scholarly journals Effect of denervation on the expression of glycogen phosphorylase in mouse skeletal muscle

1990 ◽  
Vol 272 (1) ◽  
pp. 231-237 ◽  
Author(s):  
D M Leyland ◽  
P C Turner ◽  
R J Beynon
Keyword(s):  
Skeletal Muscle ◽  
Glycogen Phosphorylase ◽  
Specific Activity ◽  
Muscular Atrophy ◽  
Cdna Probe ◽  
Rapid Decline ◽  
Mrna Levels ◽  
Control Muscle ◽  
Muscle Phosphorylase

After sciatectomy of the left hind-limb of C57BL/J mice, a denervation-induced muscular atrophy ensued and was accompanied by a decrease in the specific activity of glycogen phosphorylase to approx. 25% of control values. The cofactor of phosphorylase, pyridoxal 5′-phosphate, was used as a specific label in the determination of the degradation rate of the enzyme following nerve section. After a delay of 3-4 days, phosphorylase was degraded approx, twice as rapidly in the denervated gastrocnemius (0.20 day-1) as in the control muscle (0.12 day-1). The effect of denervation on phosphorylase mRNA was measured by quantitative Northern-blot analysis using a rat skeletal-muscle phosphorylase cDNA probe. After an initial rapid decline, phosphorylase mRNA levels stabilized in denervated muscle at 50% of the value measured in the contralateral control muscle.

scholarly journals Neural regulation of the formation of skeletal muscle phosphorylase kinase holoenzyme in adult and developing rat muscle

1997 ◽  
Vol 325 (3) ◽  
pp. 793-800 ◽  
Author(s):  
Dean C. NG ◽  
Richard C. CARLSEN ◽  
Donal A. WALSH
Keyword(s):  
Skeletal Muscle ◽  
Muscle Development ◽  
Rapid Decline ◽  
Phosphorylase Kinase ◽  
Β Subunit ◽  
Subunit Protein ◽  
Subunit Mrna ◽  
Γ Subunit ◽  
Muscle Phosphorylase ◽  
Denervated Muscles

Neural influences on the co-ordination of expression of the multiple subunits of skeletal muscle phosphorylase kinase and their assembly to form the holoenzyme complex, α4β4γ4δ4, have been examined during denervation and re-innervation of adult skeletal muscle and during neonatal muscle development. Denervation of the tibialis anterior and extensor digitorum longus muscles of the rat hindlimb was associated with a rapid decline in the mRNA for the γ subunit, and an abrupt decrease in γ-subunit protein. The levels of the α- and β-subunit proteins in the denervated muscles also declined rapidly, their time course of reduction being similar to that for the γ-subunit protein, but they did not decrease to the same extent. In contrast with the rapid decline in γ-subunit mRNA upon denervation, α- and β-subunit mRNAs stayed at control innervated levels for approx. 8–10 days, but then decreased rapidly. Their decline coincided very closely with the onset of re-innervation. Re-innervation of the denervated muscles, which occurs rapidly and uniformly after the sciatic nerve crush injury, produced an eventual slow and prolonged recovery of the mRNA for all three subunits and parallel increases in each of the subunit proteins. A similar co-ordinated increase of both subunit mRNA and subunit proteins of the phosphorylase kinase holoenzyme was observed during neonatal muscle development, during the period when the muscles were attaining their adult pattern of motor activity. The phosphorylase kinase holoenzyme remains in a non-activated form during all of these physiological changes, as is compatible with the presence of the full complement of the regulatory subunits. These data are consistent with a model whereby the transcriptional and translational expression of phosphorylase kinase γ subunit occurs only with concomitant expression of the α and β subunits. This would ensure that free and unregulated, activated γ subunit alone, which would give rise to unregulated glycogenolysis, is not produced. The data also suggest that control of phosphorylase kinase subunit expression and the formation of the holoenzyme in skeletal muscle is provided by the motor nerve, probably through imposed levels or patterns of muscle activity.

scholarly journals Adenovirus-mediated delivery into myocytes of muscle glycogen phosphorylase, the enzyme deficient in patients with glycogen-storage disease type V

1994 ◽  
Vol 304 (3) ◽  
pp. 1009-1014 ◽  
Author(s):  
S Baqué ◽  
C B Newgard ◽  
R D Gerard ◽  
J J Guinovart ◽  
A M Gómez-Foix
Keyword(s):  
Muscle Glycogen ◽  
Glycogen Phosphorylase ◽  
Recombinant Adenovirus ◽  
Genetic Diseases ◽  
C2c12 Myoblast ◽  
Mrna Levels ◽  
Phosphorylase Activity ◽  
Muscle Phosphorylase ◽  
The Impact

The feasibility of using adenovirus as a vector for the introduction of glycogen phosphorylase activity into myocytes has been examined. We used the C2C12 myoblast cell line to assay the impact of phosphorylase gene transfer on myocyte glycogen metabolism and to reproduce in vitro the two strategies proposed for the treatment of muscle genetic diseases, myoblast transplantation and direct DNA delivery. In this study, a recombinant adenovirus containing the muscle glycogen phosphorylase cDNA transcribed from the cytomegalovirus promoter (AdCMV-MGP) was used to transduce both differentiating myoblasts and nondividing mature myotube cells. Muscle glycogen phosphorylase mRNA levels and total phosphorylase activity were increased in both cell types after viral treatment although more efficiently in the differentiated myotubes. The increase in phosphorylase activity was transient (15 days) in myoblasts whereas in myotubes higher levels of phosphorylase gene expression and activity were reached, which remained above control levels for the duration of the study (20 days). The introduction of muscle phosphorylase into myotubes enhanced their glycogenolytic capacity. AdCMV MGP-transduced myotubes had lower glycogen levels under basal conditions. In addition, these engineered cells showed more extensive glycogenolysis in response to both adrenaline, which stimulates glycogen phosphorylase phosphorylation, and carbonyl cyanide m-chlorophenylhydrazone, a metabolic uncoupler. In conclusion, transfer of the muscle glycogen phosphorylase cDNA into myotubes confers an enhanced and regulatable glycogenolytic capacity. Thus this system might be useful for delivery of muscle glycogen phosphorylase and restoration of glycogenolysis in muscle cells from patients with muscle phosphorylase deficiency (McArdle's disease).

Cell surface and Golgi pools of beta-1,4-galactosyltransferase are differentially regulated during embryonal carcinoma cell differentiation

1989 ◽  
Vol 9 (6) ◽  
pp. 2370-2377
Author(s):  
L C Lopez ◽  
C M Maillet ◽  
K Oleszkowicz ◽  
B D Shur
Keyword(s):  
Cell Differentiation ◽  
Golgi Apparatus ◽  
Cell Surface ◽  
Embryonal Carcinoma ◽  
Specific Activity ◽  
Cdna Probe ◽  
Subcellular Fractionation ◽  
Specific Marker ◽  
Embryonal Carcinoma Cell ◽  
Mrna Levels

beta-1,4-Galactosyltransferase (GalTase) has two functionally distinct subcellular distributions. In the Golgi apparatus, GalTase participates in the glycosylation of secretory and membrane-bound glycoproteins, whereas on the cell surface it mediates specific aspects of intercellular adhesion. For this study, a murine GalTase clone was obtained by screening a lambda gt10 cDNA library made from F9 embryonal carcinoma cells with a heterologous bovine GalTase cDNA probe. The murine GalTase cDNA probe was used in conjunction with assays of GalTase activity to investigate the expression and distribution of GalTase during differentiation of F9 stem cells into secretory endodermal epithelium. During the initial phase of F9 cell differentiation, GalTase mRNA levels remained relatively constant; however, as differentiation progressed, as assayed by expression of the differentiation-specific marker laminin B1, GalTase mRNA levels and enzyme activity rose dramatically. Furthermore, subcellular fractionation of these cells showed that the increased GalTase levels were specifically associated with the Golgi apparatus, whereas GalTase specific activity on the plasma membrane remained constant. These results show that levels of cell surface and Golgi GalTase change relative to one another during F9 cell differentiation and suggest that these functionally distinct pools of GalTase are independently and differentially regulated.

Fish muscle glycogen phosphorylase

1968 ◽  
Vol 46 (5) ◽  
pp. 423-432 ◽  
Author(s):  
M. Yamamoto
Keyword(s):  
Skeletal Muscle ◽  
Glycogen Phosphorylase ◽  
Active Form ◽  
Maximal Activity ◽  
Fish Muscle ◽  
Salmo Gairdneri ◽  
Rabbit Muscle ◽  
Muscle Phosphorylase

Glycogen phosphorylase b was purified 70- to 90-fold from skeletal muscle of rainbow trout (Salmo gairdneri). The purified enzyme exhibited maximal activity near pH 6.8 at 37°. Of several 5′-nucleotides tested, only 5′-AMP caused stimulation of phosphorylase b. The Km value for glucose-1-phosphate was 10–15 mM, and for 5′-AMP, 0.2–0.4 mM. Glucose (25 mM) and ATP (5 mM) were both inhibitory, but glucose-6-phosphate (5 mM) had no effect. Inactive trout muscle phosphorylase was converted to the active form in vivo by subjecting a fish to physical exercise. The conversion of fish muscle phosphorylase b to a was also catalyzed in vitro with purified rabbit muscle phosphorylase b kinase in the presence of ATP and Mg++. Evidence is presented to indicate the presence of phosphorylase b kinase and phosphorylase phosphatase in trout skeletal muscle.

The sulfhydryl groups of muscle phosphorylase. V. The reactive sulfhydryl peptides

1970 ◽  
Vol 48 (7) ◽  
pp. 763-775 ◽  
Author(s):  
C. G. Zarkadas ◽  
L. B. Smillie ◽  
N. B. Madsen
Keyword(s):  
Skeletal Muscle ◽  
Molecular Weight ◽  
Muscle Glycogen ◽  
Glycogen Phosphorylase ◽  
Chromatographic Purification ◽  
Model Compounds ◽  
Rate Of Reaction ◽  
Peptic Digestion ◽  
Muscle Phosphorylase ◽  
Column Chromatographic

Skeletal muscle glycogen phosphorylase a and b have previously been shown to consist of four and two subunits, respectively, each having a molecular weight of 92 500. Our studies have indicated that there is a minimum of eight and a maximum of nine unique cysteine sequences in the enzyme providing additional evidence that the subunits are identical. From the known sequences of the nine sulfhydryl peptides it was possible to isolate and characterize the reactive sulfhydryl peptides of phosphorylase. Careful column chromatographic purification of alkylated peptides derived from peptic digestion showed that there are two cysteine residues per monomer of phosphorylase b whose rate of reaction with iodoacetamide approaches that of model compounds and whose alkylation does not affect significantly the enzymic properties of the protein. These two cysteines have been identified in sequences corresponding to numbers 2 and 5 previously elucidated. These sequences have been extended in the present work and may now be written as: No. 2 Asn–Gln–Lys–Ile–CMC–Gly–Gly–Try–Gln–Ser, and No. 5 Gly–CMC–Arg–Asp–Pro–Val–Arg–Thr–Asn–Phe.

scholarly journals Developmental regulation of the alpha-mannosidase gene in Dictyostelium discoideum: control is at the level of transcription and is affected by cell density.

1991 ◽  
Vol 11 (6) ◽  
pp. 3339-3347 ◽  
Author(s):  
J Schatzle ◽  
A Rathi ◽  
M Clarke ◽  
J A Cardelli
Keyword(s):  
Cell Density ◽  
Dictyostelium Discoideum ◽  
High Cell Density ◽  
Specific Activity ◽  
Developmental Regulation ◽  
Cdna Probe ◽  
Mrna Levels ◽  
High Cell ◽  
Enzyme Specific Activity ◽  
Negative Mutant

In Dictyostelium discoideum, there is a group of genes that are expressed following starvation and when exponentially growing cells reach high densities. We have examined the expression of one of these genes, alpha-mannosidase. Using an alpha-mannosidase cDNA probe in Northern (RNA) blot analysis, we have shown that the previously observed increase in alpha-mannosidase enzyme-specific activity during development is due to an increase in the levels of alpha-mannosidase mRNA. mRNA levels reach a maximum by 8 h of development and then begin to decline by 14 to 22 h. Using nuclear run-on analysis, we have found that this gene is regulated at the level of transcription. We also examined the effects of cell-cell contacts, cyclic AMP levels, and protein synthesis on expression of this gene and found that they were not critical in regulating its expression. However, cell density did play a major role in the expression of alpha-mannosidase. High cell density or the presence of buffer conditioned by high-density cells was sufficient to induce expression of alpha-mannosidase, indicating that this is one of the prestarvation response genes. Finally, the alpha-mannosidase gene was not expressed in aggregation-negative mutant strain HMW 404.

scholarly journals Cell surface and Golgi pools of beta-1,4-galactosyltransferase are differentially regulated during embryonal carcinoma cell differentiation.

