Evidence for a protein kinase C‐directed mechanism in the phorbol diester‐induced phospholipase D pathway of diacylglycerol generation from phosphatidylcholine

FEBS Letters

Zhang, Zu-chuan¹  Cabot, Myles C.¹  Welsh, Clement J.¹  Cao, Hui-ting¹ 

[1] W. Alton Jones Cell Science Center, Inc., Lake Placid, NY 12946, USA

关键词: Protein kinase C;    Phospholipase D;    Phorbol diester;    Phosphatidylcholine;    Diacylglycerol;    DG;    diacylglycerol;    PKC;    Ca2+-activated;    phospholipid-dependent protein kinase C;    TPA;    12-O-tetradecanoyl phorbol-13-acetate;    PC;    phosphatidylcholine;    PDBu;    phorbol dibutyrate;    Bu2cAMP;    2′-O-dibutyryladenosine 3′;    5′-cyclic monophosphate;    BSA;    bovine serum albumin;    Me2SO;    dimethyl sulfoxide;    DiC8;    dioctanoyl glycerol;    PBS;    phosphate-buffered saline;    TLC;    thin-layer chromatography;    IP3;    myo-inositol 1;    4;    5-trisphosphate;    PE;    phosphatidylethanol. In the naming of PC;    PA;    DG and PE;    we do not distinguish between the type of aliphatic linkage to glycerol (ester;    ether);   

DOI  :  10.1016/0014-5793(89)80197-8

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

In this study we provide evidence for the involvement of protein kinase C (PKC) in phorbol diester-induced phosphatidylcholine (PC) hydrolysis by the phospholipase D pathway. Rat embryo fibroblasts (REF52) were prelabeled with either tritiated choline or myristic acid; these compounds are preferentially incorporated into cellular PC. Phorbol diester-induced PC degradation was determined by measuring the release of [³H]choline, and the formation of [³H]myristoyl-containing phosphatidate (PA), diacylglycerol (DG), and phosphatidylethanol (PE). Staurosporine, a PKC inhibitor, blocked from 73 to 90% of the phorbol diester-induced PC hyrolysis. The inhibition of phorbol diester-induced choline release by staurosporine was dose dependent with an approximate ED₅₀ of 150 nM. Pretreatment of cells with phorbol diester inhibited subsequent phorbol diester-induced PC degradation by 78–92%. A close correlation between the ED₅₀ for phorbol diester-stimulated choline release and the K _d for phorbol diester binding was demonstrated. Neither forskolin nor dibutyryl cAMP elicited cellular PC degradation. In vitro experiments using phospholipase D from Streptomyces chromofuscus showed that staurosporine did not inhibit and TPA did not stimulate enzyme activity.

【 授权许可】

Unknown