Activated protein kinase C α associates with annexin VI from skeletal muscle

We have previously detected a number of protein kinase C (PKC) α-binding proteins in skeletal muscle cytosol by blot overlay assay, and now identify the major, 69 kDa binding protein as annexin VI by immunoblotting and overlay assay of hydroxyapatite chromatography fractions. Annexin VI was also detected in immunoprecipitates of PKC α. Annexin VI and PKC α are both calcium-dependent phospholipid-binding proteins, and detection of the interaction was dependent on the presence of calcium and phosphatidylserine (PS). The association probably involves specific protein-protein interactions rather than mere bridging by lipid molecules: firstly, detection of PKC α-annexin VI complexes by overlay assay was not diminished when PS concentrations were increased over a 10-fold range, while that of other PKC α-binding protein complexes was reduced or abolished; secondly, the presence in the overlay assay of a PKC pseudosubstrate peptide, analogous to a PKC sequence previously found to be involved in PKC binding activity, reduced complex formation; thirdly, we were also able to detect annexin VI interaction with PKC β by overlay of skeletal muscle cytosol, but not with PKC ϴ, the major novel PKC in this tissue, suggesting sequences specific to calcium-dependent PKC isoenzymes are involved. While other annexin isoforms may be PKC substrates or inhibitors, annexin VI phosphorylation by PKC α could not be detected after co-purification, while phosphorylation of subsequently-added histone IIIS was readily observed. Annexin VI is a major skeletal muscle protein and our data are consistent with a role for this isoform in the control of calcium-dependent PKC.

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Reversal of chronic alterations of skeletal muscle protein kinase C from fat-fed rats by BRL-49653

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1997.273.5.e915 ◽

1997 ◽

Vol 273 (5) ◽

pp. E915-E921 ◽

Cited By ~ 12

Author(s):

Carsten Schmitz-Peiffer ◽

Nicholas D. Oakes ◽

Carol L. Browne ◽

Edward W. Kraegen ◽

Trevor J. Biden

Keyword(s):

Insulin Resistance ◽

Skeletal Muscle ◽

Protein Kinase C ◽

Protein Kinase ◽

Muscle Protein ◽

Skeletal Muscle Protein ◽

Insulin Sensitizer ◽

Muscle Insulin Resistance ◽

Skeletal Muscle Insulin Resistance ◽

Kinase C

We have recently shown that the reduction in insulin sensitivity of rats fed a high-fat diet is associated with the translocation of the novel protein kinase Cε(nPKCε) from cytosolic to particulate fractions in red skeletal muscle and also the downregulation of cytosolic nPKCθ. Here we have further investigated the link between insulin resistance and PKC by assessing the effects of the thiazolidinedione insulin-sensitizer BRL-49653 on PKC isoenzymes in muscle. BRL-49653 increased the recovery of nPKC isoenzymes in cytosolic fractions of red muscle from fat-fed rats, reducing their apparent activation and/or downregulation, whereas PKC in control rats was unaffected. Because BRL-49653 also improves insulin-stimulated glucose uptake in fat-fed rats and reduces muscle lipid storage, especially diglyceride content, these results strengthen the association between lipid availability, nPKC activation, and skeletal muscle insulin resistance and support the hypothesis that chronic activation of nPKC isoenzymes is involved in the generation of muscle insulin resistance in fat-fed rats.

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Ca2+-binding proteins from bovine brain including a potent inhibitor of protein kinase C

Biochemical Journal ◽

10.1042/bj2320559 ◽

1985 ◽

Vol 232 (2) ◽

pp. 559-567 ◽

Cited By ~ 66

Author(s):

J R McDonald ◽

M P Walsh

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Large Scale ◽

Binding Proteins ◽

Potent Inhibitor ◽

Binding Protein ◽

Sodium Dodecyl ◽

Bovine Brain ◽

Heat Stable ◽

Kinase C

We have previously described the use of Ca2+-dependent hydrophobic-interaction chromatography to isolate the Ca2+ + phospholipid-dependent protein kinase (protein kinase C) and a novel heat-stable 21 000-Mr Ca2+-binding protein from bovine brain [Walsh, Valentine, Ngai, Carruthers & Hollenberg (1984) Biochem. J. 224, 117-127]. The procedure described for purification of the 21 000-Mr calciprotein to electrophoretic homogeneity has been modified to permit the large-scale isolation of this Ca2+-binding protein, enabling further structural and functional characterization. The 21 000-Mr calciprotein was shown by equilibrium dialysis to bind approx. 1 mol of Ca2+/mol, with apparent Kd approx. 1 microM. The modified large-scale purification procedure revealed three additional, previously unidentified, Ca2+-binding proteins of Mr 17 000, 18 400 and 26 000. The 17 000-Mr and 18 400-Mr Ca2+-binding proteins are heat-stable, whereas the 26 000-Mr Ca2+-binding protein is heat-labile. Use of the transblot/45CaCl2 overlay technique [Maruyama, Mikawa & Ebashi (1984) J. Biochem. (Tokyo) 95, 511-519] suggests that the 18 400-Mr and 21 000-Mr Ca2+-binding proteins are high-affinity Ca2+-binding proteins, whereas the 17 000-Mr Ca2+-binding protein has a relatively low affinity for Ca2+. Consistent with this observation, the 18 400-Mr and 21 000-Mr Ca2+-binding proteins exhibit a Ca2+-dependent mobility shift on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, whereas the 17 000-Mr Ca2+-binding protein does not. The amino acid compositions of the 17 000-Mr, 18 400-Mr and 21 000-Mr Ca2+-binding proteins show some similarities to each other and to calmodulin and other members of the calmodulin superfamily; however, they are clearly distinct and novel calciproteins. In functional terms, none of the 17 000-Mr, 18 400-Mr or 21 000-Mr Ca2+-binding proteins activates either cyclic nucleotide phosphodiesterase or myosin light-chain kinase, both calmodulin-activated enzymes. However, the 17 000-Mr Ca2+-binding protein is a potent inhibitor of protein kinase C. It may therefore serve to regulate the activity of this important enzyme at elevated cytosolic Ca2+ concentrations.

