Possible involvement of protein kinase C-epsilon in phorbol ester-induced growth inhibition of human lymphoblastic cells.
Sustained activation of members of the protein kinase C (PKC) family is known to influence the growth and differentiation of various cell types, however, the specific roles for individual isoforms mediating these cellular events have yet to be elucidated. Activation of PKC by phorbol esters leads to growth inhibition in certain cell lines. The HT58 human B lymphoblastic cell may serve as a cellular model system to investigate the participation of individual isoforms in the initial events of growth arrest induced by phorbol ester. Determination of cell cycle and investigation of apoptosis were performed by flow cytometric measurements. Phorbol ester-induced translocation and down-regulation of the conventional alpha, beta and the novel epsilon isoforms of PKC were demonstrated by Western blot analysis. At lower concentrations (o.5 ng/ml) phorbol myristate acetate (PMA) stimulated a G1 arrest with retention of viability in the human HT58 B lymphoblastic cell. The protein kinase inhibitor staurosporine at a concentration of 25 nM did not significantly alter HT58 cell viability. However, staurosporine (25 nM) induced apoptosis in cells preincubated for 4 hr with 0.5-1.0 ng/ml PMA. The translocation of PKC-epsilon was observed within 39 min exposure to 0.5 ng/ml PMA. After a 4 hr treatment, evidence for down-regulation and and altered phosphorylation state of PKC-epsilon was seen. In contrast, the conventional alpha and beta isoforms were practically uneffected by this PMA treatment. At higher PMA concentrations (50 ng/ml) the alpha and beta isoforms showed a significant down-regulation. The preferential alterations in PKC-epsilon observed under the conditions required for PMA to influence the growth and survival of HT58 cells suggest a role for the Ca(2+)-independent epsilon isoform in mediating the initial events of the phorbol ester stimulated cellular responses.
Mihalik
R
R
Department of Immunology, National Institute of Haematology, Blood Transfusion and Immunology, Budapest Hungary.
Farkas
G
G
Kopper
L
L
Benczúr
M
M
Faragó
A
A
D017209:Apoptosis / Q000187:drug effects*; D001402:B-Lymphocytes / Q000166:cytology / Q000378:metabolism*; D015153:Blotting, Western; D002273:Carcinogens / Q000494:pharmacology*; D002453:Cell Cycle / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D005434:Flow Cytometry; D006801:Humans; D010703:Phorbol Esters / Q000494:pharmacology*; D011493:Protein Kinase C / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*
D002273:Carcinogens; D010703:Phorbol Esters; D011493:Protein Kinase C