Dissociation of Phosphorylation and Translocation of a Myristoylated Protein Kinase C Substrate (MARCKS Protein) in C6 Glioma and N1E‐115 Neuroblastoma Cells

Abstract: 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 4β‐12‐O‐tetradecanoylphorbol 13‐acetate (β‐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 β‐TPA to a similar extent in both fractions (maximal, fourfold at 30 min). In contrast, β‐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 α‐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.