scholarly journals The “Glycogen Granule” Revisited

1994 ◽  
Vol 2 (7) ◽  
pp. 16-18
Author(s):  
Krystyna Rybicka
Keyword(s):  
Electron Microscopy ◽  
Glycogen Synthase ◽  
Glycogen Synthesis ◽  
Protein Component ◽  
Glycogen Granule ◽  
Regulatory Enzymes ◽  
Ionic Compounds ◽  
Cellular Organelles ◽  
Covalently Bound

There is a gap between biochemical findings and ultrastructural interpretation of “glycogen granules”. Biochemists have recognized that glycogen contains covalently bound proteins. These include enzymes involved in giycogen metabolism: glycogenin (protein primer responsible for initiation of glycogen synthesis), glycogen synthase and phosphorylase, and presumably other regulatory enzymes. The structures formed by the association of glycogen and protein have been called protein-glycogen complexes, considered as proteoglycans, or as dynamic cellular organelles, glycosomes.The question arises as to why the biochemical recognition of a protein component in glycosomes has not been acknowledged in electron microscopy (EM)? This protein is visible in every section stained by uranium (U) and lead (Pb) salts where it appears as 20-30 nm granules (Fig. 1). However, these granules are commonly interpreted as glycogen, despite the fact that glycogen does not react with ionic compounds and therefore cannot be stained by U-Pb.

scholarly journals Association of glycogen synthase phosphatase and phosphorylase phosphatase activities with membranes of hepatic smooth endoplasmic reticulum.

1979 ◽  
Vol 83 (2) ◽  
pp. 348-356 ◽  
Author(s):  
R N Margolis ◽  
R R Cardell ◽  
R T Curnow
Keyword(s):  
Endoplasmic Reticulum ◽  
Glycogen Synthase ◽  
Close Contact ◽  
Smooth Endoplasmic Reticulum ◽  
Glycogen Synthesis ◽  
Glycogen Metabolism ◽  
Hepatic Glycogen ◽  
Phosphatase Activities ◽  
Regulatory Enzymes ◽  
Glycogen Particles

A detailed investigation was conducted to determine the precise subcellular localization of the rate-limiting enzymes of hepatic glycogen metabolism (glycogen synthase and phosphorylase) and their regulatory enzymes (synthase phosphatase and phosphorylase phosphatase). Rat liver was homogenized and fractionated to produce soluble, rough and smooth microsomal fractions. Enzyme assays of the fractions were performed, and the results showed that glycogen synthase and phosphorylase were located in the soluble fraction of the livers. Synthase phosphatase and phosphorylase phosphatase activities were also present in soluble fractions, but were clearly identified in both rough and smooth microsomal fractions. It is suggested that the location of smooth endoplasmic reticulum (SER) within the cytosome forms a microenvironment within hepatocytes that establishes conditions necessary for glycogen synthesis (and degradation). Thus the location of SER in the cell determines regions of the hepatocyte that are rich in glycogen particles. Furthermore, the demonstration of the association of synthase phosphatase and phosphorylase phosphatase with membranes of SER may account for the close morphological association of SER with glycogen particles (i.e., disposition of SER membranes brings the membrane-bound regulatory enzymes in close contact with their substrates).

Scanning Secondary Electron Microscopy of Myofibrils Using Transmission Techniques

1975 ◽  
Vol 33 ◽  
pp. 556-557
Author(s):  
M. K. Lamvik
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
Carbon Films ◽  
Photographic Recording ◽  
Transmission Electron ◽  
Thin Carbon ◽  
The Common ◽  
Scanning Electron ◽  
Better Than ◽  
Cellular Organelles

When observing small objects such as cellular organelles by scanning electron microscopy, it is often valuable to use the techniques of transmission electron microscopy. The common practice of mounting and coating for SEM may not always be necessary. These possibilities are illustrated using vertebrate skeletal muscle myofibrils.Micrographs for this study were made using a Hitachi HFS-2 scanning electron microscope, with photographic recording usually done at 60 seconds per frame. The instrument was operated at 25 kV, with a specimen chamber vacuum usually better than 10-7 torr. Myofibrils were obtained from rabbit back muscle using the method of Zak et al. To show the component filaments of this contractile organelle, the myofibrils were partially disrupted by agitation in a relaxing medium. A brief centrifugation was done to clear the solution of most of the undisrupted myofibrils before a drop was placed on the grid. Standard 3 mm transmission electron microscope grids covered with thin carbon films were used in this study.

scholarly journals Pho85p, a cyclin-dependent protein kinase, and the Snf1p protein kinase act antagonistically to control glycogen accumulation in Saccharomyces cerevisiae.

1996 ◽  
Vol 16 (8) ◽  
pp. 4357-4365 ◽  
Author(s):  
D Huang ◽  
I Farkas ◽  
P J Roach
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Kinase ◽  
Kinase Activity ◽  
Glycogen Synthase ◽  
Glycogen Synthesis ◽  
Mutant Gene ◽  
Glycogen Synthase Kinase ◽  
Partial Purification ◽  
Glycogen Accumulation ◽  
Dependent Protein

In Saccharomyces cerevisiae, nutrient levels control multiple cellular processes. Cells lacking the SNF1 gene cannot express glucose-repressible genes and do not accumulate the storage polysaccharide glycogen. The impaired glycogen synthesis is due to maintenance of glycogen synthase in a hyperphosphorylated, inactive state. In a screen for second site suppressors of the glycogen storage defect of snf1 cells, we identified a mutant gene that restored glycogen accumulation and which was allelic with PHO85, which encodes a member of the cyclin-dependent kinase family. In cells with disrupted PHO85 genes, we observed hyperaccumulation of glycogen, activation of glycogen synthase, and impaired glycogen synthase kinase activity. In snf1 cells, glycogen synthase kinase activity was elevated. Partial purification of glycogen synthase kinase activity from yeast extracts resulted in the separation of two fractions by phenyl-Sepharose chromatography, both of which phosphorylated and inactivated glycogen synthase. The activity of one of these, GPK2, was inhibited by olomoucine, which potently inhibits cyclin-dependent protein kinases, and contained an approximately 36-kDa species that reacted with antibodies to Pho85p. Analysis of Ser-to-Ala mutations at the three potential Gsy2p phosphorylation sites in pho85 cells implicated Ser-654 and/or Thr-667 in PHO85 control of glycogen synthase. We propose that Pho85p is a physiological glycogen synthase kinase, possibly acting downstream of Snf1p.

Insulin promotes glycogen synthesis in the absence of GSK3 phosphorylation in skeletal muscle

2008 ◽  
Vol 294 (1) ◽  
pp. E28-E35 ◽  
Author(s):  
Michale Bouskila ◽  
Michael F. Hirshman ◽  
Jørgen Jensen ◽  
Laurie J. Goodyear ◽  
Kei Sakamoto
Keyword(s):  
Skeletal Muscle ◽  
Glucose Uptake ◽  
Muscle Glycogen ◽  
Glycogen Content ◽  
Glycogen Synthase ◽  
Glycogen Synthesis ◽  
Wild Type ◽  
Muscle Glycogen Synthesis ◽  
Mutant Forms

