Lindmar R, Löffelholz K
Department of Pharmacology, University of Mainz, Germany.
Biochem Pharmacol. 1998 Oct 1;56(7):799-805. doi: 10.1016/s0006-2952(97)00636-9.
Activation of phospholipase D (PLD) and phosphoinositide-specific phospholipase C (PI-PLC) by fluoride, to stimulate heterotrimeric G-proteins, and by phorbol esters, to stimulate protein kinase C (PKC), was studied in rat atria. Fluoride and 4beta-phorbol-12beta,13alpha-dibutyrate (PDB), in contrast to 4beta-phorbol-13alpha-acetate (PAc), activated PLD, catalyzing the formation of [3H]-phosphatidylethanol ([3H]-PETH), [3H]-phosphatidic acid ([3H]-PA), choline and sn-1,2-diacylglycerol (DAG). Basal PLD activity was resistant to drastic changes in Ca2+ and to Ro 31-8220, a PKC inhibitor, but was decreased by genistein, an inhibitor of tyrosine kinase, and increased by vanadate, a tyrosine phosphatase inhibitor; both effects were, however, very small. Fluoride-evoked PLD activity was resistant to Ro 31-8220 and to genistein, but was Ca2+-dependent. The rate of fluoride-induced PLD activation was maintained for at least 60 min. In contrast, PDB-mediated PLD activity was blocked by Ro 31-8220 and was resistant to extracellular Ca2+-depletion and desensitized within ca. 15 min. PDB markedly potentiated the fluoride-evoked generation of [3H]-phosphatidylethanol and of choline, but inhibited the formation of [3H]-inositol phosphates ([3H]-IP(1-3)). Ethanol (2%) blocked the PDB-evoked generation of both [3H]-phosphatidic acid and of sn-1,2-diacylglycerol, whereas fluoride-evoked responses were reduced only to approximately 50%. In conclusion, the trimeric G-protein-PLD pathway in heart tissue did not enclose PKC activation and was long-lasting and Ca2+-dependent; there was no evidence for an involvement of tyrosine phosphorylation. However, PKC activation modulated G-protein-coupled PLD and PI-PLC activities in opposite directions. PLD activity significantly contributed to the mass production of sn-1,2-diacylglycerol in the heart. The evidence for a pathophysiological role of PLD activation in cardiac hypertrophy and in ischemic preconditioning is discussed.
在大鼠心房中研究了氟化物对磷脂酶D（PLD）和磷酸肌醇特异性磷脂酶C（PI-PLC）的激活作用，以刺激异源三聚体G蛋白，以及佛波酯对蛋白激酶C（PKC）的激活作用。与4β-佛波醇-13α-乙酸酯（PAc）不同，氟化物和4β-佛波醇-12β,13α-二丁酸酯（PDB）激活了PLD，催化形成[3H]-磷脂酰乙醇（[3H]-PETH）、[3H]-磷脂酸（[3H]-PA）、胆碱和sn-1,2-二酰基甘油（DAG）。基础PLD活性对Ca2+的剧烈变化和PKC抑制剂Ro 31-8220具有抗性，但被酪氨酸激酶抑制剂染料木黄酮降低，并被酪氨酸磷酸酶抑制剂钒酸盐增加；然而，这两种作用都非常小。氟化物诱导的PLD活性对Ro 31-8220和染料木黄酮具有抗性，但依赖于Ca2+。氟化物诱导的PLD激活速率至少维持60分钟。相比之下，PDB介导的PLD活性被Ro 31-8220阻断，对细胞外Ca2+耗竭具有抗性，并在约15分钟内脱敏。PDB显著增强了氟化物诱导的[3H]-磷脂酰乙醇和胆碱的生成，但抑制了[3H]-肌醇磷酸（[3H]-IP(1-3)）的形成。乙醇（2%）阻断了PDB诱导的[3H]-磷脂酸和sn-1,2-二酰基甘油的生成，而氟化物诱导的反应仅降低到约50%。总之，心脏组织中的三聚体G蛋白-PLD途径不涉及PKC激活，是持久的且依赖于Ca2+；没有证据表明酪氨酸磷酸化参与其中。然而，PKC激活以相反的方向调节G蛋白偶联的PLD和PI-PLC活性。PLD活性对心脏中sn-1,2-二酰基甘油的大量产生有显著贡献。讨论了PLD激活在心脏肥大和缺血预处理中的病理生理作用的证据。