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The Glutamate Agonist Homocysteine Sulfinic Acid Stimulates Glucose Uptake through the Calcium-dependent AMPK-p38 MAPK-Protein Kinase C ζ Pathway in Skeletal Muscle Cells

Journal of Biological Chemistry ◽

10.1074/jbc.m110.149328 ◽

2010 ◽

Vol 286 (9) ◽

pp. 7567-7576 ◽

Cited By ~ 21

Author(s):

Ji Hae Kim ◽

Jung Ok Lee ◽

Soo Kyung Lee ◽

Ji Wook Moon ◽

Ga Young You ◽

...

Keyword(s):

Skeletal Muscle ◽

Protein Kinase C ◽

Protein Kinase ◽

Glucose Uptake ◽

P38 Mapk ◽

Muscle Cells ◽

Skeletal Muscle Cells ◽

Sulfinic Acid ◽

Calcium Dependent ◽

Kinase C

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Defective phosphorylation of a calmodulin-binding protein in cystic-fibrosis submandibular glands

Biochemical Journal ◽

10.1042/bj2630613 ◽

1989 ◽

Vol 263 (2) ◽

pp. 613-616 ◽

Cited By ~ 11

Author(s):

D K Shori ◽

R L Dormer ◽

M C Goodchild ◽

M A McPherson

Keyword(s):

Cystic Fibrosis ◽

Protein Kinase C ◽

Protein Kinase ◽

Binding Proteins ◽

Binding Protein ◽

Submandibular Glands ◽

Calmodulin Binding ◽

Kinase C ◽

Exogenous Protein ◽

Molecular Masses

Calmodulin-binding proteins in fractions purified from human submandibular glands by calmodulin-Sepharose were phosphorylated with [gamma-32P]ATP, in the absence of exogenous protein kinase. The major proteins phosphorylated had molecular masses of 45, 51 and 61 kDa. Phosphorylation was increased by activators of protein kinase C and inhibited by H-7. Phosphorylation of the 61 kDa band was markedly decreased in cystic-fibrosis submandibular glands.

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Ca2+-dependent hydrophobic-interaction chromatography. Isolation of a novel Ca2+-binding protein and protein kinase C from bovine brain

Biochemical Journal ◽

10.1042/bj2240117 ◽

1984 ◽

Vol 224 (1) ◽

pp. 117-127 ◽

Cited By ~ 156

Author(s):

M P Walsh ◽

K A Valentine ◽

P K Ngai ◽

C A Carruthers ◽

M D Hollenberg

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Hydrophobic Interaction ◽

Binding Proteins ◽

Binding Protein ◽

Hydrophobic Interaction Chromatography ◽

Bovine Brain ◽

Dependent Protein Kinase ◽

Kinase C ◽

Dependent Protein

Several bovine brain proteins have been found to interact with a hydrophobic chromatography resin (phenyl-Sepharose CL-4B) in a Ca2+-dependent manner. These include calmodulin, the Ca2+/phospholipid-dependent protein kinase (protein kinase C) and a novel Ca2+-binding protein that has now been purified to electrophoretic homogeneity. This latter protein is acidic (pI 5.1) and, like calmodulin and some other high-affinity Ca2+-binding proteins, exhibits a Ca2+-dependent mobility shift on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, with an apparent Mr of 22 000 in the absence of Ca2+ and Mr 21 000 in the presence of Ca2+. This novel calciprotein is distinct from known Ca2+-binding proteins on the basis of Mr under denaturing conditions, Cleveland peptide mapping and amino acid composition analysis. It may be a member of the calmodulin superfamily of Ca2+-binding proteins. This calciprotein does not activate two calmodulin-dependent enzymes, namely cyclic nucleotide phosphodiesterase and myosin light-chain kinase, nor does it have any effect on protein kinase C. It may be a Ca2+-dependent regulatory protein of an as-yet-undefined enzymic activity. The Ca2+/phospholipid-dependent protein kinase is also readily purified by Ca2+-dependent hydrophobic-interaction chromatography followed by ion-exchange chromatography, during which it is easily separated from calmodulin. A preparation of protein kinase C that lacks contaminating kinase or phosphatase activities is thereby obtained rapidly and simply. Such a preparation is ideal for the study of phosphorylation reactions catalysed in vitro by protein kinase C.

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