Insulin promotes dephosphorylation and activation of glycogen synthase (GS) by inactivating glycogen synthase kinase (GSK) 3 through phosphorylation. Insulin also promotes glucose uptake and glucose 6-phosphate (G-6- P) production, which allosterically activates GS. The relative importance of these two regulatory mechanisms in the activation of GS in vivo is unknown. The aim of this study was to investigate if dephosphorylation of GS mediated via GSK3 is required for normal glycogen synthesis in skeletal muscle with insulin. We employed GSK3 knockin mice in which wild-type GSK3α and -β genes are replaced with mutant forms (GSK3α/βS21A/S21A/S9A/S9A), which are nonresponsive to insulin. Although insulin failed to promote dephosphorylation and activation of GS in GSK3α/βS21A/S21A/S9A/S9Amice, glycogen content in different muscles from these mice was similar compared with wild-type mice. Basal and epinephrine-stimulated activity of muscle glycogen phosphorylase was comparable between wild-type and GSK3 knockin mice. Incubation of isolated soleus muscle in Krebs buffer containing 5.5 mM glucose in the presence or absence of insulin revealed that the levels of G-6- P, the rate of [14C]glucose incorporation into glycogen, and an increase in total glycogen content were similar between wild-type and GSK3 knockin mice. Injection of glucose containing 2-deoxy-[3H]glucose and [14C]glucose also resulted in similar rates of muscle glucose uptake and glycogen synthesis in vivo between wild-type and GSK3 knockin mice. These results suggest that insulin-mediated inhibition of GSK3 is not a rate-limiting step in muscle glycogen synthesis in mice. This suggests that allosteric regulation of GS by G-6- P may play a key role in insulin-stimulated muscle glycogen synthesis in vivo.

Insulin-Sensitive Phospholipid Signaling Systems and Glucose Transport. Update II

2001 ◽  
Vol 226 (4) ◽  
pp. 283-295 ◽  
Author(s):  
Robert V. Farese
Keyword(s):  
Protein Kinase ◽  
Glucose Metabolism ◽  
Glucose Transport ◽  
Intracellular Signaling ◽  
Glucose Transporter ◽  
De Novo ◽  
Glycogen Synthase ◽  
Glycogen Synthesis ◽  
Cell Types ◽  
Biologically Active

Insulin provokes rapid changes in phospholipid metabolism and thereby generates biologically active lipids that serve as intracellular signaling factors that regulate glucose transport and glycogen synthesis. These changes include: (i) activation of phosphatidylinositol 3-kinase (PI3K) and production of PIP3; (ii) PIP3-dependent activation of atypical protein kinase Cs (PKCs); (iii) PIP3-dependent activation of PKB; (iv) PI3K-dependent activation of phospholipase D and hydrolysis of phosphatidyicholine with subsequent increases in phosphatidic acid (PA) and diacyiglycerol (DAG); (v) PI3K-independent activation of glycerol-3-phosphate acylytansferase and increases in de novo synthesis of PA and DAG; and (vi) activation of DAG-sensitive PKCs. Recent findings suggest that atypical PKCs and PKB serve as important positive regulators of insulin-stimulated glucose metabolism, whereas mechanisms that result in the activation of DAG-sensitive PKCs serve mainly as negative regulators of insulin signaling through PI3K. Atypical PKCs and PKB are rapidly activated by insulin in adipocytes, liver, skeletal muscles, and other cell types by a mechanism requiring PI3K and its downstream effector, 3-phosphoinositide-dependent protein kinase-1 (PDK-1), which, in conjunction with PIP3, phosphorylates critical threonine residues in the activation loops of atypical PKCs and PKB. PIP3 also promotes increases in autophosphorylation and allosteric activation of atypical PKCs. Atypical PKCs and perhaps PKB appear to be required for insulin-induced translocation of the GLUT 4 glucose transporter to the plasma membrane and subsequent glucose transport. PKB also appears to be the major regulator of glycogen synthase. Together, atypical PKCs and PKB serve as a potent, integrated PI3K/PDK-1-directed signaling system that is used by insulin to regulate glucose metabolism.

Multiple metabolic effects of CGRP in conscious rats: role of glycogen synthase and phosphorylase

1993 ◽  
Vol 264 (1) ◽  
pp. E1-E10 ◽  
Author(s):  
L. Rossetti ◽  
S. Farrace ◽  
S. B. Choi ◽  
A. Giaccari ◽  
L. Sloan ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Glycogen Synthase ◽  
Hepatic Glucose Production ◽  
Glycogen Synthesis ◽  
Plasma Concentrations ◽  
Glucose Production ◽  
Glucose Disposal ◽  
Hepatic Glucose ◽  
Intracellular Glucose

