Protein Kinase CβII Activation by 1-β-d-Arabinofuranosylcytosine Is Antagonistic to Stimulation of Apoptosis and Bcl-2α Down-regulation

Abstract

1-␤-D-Arabinofuranosylcytosine (ara-C. ara-C also causes apoptosis in HL-60 cells which can be mimicked by exogenous ceramide. However, the signaling role for ara-C-induced diacylglycerol (DAG) is not defined. We found that Bcl-2 levels were increased by treatment of HL-60 cells with exogenous DAG or 12-O-tetradecanoylphorbol-13-acetate (TPA). In contrast, exogenous ceramide treatment caused a decrease in cellular Bcl-2 levels. Thus, ara-C stimulates the synthesis of two second messengers with opposing effects on Bcl-2. Since the effects of ara-Cinduced DAG could be due to protein kinase C (PKC) activation, we determined the effects of ara-C on PKC isozymes. ara-C caused an increase in membrane-bound PKC␤II (but not PKC␣ or PKC␦). ara-C or TPA-induced translocation of PKC␤II was inhibited by 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH 3 ), and ara-C-induced apoptosis was stimulated by pretreatment of the cells with ET-18-OCH 3 . ET-18-OCH 3 also inhibited stimulation of Bcl-2 by TPA and enhanced the decrease in Bcl-2 observed in ara-C-treated cells. These data indicate that ara-C-induced apoptosis is limited by ara-C-stimulated PKC␤II through effects on Bcl-2. To further determine the role of PKC, we used antisense oligonucleotides directed toward PKC␤II. The antisense, but not the sense, oligonucleotide inhibited PKC␤II activation and enhanced ara-C-induced apoptosis. These data demonstrate that the stimulation of apoptosis by ara-C is self-limiting and can be enhanced by inhibition of PKC.

The roles of signaling by the p42/p44 mitogen-activated protein (MAP) kinase pathway; a potential route to radio- and chemo-sensitization of tumor cells resulting in the induction of apoptosis and loss of clonogenicity

“…In agreement with this hypothesis, we and others have found that pharmacologic agents which directly inhibit PKC (eg staurosporine), 99a down-regulate PKC upon chronic exposure (eg bryostatin 1), or ablate PKC expression (eg PKC antisense oligonucleotides) have all been shown to enhance the lethal actions of Ara-C. 93,95,100 Interestingly, in the latter study, ceramide exposure decreased whereas diglyceride exposure increased expression of the anti-apoptotic protein Bcl-2 in leukemic cells. 97,100 More recently, bryostatin 1 and the PKC inhibitor safingol have been shown to block Ara-Cmediated MAP kinase activation in association with potentiation of leukemic cell apoptosis, effects which have found to be mimicked by the MEK1 inhibitor PD98059. 92,93,101 Bryostatin 1 and PD98059 have also been found to promote taxolinduced apoptosis in leukemic cells.…”

Human munc13 Is a Diacylglycerol Receptor that Induces Apoptosis and May Contribute to Renal Cell Injury in Hyperglycemia

“…PDBu is a reagent known to be a tumor promoter capable of stimulating cell proliferation through PKC activation (Mochly-Rosen and Kauvar, 1998). Although the role of PKC in apoptosis is not consistent in the literature (Lavin et al, 1996;Deacon et al, 1997), the bulk of evidence suggests that PKC, especially PKC␣, activated by phorbol esters such as phorbol 12-myristate 13-acetate and PDBu, inhibits apoptosis (Lavin et al, 1996;Deacon et al, 1997;Whitman et al, 1997;Mochly-Rosen and Kauvar, 1998). In the case of PKCinduced apoptosis, there is evidence suggesting that down-regulation rather than DAG activation of PKC is responsible for this effect, because prolonged exposure to a stable phorbol ester form could result in downregulation of PKC expression (Leszczynski, 1996;Deacon et al, 1997).…”

Protein kinase C-α overexpression stimulates Akt activity and suppresses apoptosis induced by interleukin 3 withdrawal

“…To date, dierent genes including Bcl-2 family members, caspases, and other signaling pathways have been implicated in apoptosis (Cory, 1995;Korsmeyer, 1995). Several reports indicate that dierent PKCs are also involved in apoptotic signaling pathways (Barr et al, 1997;Berra et al, 1997;Datta et al, 1997;DiazMeco et al, 1996;Emoto et al, 1995Emoto et al, , 1996Ghayur et al, 1996;Mizuno et al, 1997;Whelan and Parker, 1998;Whitman et al, 1997). When U937 cells were exposed to ionizing radiation, TNF-a or anti-Fas antibody, cells underwent apoptosis (Emoto et al, 1995).…”

“…These data suggest that activation of nPKCs by protease cleavage may allow these PKC isoenzymes to exert pro-apoptotic eects. In contrast, PKC-bII was demonstrated to be critical in antagonizing Ara-Cinduced apoptosis in the HL-60 acute myelogenous leukemia line (Whitman et al, 1997) and in inhibiting c-myc-induced apoptosis of small cell lung cancer cells (Barr et al, 1997).…”