Diacylglycerol kinase activity in rat liver nuclei.
Membrane-depleted rat liver nuclei contain diacylglycerol (DAG) kinase showing a specific activity which doubles that of the whole homogenate. In contrast, cytoplasmic and plasma membrane marker enzymes attain a specific activity of 0.4% at the most, when nuclear DAG kinase approaches 4.5% of the total tissue activity. The enzyme shows a Km of 161 and 200 microM for ATP in both nuclei and microsomes whereas the Km for DAG is 75 microM in nuclei and 658 microM in microsomes. Octylglucoside, CHAPS and Triton X-100 behave mainly as inhibitors, while deoxycholate stimulates the enzyme activity in both cellular fractions, increasing specific activity (3.2-fold in nuclei and 29.1-fold in microsomes) and decreasing Km for DAG (39 microM in nuclei and 237 microM in microsomes). Phospholipids and ceramide stimulate the enzyme activity in isolated nuclei, while no effect occurs in the microsomal fraction. At variance, sphingosine behaves as an inhibitor in both cellular fractions. DAG kinase also utilizes endogenous substrates mobilized by Bacillus cereus phospholipase C, which hydrolyses nuclear phosphatidylcholine and phosphatidylethanolamine and by phosphatidylinositol-specific phospholipase C, which hydrolyses nuclear PI and PIP. These data indicate that nuclear DAG can be controlled by converting it into phosphatidic acid by the action of a nuclear enzyme and support the contention that protein kinase C activity can be modulated at the nuclear level by a discrete system involving phospholipase C and DAG kinase that could operate independently from the cytoplasm.
Previati
M
M
Istituto di Anatomia Umana Normale, Università di Ferrara, Italy.
Bertagnolo
V
V
Mazzoni
M
M
Osti
F
F
Borgatti
P
P
Capitani
S
S
D000818:Animals; D002467:Cell Nucleus / Q000201:enzymology*; D003902:Detergents / Q000494:pharmacology; D019852:Diacylglycerol Kinase; D004789:Enzyme Activation; D004791:Enzyme Inhibitors; D004795:Enzyme Stability; D008099:Liver / Q000201:enzymology*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010972:Platelet Activating Factor / Q000037:antagonists & inhibitors; D011744:Pyrimidinones / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013379:Substrate Specificity; D013844:Thiazoles / Q000494:pharmacology
D003902:Detergents; D004791:Enzyme Inhibitors; D010972:Platelet Activating Factor; D011744:Pyrimidinones; D013844:Thiazoles; C047505:R 59022; D017853:Phosphotransferases (Alcohol Group Acceptor); D019852:Diacylglycerol Kinase