Protein kinase C in primary astrocyte cultures: cytoplasmic localization and translocation by a phorbol ester.
The distribution of calcium-activated, phospholipid-dependent protein kinase (protein kinase C) in supernatant and particulate fractions of primary cultures of rat astrocytes and its translocation by a phorbol ester were studied. We observed that 91% of protein kinase C activity in astrocytes was in the supernatant fraction, as measured by lysine-rich histone phosphorylation assay. Attempts to uncover latent activity in the particulate fraction were unsuccessful. Approximately 75% of the supernatant protein kinase C activity could be translocated to the particulate fraction by prior treatment (30-60 min) of the cultures with 100 nM 12-O-tetradecanoyl-phorbol 13-acetate (TPA), but not with 4 alpha-phorbol, an inactive phorbol ester. Investigation of endogenous substrates for protein kinase C showed that TPA treatment brought about an increase in phosphorylation in membrane proteins and a decrease in phosphorylation of supernatant proteins. These findings indicate that the distribution of protein kinase C in astrocytes differs substantially from that in whole brain tissue, where approximately two-thirds of the protein kinase C activity is associated with the particulate fraction. Because protein kinase C is concentrated in the cytosol of astrocytes and most of this activity can be translocated to membranes, astrocytes may be particularly well-suited to respond to signals that activate phosphoinositide-linked receptors in brain.
Neary
J T
JT
Laboratory of Neuropathology, Veterans Administration Medical Center, Miami, FL 33125.
Norenberg
L O
LO
Norenberg
M D
MD
D000818:Animals; D000831:Animals, Newborn / Q000378:metabolism; D001253:Astrocytes / Q000201:enzymology*; D001692:Biological Transport / Q000187:drug effects; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D002852:Chromatography, Ion Exchange; D003593:Cytoplasm / Q000201:enzymology; D003902:Detergents / Q000494:pharmacology; D007700:Kinetics; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D003902:Detergents; D010750:Phosphoproteins; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate