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 homogenat e. In contrast, cytoplasmic and plasma membrane marker enzymes attain a specific activity of 0.4% at the most, when nuclear DAG kinase appro aches 4.5% of the total tissue activity. The enzyme shows a K-m of 161 and 200 mu M for ATP in both nuclei and microsomes whereas the K, for DAG is 75 mu M in nuclei and 658 mu M 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) an d decreasing K-m for DAG (39 mu M in nuclei and 237 mu M in microsomes ). Phospholipids and ceramide stimulate the enzyme activity in isolate d nuclei, while no effect occurs in the microsomal fraction. At varian ce, sphingosine behaves as an inhibitor in both cellular fractions. DA G kinase also utilizes endogenous substrates mobilized by Bacillus cer eus phospholipase C, which hydrolyses nuclear phosphatidylcholine and phosphatidylethanolamine and by phosphatidlyinositol-specific phosphol ipase 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 pro tein kinase C activity can be modulated at the nuclear level by a disc rete system involving phospholipase C and DAG kinase that could operat e independently from the cytoplasm.