1989 ◽  
Vol 9 (6) ◽  
pp. 2370-2377 ◽  
Author(s):  
L C Lopez ◽  
C M Maillet ◽  
K Oleszkowicz ◽  
B D Shur
Keyword(s):  
Cell Differentiation ◽  
Golgi Apparatus ◽  
Cell Surface ◽  
Embryonal Carcinoma ◽  
Specific Activity ◽  
Cdna Probe ◽  
Subcellular Fractionation ◽  
Specific Marker ◽  
Embryonal Carcinoma Cell ◽  
Mrna Levels

beta-1,4-Galactosyltransferase (GalTase) has two functionally distinct subcellular distributions. In the Golgi apparatus, GalTase participates in the glycosylation of secretory and membrane-bound glycoproteins, whereas on the cell surface it mediates specific aspects of intercellular adhesion. For this study, a murine GalTase clone was obtained by screening a lambda gt10 cDNA library made from F9 embryonal carcinoma cells with a heterologous bovine GalTase cDNA probe. The murine GalTase cDNA probe was used in conjunction with assays of GalTase activity to investigate the expression and distribution of GalTase during differentiation of F9 stem cells into secretory endodermal epithelium. During the initial phase of F9 cell differentiation, GalTase mRNA levels remained relatively constant; however, as differentiation progressed, as assayed by expression of the differentiation-specific marker laminin B1, GalTase mRNA levels and enzyme activity rose dramatically. Furthermore, subcellular fractionation of these cells showed that the increased GalTase levels were specifically associated with the Golgi apparatus, whereas GalTase specific activity on the plasma membrane remained constant. These results show that levels of cell surface and Golgi GalTase change relative to one another during F9 cell differentiation and suggest that these functionally distinct pools of GalTase are independently and differentially regulated.

scholarly journals Tissue-specific activity of lipoprotein lipase in skeletal muscle regulates the expression of uncoupling protein 3 in transgenic mouse models

2001 ◽  
Vol 355 (3) ◽  
pp. 647-652 ◽  
Author(s):  
Dagmar KRATKY ◽  
Juliane G. STRAUSS ◽  
Rudolf ZECHNER
Keyword(s):  
Skeletal Muscle ◽  
Lipoprotein Lipase ◽  
Specific Activity ◽  
Uncoupling Protein ◽  
Large Body ◽  
Mrna Levels ◽  
Expression Levels ◽  
Uncoupling Protein 3 ◽  
Triacylglycerol Hydrolysis ◽  
Brown Adipose

Uncoupling protein (UCP)-2 and UCP-3 are two recently discovered proteins similar to UCP-1, which regulates thermogenesis in brown adipose tissue (BAT). Whereas UCP-1 expression is restricted to BAT, UCP-2 is widely expressed. UCP-3 is found mainly in skeletal muscle and BAT. A large body of evidence exists that the expression of UCP-2 and UCP-3 in skeletal muscle of mice is regulated by feeding/fasting, and some studies have suggested that this effect might be caused by the changing concentration of plasma non-esterified fatty acids (NEFAs). In an attempt to determine whether the increased import of triacylglycerol-derived NEFAs can also affect UCP expression, we determined the mRNA levels of UCP-1, UCP-2 and UCP-3 in BAT and muscle of induced mutant mouse lines that overexpressed or lacked lipoprotein lipase (LPL) in these tissues. The expression levels of UCP-1 and UCP-2 in BAT and in skeletal and cardiac muscle respectively were not affected by variations in tissue LPL activities. In contrast, UCP-3 mRNA levels were induced 3.4-fold in mice with high levels of LPL in skeletal muscle, and down-regulated in mice that lacked LPL in skeletal muscle. The presence or absence of LPL in BAT had no effect on UCP-3 expression levels. The response of UCP-3 mRNA expression to variations in LPL activity in skeletal muscle was independent of the feeding status or of plasma NEFA concentrations. These findings indicated that NEFAs as lipolytic products of LPL-mediated triacylglycerol hydrolysis markedly affect UCP-3 expression and that increased LPL activities occurring during fasting in skeletal muscle contribute to the induction of UCP-3 expression by promoting the increased uptake of NEFAs. In addition, our results demonstrate that UCP-2 and UCP-3 are differentially regulated in response to LPL-mediated NEFA uptake in skeletal muscle of mice.

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