Regulation of the dephosphorylation of glycogen phosphorylase a and synthase b by glucose and caffeine in isolated hepatocytes

1981 ◽  
Vol 59 (6) ◽  
pp. 387-395 ◽  
Author(s):  
Peter J. Kasvinsky ◽  
Robert J. Fletterick ◽  
Neil B. Madsen
Keyword(s):  
Glycogen Phosphorylase ◽  
Glycogen Synthase ◽  
Physiological Role ◽  
Glycogen Metabolism ◽  
Isolated Hepatocytes ◽  
Effector Site ◽  
Synergistic Regulation ◽  
Hepatic Regulation

Synergistic regulation of glycogen phosphorylase a by the competitive inhibitors glucose and caffeine in vitro indicates a possible physiological role for the negative effector site which binds caffeine (nucleoside site). In intact viable hepatocytes glucose promotes the phosphorylase a to b conversion by phosphorylase phosphatase. This conversion is considered to be a necessary prelude to the activation of glycogen synthase by phosphatase and of importance in hepatic regulation of glucose homeostasis. The effects of glucose and(or) caffeine on the conversion of phosphorylase a to b and synthase b to a were studied. Assays of phosphorylase a were used which limited synergistic inhibition (in the assay) by these ligands. Such an approach is necessary to achieve an accurate measure of phosphatase activity in the viable hepatocyte when the combination of ligands is used. The data indicate that in the presence of caffeine and glucose together, the rate of loss of phosphorylase a is significantly increased (1.7-fold) over that in the presence of glucose alone. Phosphorylase phosphatase is activated. The sequential activation of glycogen synthase was also accelerated in the presence of both ligands. The results are consistent with an in vivo function for the nucleoside site, similar to that of glucose. A controlling role for phosphorylase in the regulation of glycogen metabolism by glucose is supported. Although the existence and nature of an intracellular effector is as yet unknown, crystallographic analyses of phosphorylase a crystals soaked in perchloric acid extracts of liver demonstrate that the negative effector site binds a natural metabolite.

scholarly journals Diverse effects of two allosteric inhibitors on the phosphorylation state of glycogen phosphorylase in hepatocytes

2002 ◽  
Vol 368 (1) ◽  
pp. 309-316 ◽  
Author(s):  
Theodore LATSIS ◽  
Birgitte ANDERSEN ◽  
Loranne AGIUS
Keyword(s):  
Glycogen Phosphorylase ◽  
Potent Inhibitor ◽  
Glycogen Synthase ◽  
Glycogen Synthesis ◽  
Glycogen Metabolism ◽  
Small Degree ◽  
Allosteric Inhibitors ◽  
Phosphorylation State ◽  
Stimulation Of

Two distinct allosteric inhibitors of glycogen phosphorylase, 1,4-dideoxy-1,4-imino-d-arabinitol (DAB) and CP-91149 (an indole-2-carboxamide), were investigated for their effects on the phosphorylation state of the enzyme in hepatocytes in vitro. CP-91149 induced inactivation (dephosphorylation) of phosphorylase in the absence of hormones and partially counteracted the phosphorylation caused by glucagon. Inhibition of glycogenolysis by CP-91149 can be explained by dephosphorylation of phosphorylase a. This was associated with activation of glycogen synthase and stimulation of glycogen synthesis. DAB, in contrast, induced a small degree of phosphorylation of phosphorylase. This was associated with inactivation of glycogen synthase and inhibition of glycogen synthesis. Despite causing phosphorylation (activation) of phosphorylase, DAB is a very potent inhibitor of glycogenolysis in both the absence and presence of glucagon. This is explained by allosteric inhibition of phosphorylase a, which overrides the increase in activation state. In conclusion, two potent phosphorylase inhibitors exert different effects on glycogen metabolism in intact hepatocytes as a result of opposite effects on the phosphorylation state of both phosphorylase and glycogen synthase.

scholarly journals Identification of Domains Responsible for Ubiquitin-Dependent Degradation of dMyc by Glycogen Synthase Kinase 3β and Casein Kinase 1 Kinases

2009 ◽  
Vol 29 (12) ◽  
pp. 3424-3434 ◽  
Author(s):  
Margherita Galletti ◽  
Sara Riccardo ◽  
Federica Parisi ◽  
Carlina Lora ◽  
Mahesh Kumar Saqcena ◽  
...  
Keyword(s):  
Compound Eye ◽  
Glycogen Synthase ◽  
Physiological Role ◽  
Casein Kinase ◽  
Glycogen Synthase Kinase ◽  
Glycogen Synthase Kinase 3Β ◽  
Casein Kinase 1 ◽  
The One

