. Glycogen branches out: new perspectives on the role of glycogen metabolism in the integration of metabolic pathways. Am J Physiol Endocrinol Metab 291: E1-E8, 2006; doi:10.1152/ajpendo.00652.2005.-Glycogen is the storage form of carbohydrate for virtually every organism from yeast to primates. Most mammalian tissues store glucose as glycogen, with the major depots located in muscle and liver. The French physiologist Claude Bernard first identified a starch-like substance in liver and muscle and coined the term glycogen, or "sugar former," in the 1850s. During the 150 years since its identification, researchers in the field of glycogen metabolism have made numerous discoveries that are now recognized as significant milestones in biochemistry and cell signaling. Even so, more questions remain, and studies continue to demonstrate the complexity of the regulation of glycogen metabolism. Under classical definitions, the functions of glycogen seem clear: muscle glycogen is degraded to generate ATP during increased energy demand, whereas hepatic glycogen is broken down for release of glucose into the bloodstream to supply other tissues. However, recent findings demonstrate that the roles of glycogen metabolism in energy sensing, integration of metabolic pathways, and coordination of cellular responses to hormonal stimuli are far more complex. GLYCOGEN STRUCTURE AND KEY ENZYMES OF ITS METABOLISMGLYCOGEN SYNTHESIS follows a simple but strictly ordered process, resulting in a complex structure (38, 54). The initiation of glycogen synthesis is provided by self-glucosylation of glycogenin (55). To this oligosaccharide primer, glycogen synthase utilizes UDP-glucose to add glucose molecules by ␣1-4 linkages, which is the rate-controlling step of glycogen synthesis. Glycogen synthase activity is controlled by covalent modification, allosteric activation, and enzymatic translocation. The enzyme is phosphorylated on up to nine residues, resulting in its progressive inactivation and decreased sensitivity to allosteric activators (31). Binding of glucose 6-phosphate (G-6-P) to glycogen synthase causes unfolding of the enzyme, resulting in allosteric activation that overrides inhibition by phosphorylation. In addition, G-6-P binding promotes conformational changes that favor dephosphorylation of the enzyme. Translocation of glycogen synthase to glycogen particles in response to stimuli such as insulin represents a third mechanism by which enzymatic activity is regulated (13,42,49). When the elongating glycogen chain consists of at least 11 residues, branching enzyme transfers a chain of seven molecules to another chain by an ␣1-6 bond. Thus glycogen synthase elongates the glycogen chain, and branching enzyme produces new branches, creating a molecule with a helical structure of 12 concentric tiers (38).For glycogen degradation, the synchronous activities of glycogen phosphorylase and debranching enzyme are required.Glycogen phosphorylase, which catalyzes the rate-limiting step of glycogenolysis, cleaves ␣1-4 linkages to remove gl...

