Hatch G M, Vance D E
Faculty of Medicine, Heritage Medical Research Center, University of Alberta, Edmonton, Canada.
Biochem Cell Biol. 1991 Aug;69(8):515-22. doi: 10.1139/o91-076.
The effect of CPT-cAMP and okadaic acid on phosphatidylcholine catabolism in suspension cultures of choline-deficient rat hepatocytes was investigated. Choline-deficient hepatocytes were pulse-labeled for 30 min with [methyl-3H]choline and subsequently chased for up to 60 min with choline in the absence or presence of 0.5 mM CPT-cAMP or 0.5 microM okadaic acid. Radioactivity in phosphatidylcholine and lysophosphatidylcholine were unchanged during the chase. However, the radioactivity incorporated into glycerophosphocholine was significantly increased (P less than 0.05) 59 and 77% after 60 min of chase in hepatocytes incubated with either okadaic acid or CPT-cAMP, respectively. Incubation of choline-deficient hepatocytes with both okadaic acid and CPT-cAMP produced an additive effect on radioactivity incorporated ino glycerophosphocholine. Crude mitochondrial, microsomal, and cytosolic phospholipaselysophospholipase activities, assayed in the presence of exogenously labeled phosphatidylcholine, were unchanged in both CPT-cAMP and okadaic acid treated hepatocytes compared with control. Phospholipase-lysophospholipase activity, assayed with endogenously labeled phosphatidylcholine, was increased 28 and 47% (P less than 0.05) in the crude mitochondrial fraction of hepatocytes treated with either okadaic acid or CPT-cAMP, respectively, compared with the control. Incubation of choline-deficient hepatocytes, labeled with L-[methyl-3H]methionine, with CPT-cAMP or okadaic acid caused a 31 and 20% increase (P less than 0.05) in the radioactivity incorporated into glycerophosphocholine, respectively, compared with the control. We postulate that phosphatidylcholine catabolism in choline-deficient hepatocytes may be regulated by a phosphorylation-dephosphorylation mechanism mediated through cAMP-dependent protein kinase and phosphoprotein phosphatase activities.
研究了CPT-cAMP和冈田酸对胆碱缺乏大鼠肝细胞悬浮培养物中磷脂酰胆碱分解代谢的影响。胆碱缺乏的肝细胞用[甲基-3H]胆碱脉冲标记30分钟，随后在不存在或存在0.5 mM CPT-cAMP或0.5 μM冈田酸的情况下，用胆碱追踪长达60分钟。在追踪过程中，磷脂酰胆碱和溶血磷脂酰胆碱中的放射性没有变化。然而，在用冈田酸或CPT-cAMP孵育的肝细胞中，追踪60分钟后，掺入甘油磷酸胆碱中的放射性分别显著增加（P<0.05）59%和77%。同时用冈田酸和CPT-cAMP孵育胆碱缺乏的肝细胞，对掺入甘油磷酸胆碱中的放射性产生相加作用。与对照相比，在CPT-cAMP和冈田酸处理的肝细胞中，在外源标记的磷脂酰胆碱存在下测定的粗线粒体、微粒体和胞质磷脂酶-溶血磷脂酶活性没有变化。用内源性标记的磷脂酰胆碱测定时，与对照相比，用冈田酸或CPT-cAMP处理的肝细胞粗线粒体部分的磷脂酶-溶血磷脂酶活性分别增加28%和47%（P<0.05）。与对照相比，用L-[甲基-3H]蛋氨酸标记的胆碱缺乏肝细胞与CPT-cAMP或冈田酸孵育，分别使掺入甘油磷酸胆碱中的放射性增加31%和20%（P<0.05）。我们推测，胆碱缺乏肝细胞中的磷脂酰胆碱分解代谢可能受cAMP依赖性蛋白激酶和磷蛋白磷酸酶活性介导的磷酸化-去磷酸化机制调节。
