Free cholesterol induces activation but not translocation of protein kinase C in cultured ascites tumour cells.
Ascites tumour cells, cultivated in a serum replacement medium with low mitogenic activity supplemented with free cholesterol or cholesterol complexed to digitonine, were used to study protein kinase C (PKC) activation and its translocation from the cytosol to the membrane compartment. Activity of partially purified PKC was assessed by the Ca2+/phospholipid-dependent phosphorylation of histone as substrate. Staurosporine-sensitive PKC activity was increased by a factor of about three in the membrane compartment as the result of cholesterol supplementation with about 90% of total activity in the membrane, and the remaining 10% in the cytosolic fraction. A translocation of enzyme mass from the cytosol to the membrane compartment was not induced under these conditions, as documented by the enzyme-linked immuno adsorbent assay using a monoclonal anti-PKC antibody. In cells cultivated in the presence of the cholesterol-digitonide complex PKC activation was not observed, suggesting an inadequate utilization of cholesterol as membrane constituent.
Haeffner
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Forschungsschwerpunkt Diagnostik und Experimentelle Therapie, Deutsches Krebsforschungszentrum, Heidelberg, Germany.
Wittmann
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U
D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002286:Carcinoma, Ehrlich Tumor; D002462:Cell Membrane / Q000201:enzymology; D002784:Cholesterol / Q000494:pharmacology*; D003600:Cytosol / Q000201:enzymology; D004072:Digitonin / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019311:Staurosporine; D014407:Tumor Cells, Cultured
D000470:Alkaloids; D002784:Cholesterol; D011493:Protein Kinase C; D019311:Staurosporine; D004072:Digitonin