ABSTRACT In the present study, we report that ubiquitin-mediated degradation of dMyc, the Drosophila homologue of the human c-myc proto-oncogene, is regulated in vitro and in vivo by members of the casein kinase 1 (CK1) family and by glycogen synthase kinase 3β (GSK3β). Using Drosophila S2 cells, we demonstrate that CK1α promotes dMyc ubiquitination and degradation with a mechanism similar to the one mediated by GSK3β in vertebrates. Mutation of ck1α or -ε or sgg/gsk3β in Drosophila wing imaginal discs results in the accumulation of dMyc protein, suggesting a physiological role for these kinases in vivo. Analysis of the dMyc amino acid sequence reveals the presence of conserved domains containing potential phosphorylation sites for mitogen kinases, GSK3β, and members of the CK1 family. We demonstrate that mutations of specific residues within these phosphorylation domains regulate dMyc protein stability and confer resistance to degradation by CK1α and GSK3β kinases. Expression of the dMyc mutants in the compound eye of the adult fly results in a visible defect that is attributed to the effect of dMyc on growth, cell death, and inhibition of ommatidial differentiation.

Prolonged exposure to halothane and associated changes in carbohydrate metabolism in rat muscles in vivo

1998 ◽  
Vol 84 (4) ◽  
pp. 1470-1474 ◽  
Author(s):  
Luis D. M. C.-B. Ferreira ◽  
T. Norman Palmer ◽  
Paul A. Fournier
Keyword(s):  
Skeletal Muscles ◽  
Glycogen Phosphorylase ◽  
Prolonged Exposure ◽  
Glycogen Metabolism ◽  
Activity Ratios ◽  
Fructose 1,6 Bisphosphate ◽  
Glycogen Breakdown ◽  
Stimulation Of

Halothane, an anesthetic presently used in animal experimentation, is reported to stimulate glycogen breakdown in isolated preparations of rat skeletal muscles, suggesting that it may not be a suitable anesthetic for the study of glycogen metabolism in rats in vivo. The purpose of this study was to establish whether prolonged exposure to halothane in rats in vivo is associated with accelerated glycogenolysis. Exposure of rats to halothane for up to 1 h was not accompanied by either any change in the levels of glycogen or increase in activity ratios of glycogen phosphorylase in muscles, irrespective of their fiber compositions. In marked contrast, the levels of lactate, inorganic phosphate, glucose 1-phosphate, glucose 6-phosphate, fructose 1,6-bisphosphate, and fructose 2,6-bisphosphate changed progressively during anesthesia. Accordingly, the interpretation of muscle metabolite levels must be performed with caution in experiments involving prolonged exposure to halothane. Overall, our findings indicate that the reported halothane-mediated stimulation of glycogen breakdown in vitro is likely to be an artifact and that halothane is a suitable anesthetic for experiments concerned with glycogen metabolism in rats.

scholarly journals The stimulation of glycogenolysis in isolated hepatocytes by opioid peptides

1986 ◽  
Vol 238 (2) ◽  
pp. 531-535 ◽  
Author(s):  
R P Leach ◽  
M A Titheradge
Keyword(s):  
Cyclic Amp ◽  
Opioid Peptides ◽  
Glycogen Phosphorylase ◽  
Glycogen Synthase ◽  
Glycogen Metabolism ◽  
Isolated Hepatocytes ◽  
Dependent Protein ◽  
Simultaneous Decrease ◽  
Glycogen Phosphorylase Activity ◽  
Stimulation Of

The opioid peptides [Leu]enkephalin and dynorphin-(1-13) were shown to enhance glycogen breakdown when added directly to hepatocytes. This was the result of a concerted effect on the enzymes of glycogen metabolism, with a stimulation of glycogen phosphorylase activity and a simultaneous decrease in glycogen synthase I activity. The latter only became significant when the enzyme was activated by incubating the cells in presence of 20 mM- or 40 mM-glucose. The effect of the opioid peptides was independent of an increase in cyclic AMP or any change in the activity ratio of the cyclic AMP-dependent protein kinase and was abolished by depleting the cells of Ca2+. Both [Leu]enkephalin and dynorphin-(1-13) produced a significant decrease in cyclic AMP formation, suggesting that in liver, as in neuronal tissue, they may act by inhibiting adenylate cyclase activity.

scholarly journals Effects of C-1-substituted glucose analogue on the activation states of glycogen synthase and glycogen phosphorylase in rat hepatocytes

1995 ◽  
Vol 311 (3) ◽  
pp. 845-852 ◽  
Author(s):  
M Board ◽  
M Bollen ◽  
W Stalmans ◽  
Y Kim ◽  
G W J Fleet ◽  
...  
Keyword(s):  
Glycogen Phosphorylase ◽  
Kinase Inhibitor ◽  
Glycogen Synthase ◽  
Kinetic Studies ◽  
Rat Hepatocytes ◽  
Glycogen Metabolism ◽  
Isolated Hepatocytes ◽  
Effective Control ◽  
Subsequent Addition ◽  
Glucose Analogue

A series of glucose-analogue inhibitors of glycogen phosphorylase b (GPb) has been designed, synthesized and investigated in crystallographic binding and kinetic studies. The aim is to produce a compound that may exert more effective control over glycogen metabolism than the parent glucose molecule and which could alleviate hyperglycaemia in Type-II diabetes. N-Acetyl-beta-D-glucopyranosylamine (1-GlcNAc) has a Ki for muscle GPb in crude extracts of 30 microM, 367-fold lower than that of beta-D-glucose [Board, Hadwen and Johnson (1995) Eur. J. Biochem. 228, 753-761]. In the current work, the effects of 1-GlcNAc on the activation states of GP and glycogen synthase (GS) in cell-free preparations and in isolated hepatocytes are reported. In gel-filtered extracts of liver, which lack ATP for kinase activity, 1-GlcNAc produced a rapid and time-dependent inactivation of GP with a subsequent activation of GS. Effects of 1-GlcNAc on both enzymes were stronger than those of glucose, with 0.8 mM 1-GlcNAc being equipotent with 50 mM glucose. At 1 mM, 1-GlcNAc enhanced the dephosphorylation of exogenous GPa by liver extracts (600%) and by muscle extracts (75%). This represents an approximately 500-fold improvement on glucose for the liver activity and 40-fold for the muscle activity. In whole hepatocytes, 1-GlcNAc showed an approximately 5-fold enhancement of glucose effects for GP inactivation but failed to elicit activation of GS. Glucose-induced activation of GS in whole hepatocytes was reversed by subsequent addition of 1-GlcNAc. However, when GS activation was achieved via the adenosine analogue and kinase inhibitor, 5′-iodotubercidin (ITU), subsequent addition of 1-GlcNAc allowed continued activation of GS. Phosphorylation of 1-GlcNAc in rat hepatocytes was established using radiolabelled material. The rate of phosphorylation was 1.60 nmol/min per 10(6) cells at 20 mM 1-GlcNAc but was reduced by the presence of 50 microM ITU (0.775 nmol/min per 10(6) cells). It is suggested that the phosphorylated derivative of 1-GlcNAc formed in hepatocytes is 1-GlcNAc 6-phosphate and that the presence of this species is responsible for the failure of 1-GlcNAc to activate GS. The relative importance of the reduction in concentration of GPa versus increased glucose 6-phosphate levels for activation of GS is discussed.

Targeting Tau Hyperphosphorylation via Kinase Inhibition: Strategy to Address Alzheimer's Disease

2020 ◽  
Vol 20 (12) ◽  
pp. 1059-1073 ◽  
Author(s):  
Ahmad Abu Turab Naqvi ◽  
Gulam Mustafa Hasan ◽  
Md. Imtaiyaz Hassan
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Tau Protein ◽  
Glycogen Synthase ◽  
Paired Helical Filaments ◽  
Tau Hyperphosphorylation ◽  
Microtubule Stabilization ◽  
Extracellular Signal

Microtubule-associated protein tau is involved in the tubulin binding leading to microtubule stabilization in neuronal cells which is essential for stabilization of neuron cytoskeleton. The regulation of tau activity is accommodated by several kinases which phosphorylate tau protein on specific sites. In pathological conditions, abnormal activity of tau kinases such as glycogen synthase kinase-3 β (GSK3β), cyclin-dependent kinase 5 (CDK5), c-Jun N-terminal kinases (JNKs), extracellular signal-regulated kinase 1 and 2 (ERK1/2) and microtubule affinity regulating kinase (MARK) lead to tau hyperphosphorylation. Hyperphosphorylation of tau protein leads to aggregation of tau into paired helical filaments like structures which are major constituents of neurofibrillary tangles, a hallmark of Alzheimer’s disease. In this review, we discuss various tau protein kinases and their association with tau hyperphosphorylation. We also discuss various strategies and the advancements made in the area of Alzheimer's disease drug development by designing effective and specific inhibitors for such kinases using traditional in vitro/in vivo methods and state of the art in silico techniques.

