Modulation of protein kinase C activity and calcium-sensitive isoform expression in human myeloid leukemia cells by bryostatin 1: relationship to differentiation and ara-C-induced apoptosis.
Previous studies have shown that pretreatment of human myeloid leukemia cells (HL-60) with the protein kinase C (PKC) activator bryostatin 1 potentiates ara-C-induced apoptosis. To test the hypothesis that this capacity stems from down-regulation of PKC activity and/or Ca2+-dependent (group-I; cPKC) isoform expression, comparisons were made between the effects of this agent and the stage-2 tumor promoter mezerein under conditions favoring either cellular differentiation or drug-induced apoptosis. Twenty-four-hour pretreatment of HL-60 cells with 10 nM bryostatin 1, which does not induce differentiation in this cell line, led to a profound reduction in membrane and cytosolic PKC activity, decreased expression of cPKC isoforms (alpha, betaI, betaII, gamma), and a marked increase in ara-C induced apoptosis. In contrast, 10 nM mezerein, which induces HL-60 cell differentiation, was less effective in down-regulating membrane and cytosolic PKC activity as well as alpha, betaI, and gamma cPKC isoform expression, and failed to potentiate ara-C-related apoptosis. The effects of bryostatin 1 were dominant to those of mezerein, in that the combination resulted in down-regulation of PKC activity and expression and potentiation of ara-C-induced apoptosis, but not cellular maturation. However, coadministration of the Ca2+ ionophore A23187 (250 nM) restored bryostatin 1's differentiating ability while antagonizing its capacity to augment apoptosis, despite failing to reverse bryostatin 1-induced down-regulation of PKC activity and cPKC isoform expression. Furthermore, pretreatment of differentiation-responsive monocytic leukemia cells (U937) with bryostatin 1 substantially reduced PKC activity and cPKC isoform expression, but exerted minimal effects on ara-C-related apoptosis. In contrast, exposure of U937 cells to bryostatin 1 after ara-C dramatically increased apoptosis, a phenomenon that did not occur in differentiation-unresponsive HL-60 cells. Collectively, these observations indicate that down-regulation of total assayable PKC activity and cPKC expression by bryostatin 1 are insufficient, by themselves, to account for potentiation of leukemic cell apoptosis, at least under conditions in which differentiation occurs. They also provide further evidence that a reciprocal and highly schedule-dependent relationship exists between leukemic cell differentiation and drug-induced apoptosis.
Department of Medicine, Medical College of Virginia, Richmond 23298, USA.
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S
Grant
A J
AJ
Turner
A J
AJ
Freemerman
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Z
Wang
L
L
Kramer
W D
WD
Jarvis
D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D054713:Bryostatins; D002118:Calcium / Q000378:metabolism*; D002273:Carcinogens / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D003561:Cytarabine / Q000494:pharmacology; D004224:Diterpenes; D004789:Enzyme Activation / Q000187:drug effects; D018922:HL-60 Cells; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D007783:Lactones / Q000494:pharmacology*; D018942:Macrolides; D011493:Protein Kinase C / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013729:Terpenes / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000964:Antimetabolites, Antineoplastic; D054713:Bryostatins; D002273:Carcinogens; D004224:Diterpenes; D007527:Isoenzymes; D007783:Lactones; D018942:Macrolides; D013729:Terpenes; D003561:Cytarabine; C011309:mezerein; C046785:bryostatin 1; D011493:Protein Kinase C; D002118:Calcium