Calcitonin gene-related peptide (CGRP) is a neuropeptide that is released at the neuromuscular junction in response to nerve excitation. To examine the relationship between plasma CGRP concentration and intracellular glucose metabolism in conscious rats, we performed insulin (22 pmol.kg-1.min-1) clamp studies combined with the infusion of 0, 20, 50, 100, 200, and 500 pmol.kg-1.min-1 CGRP (plasma concentrations ranging from 2 x 10(-11) to 5 x 10(-9) M). CGRP antagonized insulin's suppression of hepatic glucose production at plasma concentrations (approximately 10(-10) M) that are only two- to fivefold its basal portal concentration. Insulin-mediated glucose disposal was decreased by 20-32% when CGRP was infused at 50 pmol.kg-1.min-1 (plasma concentration 3 x 10(-10) M) or more. The impairment in insulin-stimulated glycogen synthesis in skeletal muscle accounted for all of the CGRP-induced decrease in glucose disposal, while whole body glycolysis was increased despite the reduction in total glucose uptake. The muscle glucose 6-phosphate concentration progressively increased during the CGRP infusions. CGRP inhibited insulin-stimulated glycogen synthase in skeletal muscle with a 50% effective dose of 1.9 +/- 0.36 x 10(-10) M. This effect on glycogen synthase was due to a reduction in enzyme affinity for UDP-glucose, with no changes in the maximal velocity. In vitro CGRP stimulated both hepatic and skeletal muscle adenylate cyclase in a dose-dependent manner. These data suggest that 1) CGRP is a potent antagonist of insulin at the level of muscle glycogen synthesis and hepatic glucose production; 2) inhibition of glycogen synthase is its major biochemical action in skeletal muscle; and 3) these effects are present at concentrations of the peptide that may be in the physiological range for portal vein and skeletal muscle. These data underscore the potential role of CGRP in the physiological modulation of intracellular glucose metabolism.

scholarly journals STUDIES OF THE ULTRASTRUCTURE AND RIBOSOMAL ARRANGEMENTS OF THE PLEUROPNEUMONIA-LIKE ORGANISM A5969

1965 ◽  
Vol 25 (1) ◽  
pp. 139-150 ◽  
Author(s):  
Jack Maniloff ◽  
Harold J. Morowitz ◽  
Russell J. Barrnett
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Chemical Analysis ◽  
Thin Section ◽  
Differential Centrifugation ◽  
Thin Section Electron Microscopy ◽  
Section Electron ◽  
Section Electron Microscopy ◽  
Cellular Organelles

Thin-section electron microscopy, together with isolation of cellular organelles by differential centrifugation and chemical analysis, has been used to investigate the ultrastructure of the avian pleuropneumonia-like organism A5969. Each cell (approximate diameter 5500 A) was surrounded by a 150 A plasma membrane. In the center of the cell was an unbounded area, granular in appearance and containing the cell's DNA. The periphery of the cell contained granules of several different sizes and densities. The most dense particles (150 A) corresponded to the 78S ribosomes. These particles exhibited two predominant arrangements: (a) sometimes they showed cubic packing; (b) most arrays, however, were consistent with cylindrical arrangements of approximately 50 particles. Bundles of up to 18 arrays were observed. Structured blebs have been found protruding from the surface of log phase cells.

Metabolic fate of lactate after vigorous activity in the leopard frog, Rana pipiens

1992 ◽  
Vol 262 (2) ◽  
pp. R245-R254 ◽  
Author(s):  
P. A. Fournier ◽  
H. Guderley
Keyword(s):  
Muscle Glycogen ◽  
Glycogen Synthesis ◽  
Physiological Saline ◽  
Frog Muscle ◽  
Leopard Frog ◽  
Strenuous Exercise ◽  
Regulatory Enzymes ◽  
Time Courses