Protein phosphatase-1 (PP1) plays an important role in the regulation of glycogen synthesis by insulin. Protein targeting to glycogen (PTG) enhances glycogen accumulation by increasing PP1 activity against glycogenmetabolizing enzymes. However, the specificity of PTG's effects on cellular dephosphorylation and glucose metabolism is unclear. Overexpression of PTG in 3T3-L1 adipocytes using a doxycycline-controllable adenoviral construct resulted in a 10 -20-fold increase in PTG levels and an 8-fold increase in glycogen levels. Inclusion of 1 g/ml doxycycline in the media suppressed PTG expression, and fully reversed all PTG-dependent effects. Infection of 3T3-L1 adipocytes with the PTG adenovirus caused a marked dephosphorylation and activation of glycogen synthase. The effects of PTG seemed specific, because basal and insulin-stimulated phosphorylation of a variety of signaling proteins was unaffected. Indeed, glycogen synthase was the predominant protein whose phosphorylation state was decreased in 32 P-labeled cells. PTG overexpression did not alter PP1 protein levels but increased PP1 activity 6-fold against phosphorylase in vitro. In contrast, there was no change in PP1 activity measured using myelin basic protein, suggesting that PTG overexpression specifically directed PP1 activity against glycogen-metabolizing enzymes. To investigate the metabolic consequences of altering PTG levels, glucose uptake and storage in 3T3-L1 adipocytes was measured. PTG overexpression did not affect 2-deoxy-glucose transport rates in basal and insulin-stimulated cells but dramatically enhanced glycogen synthesis rates under both conditions. Despite the large increases in cellular glucose flux upon PTG overexpression, basal and insulin-stimulated glucose incorporation into lipid were unchanged. Cumulatively, these data indicate that PTG overexpression in 3T3-L1 adipocytes discretely stimulates PP1 activity against glycogen synthase and phosphorylase, resulting in a marked and specific increase in glucose uptake and storage as glycogen.Insulin exerts its anabolic actions by promoting the uptake and storage of glucose and lipids in target tissues. Insulin coordinates the stimulation of the translocation of both glucose and fatty acid transporters to the cell surface (1, 2) and the modulation of the activity of key metabolic enzymes (3). In addition, insulin inhibits hepatic glycogenolysis and adipocytic triglyceride breakdown, further lowering excess glucose and free fatty acids levels in the bloodstream. Disruption of the complex interplay between lipid and carbohydrate metabolism results in the development of insulin resistance and type II diabetes (4, 5). However, the molecular mechanisms by which insulin potently regulates energy uptake and storage in vivo are not fully understood.Glycogen synthase activity is stimulated by insulin in liver, muscle, and adipose tissue via protein dephosphorylation, allosteric activation, and enzymatic translocation. Glycogen synthase is phosphorylated on up to nine residues by a variety of ki...

High levels of saturated, branched-chain fatty acids are deleterious to cells and animals, resulting in lipid accumulation and cytotoxicity. Although fatty acid binding proteins (FABPs) are thought to be protective, this hypothesis has not previously been examined. Phytanic acid (branched chain, 16-carbon backbone) induced lipid accumulation in L cell fibroblasts similar to that observed with palmitic acid (unbranched, C16): triacylglycerol ≫ free fatty acid > cholesterol > cholesteryl ester ≫ phospholipid. Although expression of sterol carrier protein (SCP)-2, SCP-x, or liver FABP (L-FABP) in transfected L cells reduced [3H]phytanic acid uptake (57–87%) and lipid accumulation (21–27%), nevertheless [3H]phytanic acid oxidation was inhibited (74–100%) and phytanic acid toxicity was enhanced in the order L-FABP ≫ SCP-x > SCP-2. These effects differed markedly from those of [3H]palmitic acid, whose uptake, oxidation, and induction of lipid accumulation were not reduced by L-FABP, SCP-2, or SCP-x expression. Furthermore, these proteins did not enhance the cytotoxicity of palmitic acid. In summary, intracellular FABPs reduce lipid accumulation induced by high levels of branched-chain but not straight-chain saturated fatty acids. These beneficial effects were offset by inhibition of branched-chain fatty acid oxidation that correlated with the enhanced toxicity of high levels of branched-chain fatty acid.

