Dissociation of phosphorylation and translocation of a myristoylated protein kinase C substrate (MARCKS protein) in C6 glioma and N1E-115 neuroblastoma cells.
An 80-kDa protein labeled with [3H]myristic acid in C6 glioma and N1E-115 neuroblastoma cells has been identified as the myristoylated alanine-rich C kinase substrate (MARCKS protein) on the basis of its calmodulin-binding, acidic nature, heat stability, and immunochemical properties. When C6 cells preincubated with [3H]myristate were treated with 200 nM 4 beta-12-O-tetradecanoylphorbol 13-acetate (beta-TPA), labeled MARCKS was rapidly increased in the soluble digitonin fraction (maximal, fivefold at 10 min) with a concomitant decrease in the Triton X-100-soluble membrane fraction. However, phosphorylation of this protein was increased in the presence of beta-TPA to a similar extent in both fractions (maximal, fourfold at 30 min). In contrast, beta-TPA-stimulated phosphorylation of MARCKS in N1E-115 cells was confined to the membrane fraction only and no change in the distribution of the myristoylated protein was noted relative to alpha-TPA controls. These results indicate that although phosphorylation of MARCKS by protein kinase C occurs in both cell lines, it is not directly associated with translocation from membrane to cytosol, which occurs in C6 cells only. The cell-specific translocation of MARCKS appears to correlate with previously demonstrated differential effects of phorbol esters on stimulation of phosphatidylcholine turnover in these two cell lines.
Byers
D M
DM
Department of Pediatrics, Dalhousie University, Halifax, Nova Scotia, Canada.
Palmer
F B
FB
Spence
M W
MW
Cook
H W
HW
D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004355:Drug Stability; D005910:Glioma / Q000378:metabolism*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D047908:Intracellular Signaling Peptides and Proteins; D007700:Kinetics; D008565:Membrane Proteins; D051379:Mice; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D009447:Neuroblastoma / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D002147:Calmodulin; D047908:Intracellular Signaling Peptides and Proteins; C000619711:Marcks protein, mouse; C000619712:Marcks protein, rat; D008565:Membrane Proteins; D009227:Myristic Acids; D011506:Proteins; D019814:Myristic Acid; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium