Bombesin and phorbol ester stimulate phosphatidylcholine hydrolysis by phospholipase C: Evidence for a role of protein kinase C

Abstract

Bombesin caused a marked stimulation of ³²P_i into phosphatidylinositol (Pl), with no apparent lag, and into phosphatidylcholine (PC), after a lag of about 20 min. Stimualtion was blocked by the bombesin receptor antagonist, [D-Arg¹, D-Pro², D-Trp^7,9, Leu¹¹] substance P, indicating that the effects on both Pl and PC were mediated through the same receptor. The tumor-promoting phorbol ester 12-0-tetradecanoylphorbol-13-acetate (TPA) and dioctanoylglycerol (diC₈) both directly activate protein kinase C and in this report were shown to stimulate ³²P_i incorporation into PC but not into Pl. In addition, TPA stimulated the release of [³H]choline and [³H]phosphocholine and the accumulation of [³H]diacylglycerol from prelabelled cells. These results strongly suggest that TPA activates a phospholipase C specific for PC. Pretreatment of cells with phorbol-12, 13-dibutyrate (PDBu) for 24 h depleted cellular protein kinase C activity and inhibited the ability of TPA to induce these effects suggesting a direct involvement of protein kinase C. Similarly the bombesin stimulation of ³²P_i into PC and of [³H]choline and [³H]phosphocholine release was inhibited by PDBu pretreatment. DiC₈ and, to a lesser extent, TPA stimulated the translocation of CTP:phosphocholine cytidylyltransferase from the cytosolic to the particulate fraction. DiC₈ also stimulated this translocation in cells depleted of protein kinase C. It was concluded that both bombesin and TPA activated protein kinase C leading to activation of a phospholipase C specific for PC.