhibits Smad3-stimulated PAI-1 promoter activity. Taken together, our results indicate that FoxO1 inhibits PAI-1 expression through the inhibition of TGF-β/Smad-mediated signaling pathways. Our data also suggest that in the hyperinsulinemic state, FoxO1 is inactivated by increased levels of insulin, and does not function as an inhibitor of TGF-β-induced PAI-1 expression.