scholarly journals GSK-3β in Pancreatic Cancer: Spotlight on 9-ING-41, Its Therapeutic Potential and Immune Modulatory Properties

Biology ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 610
Author(s):  
Robin Park ◽  
Andrew L. Coveler ◽  
Ludimila Cavalcante ◽  
Anwaar Saeed
Keyword(s):  
Pancreatic Cancer ◽  
Therapeutic Potential ◽  
Glycogen Synthase ◽  
Glycogen Synthase Kinase ◽  
Glycogen Synthase Kinase 3 ◽  
In Vivo Models ◽  
Gsk 3Β ◽  
Early Phase Clinical Trials

Glycogen synthase kinase-3 beta is a ubiquitously and constitutively expressed molecule with pleiotropic function. It acts as a protooncogene in the development of several solid tumors including pancreatic cancer through its involvement in various cellular processes including cell proliferation, survival, invasion and metastasis, as well as autophagy. Furthermore, the level of aberrant glycogen synthase kinase-3 beta expression in the nucleus is inversely correlated with tumor differentiation and survival in both in vitro and in vivo models of pancreatic cancer. Small molecule inhibitors of glycogen synthase kinase-3 beta have demonstrated therapeutic potential in pre-clinical models and are currently being evaluated in early phase clinical trials involving pancreatic cancer patients with interim results showing favorable results. Moreover, recent studies support a rationale for the combination of glycogen synthase kinase-3 beta inhibitors with chemotherapy and immunotherapy, warranting the evaluation of novel combination regimens in the future.

scholarly journals Propofol attenuates lung ischemia/reperfusion injury though the involvement of the MALAT1/microRNA-144/GSK3β axis

2021 ◽  
Vol 27 (1) ◽  
Author(s):  
Jian-Ping Zhang ◽  
Wei-Jing Zhang ◽  
Miao Yang ◽  
Hua Fang
Keyword(s):  
Cell Apoptosis ◽  
Ischemia Reperfusion Injury ◽  
Glycogen Synthase ◽  
Ischemia Reperfusion ◽  
Inflammatory Factors ◽  
Therapeutic Effects ◽  
Protective Effects ◽  
Loss Of Function

Abstract Background Propofol, an intravenous anesthetic, was proven to protect against lung ischemia/reperfusion (I/R) injury. However, the detailed mechanism of Propofol in lung I/R injury is still elusive. This study was designed to explore the therapeutic effects of Propofol, both in vivo and in vitro, on lung I/R injury and the underlying mechanisms related to metastasis-associated lung adenocarcinoma transcript 1 (MALAT1)/microRNA-144 (miR-144)/glycogen synthase kinase-3β (GSK3β). Methods C57BL/6 mice were used to establish a lung I/R injury model while pulmonary microvascular endothelial cells (PMVECs) were constructed as hypoxia/reperfusion (H/R) cellular model, both of which were performed with Propofol treatment. Gain- or loss-of-function approaches were subsequently employed, followed by observation of cell apoptosis in lung tissues and evaluation of proliferative and apoptotic capabilities in H/R cells. Meanwhile, the inflammatory factors, autophagosomes, and autophagy-related proteins were measured. Results Our experimental data revealed that Propofol treatment could decrease the elevated expression of MALAT1 following I/R injury or H/R induction, indicating its protection against lung I/R injury. Additionally, overexpressing MALAT1 or GSK3β promoted the activation of autophagosomes, proinflammatory factor release, and cell apoptosis, suggesting that overexpressing MALAT1 or GSK3β may reverse the protective effects of Propofol against lung I/R injury. MALAT1 was identified to negatively regulate miR-144 to upregulate the GSK3β expression. Conclusion Overall, our study demonstrated that Propofol played a protective role in lung I/R injury by suppressing autophagy and decreasing release of inflammatory factors, with the possible involvement of the MALAT1/miR-144/GSK3β axis.

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