Translocation of protein kinase C in human leukemia cells susceptible or resistant to differentiation induced by phorbol 12-myristate 13-acetate.

Abstract

We investigated the possible relationship between the susceptibility of cells to differentiation induced by phorbol 12-myristate 13-acetate (PMA) and the subcellular translocation of calcium-and phospholipid-dependent protein kinase (protein kinase C) activity from the cytosol to the membrane. These two events were analyzed in a number of human leukemia cell lines, including four cell variants of the promyelocytic cell line HL-60 that exhibit different degrees of susceptibility to PMA-induced differentiation. The phenotype of the differentiated cells was characterized by increased reactivity with monoclonal antibodies against maturationspecific cell surface antigens, increased nonspecific esterase activity, and acquisition of morphological cell maturation. Analysis of the subcellular distribution of protein kinase C activity in each of these cell types revealed that 90% of the kinase activity was present in the cytosolic fraction, with the remaining activity in the membrane fraction. Treatment of the differentiation-susceptible cells with 160 nM PMA resulted, within 5 min after treatment, in a >60% decrease in protein kinase C activity in the cytosolic fraction and a >1500% increase in the activity in the membrane fraction. No such subcellular redistribution of protein kinase C activity was found after treatment of the differentiation-resistant cells. On the basis of these findings, we suggest that the process of subcellular translocation of protein kinase C activity, initiated after the binding of PMA to this kinase, is required for the induction of cell differentiation by this phorbol diester.

Alkyllysophospholipid ET‐18‐OCH3 acts as an activator of protein kinase C in HL‐60 cells

“…The incorporated radioactivity was determined as described above. I-IL-60 cells measured with (partially) purified PK-C [17] showing that our model is very useful to investigate the activity of PK-C in its native environxnent. The action of ET-l&OCH, on PK-C in I-IL-60 cells was investigated at subtoxic concentrations of ET-1 g-QCI-I,, 2O,@nl for 4 h at 37°C [18].…”

A lineage-specific protein kinase crucial for myeloid maturation

“…Human myeloid leukemia cell line HL-60 was originally obtained from R. C. Gallo (National Cancer Institute, Bethesda, MD). The differentiation-resistant cell variant HL-525 was obtained by subculturing HL-60 cells 102 times in the presence of increasing concentrations (up to 3 M) of PMA at 5-to 8-day intervals (12). Human monocytes, granulocytes, and lymphocytes were isolated from peripheral whole venous blood by Ficoll-Hypaque density gradient (1.077 g͞ml) centrifugation as previously described (17).…”

“…We have developed a variant of HL-60 cells that is deficient in PKC-␤, an abundant isoform of PKC in the HL-60 cell line (13,14). This cell variant, HL-525, is resistant to macrophage maturation induced by a known PKC activator, phorbol 12-myristate 13-acetate (PMA), but sensitive to inducers of monocyte and granulocyte differentiation (12,14).…”

“…The human myeloid leukemia HL-60 cell line is commonly used as a model system for studying myeloid cell development (7)(8)(9)(10)(11)(12)(13)(14)(15) because these cells can be induced to develop into macrophages (7,8), monocytes (9), and granulocytes (10,11) and exhibit useful functional and morphological markers upon maturation. Macrophage maturation in HL-60 cells is initiated upon activation of protein kinase C (PKC), which acts as both a specific receptor and a functional signal transduction protein (12,13).…”

“…Macrophage maturation in HL-60 cells is initiated upon activation of protein kinase C (PKC), which acts as both a specific receptor and a functional signal transduction protein (12,13). We have developed a variant of HL-60 cells that is deficient in PKC-␤, an abundant isoform of PKC in the HL-60 cell line (13,14).…”