Although the ability of isolated frog muscle to synthesize glycogen from lactate has long been known, it has never been demonstrated that this metabolic activity occurs in the intact frog. Our results clearly indicate that lactate glycogenesis in frog muscle occurs to a significant extent in vivo. During recovery from strenuous exercise, most of the lactate accumulated by frogs seems to be recycled into muscle glycogen because the lactate that disappears during recovery could account nearly stoichiometrically for the glycogen that accumulates in muscle. Furthermore, the decrease in body lactate and the increase in muscle glycogen follow corresponding time courses, suggesting a precursor-product relationship between lactate and glycogen. During recovery from intense exercise, hepatectomized and normal frogs have nearly identical extents of lactate elimination and glycogen synthesis. This suggests that muscle is the main tissue responsible for the recycling of lactate into muscle glycogen and that liver plays a negligible role in lactate disposal. The negligible hepatic contribution to lactate recycling results in part from the liver's incapacity to produce glucose from lactate. In support of this proposition, we show that frog liver perfused in vitro is unable to incorporate any detectable labeled lactate into glucose despite its excellent physiological integrity. Changes in dietary status, training state, season at which the experiments were done, exercise status, and composition of the perfusion media (pH, hormonal composition, physiological saline vs. culture medium) did not give rise to lactate gluconeogenesis. Because frog liver contains all the regulatory enzymes of the gluconeogenic pathway, its inability to synthesize glucose from lactate is not due to an absence of pyruvate carboxylase. A limited ability for lactate uptake may explain why frog liver cannot produce glucose from lactate.

scholarly journals Antagonistic Controls of Autophagy and Glycogen Accumulation by Snf1p, the Yeast Homolog of AMP-Activated Protein Kinase, and the Cyclin-Dependent Kinase Pho85p

2001 ◽  
Vol 21 (17) ◽  
pp. 5742-5752 ◽  
Author(s):  
Zhong Wang ◽  
Wayne A. Wilson ◽  
Marie A. Fujino ◽  
Peter J. Roach
Keyword(s):  
Protein Kinase ◽  
Stationary Phase ◽  
Glycogen Synthase ◽  
Glycogen Synthesis ◽  
Glycogen Storage ◽  
Wild Type ◽  
Gene Encoding ◽  
Glycogen Accumulation ◽  
Yeast Homolog ◽  
Glycogen Stores

ABSTRACT In the yeast Saccharomyces cerevisiae, glycogen is accumulated as a carbohydrate reserve when cells are deprived of nutrients. Yeast mutated in SNF1, a gene encoding a protein kinase required for glucose derepression, has diminished glycogen accumulation and concomitant inactivation of glycogen synthase. Restoration of synthesis in an snf1 strain results only in transient glycogen accumulation, implying the existence of otherSNF1-dependent controls of glycogen storage. A genetic screen revealed that two genes involved in autophagy, APG1and APG13, may be regulated by SNF1. Increased autophagic activity was observed in wild-type cells entering the stationary phase, but this induction was impaired in ansnf1 strain. Mutants defective for autophagy were able to synthesize glycogen upon approaching the stationary phase, but were unable to maintain their glycogen stores, because subsequent synthesis was impaired and degradation by phosphorylase, Gph1p, was enhanced. Thus, deletion of GPH1 partially reversed the loss of glycogen accumulation in autophagy mutants. Loss of the vacuolar glucosidase, SGA1, also protected glycogen stores, but only very late in the stationary phase. Gph1p and Sga1p may therefore degrade physically distinct pools of glycogen. Pho85p is a cyclin-dependent protein kinase that antagonizes SNF1control of glycogen synthesis. Induction of autophagy inpho85 mutants entering the stationary phase was exaggerated compared to the level in wild-type cells, but was blocked in apg1 pho85 mutants. We propose that Snf1p and Pho85p are, respectively, positive and negative regulators of autophagy, probably via Apg1 and/or Apg13. Defective glycogen storage in snf1cells can be attributed to both defective synthesis upon entry into stationary phase and impaired maintenance of glycogen levels caused by the lack of autophagy.

Sign in / Sign up

Export Citation Format

Share Document