Diacylglycerols mimic phorbol diester induction of leukemic cell differentiation.

Abstract

Activation of cellular protein kinase C appears to be involved in the mechanism by which phorbol diesters induce differentiation of human myeloid leukemia cells . Protein kinase C is thought to be physiologically activated by diacylglycerol derived from receptor-mediated phosphatidylinositol hydrolysis. sn-1,2-diacylglycerols with short saturated acyl side chains (C4-C10) were synthesized and found to be potent activators of protein kinase C partially purified from HL-60 cells. These diacylglycerols were also competitive inhibitors of [3Hlphorbol dibutyrate binding to the soluble phorbol diester receptor. The most potent diacylglycerol, sn-1,2-dioctanoylglycerol, displaced >90% of[3H]phorbol dibutyrate from the phorbol diester receptor of intact HL-60 cells. Because of probable cellular metabolism of sn-1,2-dioctanoylglycerol, hourly doses were required to maintain persistent occupancy of the phorbol diester binding site. Treatment of HL-60 cells with either phorbol 12-myristate 13-acetate or sn-1,2-dioctanoylglycerol produced identical phosphoprotein changes. Finally, sn-1,2-dioctanoylglycerol induced differentiation of the HL-60 cells into cells with morphologic characteristics of macrophages. Substitution of the hydroxyl group at position 3 with a hydrogen, chloro, or sulfhydryl moiety inactivated sn-1,2-dioctanoylglycerol. These data strengthen the hypothesis that protein kinase C activation plays a role in macrophage differentiation.The phorbol diester tumor promoters are plant products that enhance tumor formation in two-stage models of skin carcinogenesis (1). Paradoxically, these same compounds will rapidly induce most myeloid leukemia cells (cell lines and fresh explants from patients) to cease proliferation and to differentiate into cells with the morphologic, antigenic, biochemical, and functional properties of macrophages (2, 3). The phorbol diesters exert these effects via a specific receptor (4, 5). Data from several laboratories indicate that this receptor is the Ca2+-phospholipid-dependent protein kinase, termed protein kinase C (6-9). We have recently presented data that suggest activation of protein kinase C is involved in differentiation of human promyelocytic leukemia cells (HL-60) induced by phorbol diesters (10).Protein kinase C is regulated in vitro by Ca2 + and diacylglycerol, and is presumably activated in vivo in response to receptor-mediated phosphatidylinositol turnover, which gives rise to diacylglycerol, and/or Ca2+ flux (11). In vitro, the phorbol diesters activate the kinase by interaction at the diacylglycerol site (12). In intact cells, the phorbol diesters bypass the receptor-phosphatidylinositol-mediated regulatory mechanisms and activate the kinase directly (11). Discrete and sustained changes in cellular phosphoproteins have been shown to occur during phorbol diester-induced differentiation of HL-60 cells, although evidence that these phosphorylation events are protein kinase C dependent or are involved in the process of differentiation has not been presented (13,14)...

Hypoxia-induced accumulation of erythropoietin mRNA in isolated hepatocytes is inhibited by protein kinase C

“…However, the results obtained so far appear entirely compatible with an inhibitory effect of the activation of PKC alpha on EPO mRNA accumulation in hepatocytes. A low efficacy of synthetic diacylglyerol analogues in inducing PKC-mediated reactions in hepatocytes has previously been observed [11,27], possibly due to high rates of metabolism of these substances [14]. Thus the failure to lower EPO mRNA levels with DOG or OAG appears no compelling argument against PKC-mediated reduction of EPO gene expression.…”

Near neighbour analysis of variant cell lines derived from the promyeloid cell line HL60

“…However, this also poses the question of availability of appropriate kinases. Protein kinase C has been implicated in the differentiation of HL60 cells towards monocytes (Ebeling et al, 1985;Vandenbark et al, 1984) and two groups have recently described multiple protein kinase C genes (Knopf et al, 1986;Coussens et al, 1986). It is interesting to speculate that the sequential expression or activation of particular protein kinases C and/ or other protein kinases and their co-ordinated action may play a vital role in the commitment of HL60 cells to one pathway or another of differentation.…”

Inhibition of vasopressin-stimulated water flow in toad bladder by phorbol myristate acetate, dioctanoylglycerol, and RHC-80267. Evidence for modulation of action of vasopressin by protein kinase C.

“…Furthermore, cell-permeable diglycerides such as dioctanoylglycerol (DiC8) can also mimic the action of phorbol esters or of diglycerides generated in the cell (10). We have recently shown that AVP can influence phosphatidylinositol turnover in toad bladder epithelial cells (I 1), thereby suggesting a role for the phospholipid-calcium-protein kinase C system in the physiologic response to AVP here as well.…”