Epigallocatechin gallate (EGCG), the major polyphenol in green tea, acutely stimulates production of nitric oxide (NO) from vascular endothelium to reduce hypertension, and improve endothelial dysfunction in SHR rats. Herein, we explored additional mechanisms whereby EGCG may mediate beneficial cardiovascular actions. When compared with vehicle-treated controls, EGCG treatment (2.5 μM, 8 h) of human aortic endothelial cells (HAEC) caused a ~3-fold increase in hemeoxygenase-1 (HO-1) mRNA and protein with comparable increases in HO-1 activity. This was unaffected by pre-treatment of cells with wortmannin, LY294002, PD98059, or L-NAME (PI 3-kinase, MEK, and NO synthase inhibitors, respectively). Pre-treatment of HAEC with SB203580 (p38 MAPK inhibitor) or siRNA knockdown of p38 MAPK completely blocked EGCG-stimulated induction of HO-1. EGCG treatment also inhibited TNF-α-stimulated expression of VCAM-1 and decreased adhesion of monocytes to HAEC. siRNA knockdown of HO-1, p38 MAPK, or Nrf-2 blocked these inhibitory actions of EGCG. In HAEC transiently transfected with a human HO-1 promoter luciferase reporter (or an isolated Nrf-2 responsive region), luciferase activity increased in response to EGCG. This was inhibitable by SB203580 pre-treatment. EGCG-stimulated expression of HO-1 and Nrf-2 was blocked by siRNA knockdown of Nrf-2 or p38 MAPK. Finally, liver from mice chronically treated with EGCG had increased HO-1 and decreased VCAM-1 expression. Thus, in vascular endothelium, EGCG requires p38 MAPK to increase expression of Nrf-2 that drives expression of HO-1 resulting in increased HO-1 activity. Increased HO-1 expression may underlie anti-inflammatory actions of EGCG in vascular endothelium that may help mediate beneficial cardiovascular actions of green tea.

-Adipocytes express the rate-limiting enzymes required for glycogen metabolism and increase glycogen synthesis in response to insulin. However, the physiological function of adipocytic glycogen in vivo is unclear, due in part to the low absolute levels and the apparent biophysical constraints of adipocyte morphology on glycogen accumulation. To further study the regulation of glycogen metabolism in adipose tissue, transgenic mice were generated that overexpressed the protein phosphatase-1 (PP1) glycogen-targeting subunit (PTG) driven by the adipocyte fatty acid binding protein (aP2) promoter. Exogenous PTG was detected in gonadal, perirenal, and brown fat depots, but it was not detected in any other tissue examined. PTG overexpression resulted in a modest redistribution of PP1 to glycogen particles, corresponding to a threefold increase in the glycogen synthase activity ratio. Glycogen synthase protein levels were also increased twofold, resulting in a combined greater than sixfold enhancement of basal glycogen synthase specific activity. Adipocytic glycogen levels were increased 200-to 400-fold in transgenic animals, and this increase was maintained to 1 yr of age. In contrast, lipid metabolism in transgenic adipose tissue was not significantly altered, as assessed by lipogenic rates, weight gain on normal or high-fat diets, or circulating free fatty acid levels after a fast. However, circulating and adipocytic leptin levels were doubled in transgenic animals, whereas adiponectin expression was unchanged. Cumulatively, these data indicate that murine adipocytes are capable of storing far higher levels of glycogen than previously reported. Furthermore, these results were obtained by overexpression of an endogenous adipocytic protein, suggesting that mechanisms may exist in vivo to maintain adipocytic glycogen storage at a physiological set point.insulin; glycogen synthesis; lipogenesis; protein phosphatase-1; targeting subunit CIRCULATING ENERGY SOURCES are maintained in a narrow homeostatic range across a wide variety of physiological conditions through the coordinate regulation of energy uptake, consumption, storage, and release by the principal metabolic tissues, namely adipose tissue, liver, and skeletal muscle. During times of energy deficit, adipose tissue is a primary site for energy provision via the hydrolysis of stored triglyceride to release free fatty acid (FFA) for ATP production and glycerol for hepatic gluconeogenesis. Conversely, following a meal, dietary lipid is stored by adipose tissue, and glucose is disposed of as glycogen by the skeletal muscle and to a lesser extent by the liver. Alternately, glucose can be utilized postprandially by the liver and adipocytes for de novo lipogenesis and long-term storage as triglyceride. However, adipocytes also store glucose as glycogen, albeit at substantially lower rates than in skeletal muscle and liver, so the physiological role of adipocytic glycogen metabolism remains unclear.In addition to its role in lipid metabolism, adipose tissue functions as an...