| Abstract: |
Phorbol esters have been shown to stimulate phosphatidylcholine synthesis via the CDP-choline pathway. The present study compares the effects of phorbol esters and thyrotropin-releasing hormone (TRH) on phosphatidylcholine metabolism in GH3 pituitary cells. In a previous study (Kolesnick, R.N., and Paley, A.E. (1987) J. Biol. Chem. 262, 9204-9210), the potent phorbol ester, 12-O-tetradecanoylphorbol 13-acetate (TPA) induced time- and concentration-dependent incorporation of 32P(i) and [3H]choline into phosphatidylcholine in short-term labeling experiments. In this study, TPA is shown to activate choline-phosphate cytidylyltransferase (EC 2.7.7.15), the regulatory enzyme of the CDP-choline pathway, by stimulating redistribution of the inactive cytosolic form of the enzyme to the membrane. Redistribution was quantitative. TPA reduced cytosolic activity from 3.5 ± 0.4 to 1.5 ± 0.3 nmol·min-1·107 cells-1 and enhanced particulate activity from 2.5 ± 0.4 to 4.9 ± 0.6 nmol·min-1·107 cells-1. TRH also stimulated time- and concentration-dependent 32P(i) and [3H]choline incorporation into phosphatidylcholine. An increase was detectable after 5 min; and after 30 min, the levels were 164 ± 9 and 150 ± 11% of control, respectively; EC50 ~ 2 x 10-10 M TRH. These events correlated directly with TRH-induced 32P(i) incorporation into phosphatidylcholine. TRH also stimulated redistribution of cytidylyltransferase specific activity. TRH reduced cytosolic activity 45% and enhanced particulate activity 51%. Neither TRH nor TPA stimulated phosphatidylcholine degradation. In cells down-modulated for protein kinase C (Ca2+/phospholipid-dependent protein kinase), the effects of TPA and TRH on 32P(i) incorporation into phosphatidylcholine were abolished. However, TRH-induced incorporation into phosphatidylinositol still occurred. These studies provide evidence that hormones may regulate phosphatidylcholine metabolism via the protein kinase C pathway. |
