TY - JOUR
T1 - Transcriptional activation of the gp91phox NADPH oxidase subunit by TPA in HL-60 cells
AU - Samuelson, David J.
AU - Powell, Marianne B.
AU - Lluria-Prevatt, Maria
AU - Romagnolo, Donato F.
PY - 2001
Y1 - 2001
N2 - The exposure to epigenetic effectors capable of inducing copious production of reactive oxygen species (ROS) has been associated with chronic inflammation, tumor initiation, and promotion. The objective of this study was to examine the regulation of gp91phox, the catalytic subunit of the NADPH oxidase, and the kinetics of ROS production in promyelocytic leukemia HL-60 cells induced with 12-O-tetradeconylphorbol-13-acetate (TPA). The treatment of HL-60 cells with TPA (0.1 μM) induced cellular differentiation, which was followed after 48 h by a tenfold increase in chemiluminescence from lucigenin and a 2.5-fold increase in the intracellular oxidation of 2′,7′-dicholorofluorescin (DCFH). Whereas higher concentrations (1.0 μM) of TPA did not stimulate further ROS production, repeated stimulation with 0.1 μM TPA of differentiated cells induced a modest (1.2-fold) but rapid (15 min) increase in chemiluminescence. In cells treated with TPA, the burst in ROS at 48 h was preceded by accumulation at 12 h of gp91phox (8.8-fold) and p47phox mRNA (threefold), whereas untreated cells contained steady-state levels of both transcripts. Time-course experiments with actinomycin D to inhibit transcription revealed that TPA did not improve the stability of gp91phox. In transient transfections, luciferase reporter activity directed from a 1.5-kb gp91phox promoter fragment was enhanced threefold upon treatment with TPA for 24 h. We conclude that TPA can commit HL-60 cells to differentiation and elicit transcription from the proximal gp91phox promoter.
AB - The exposure to epigenetic effectors capable of inducing copious production of reactive oxygen species (ROS) has been associated with chronic inflammation, tumor initiation, and promotion. The objective of this study was to examine the regulation of gp91phox, the catalytic subunit of the NADPH oxidase, and the kinetics of ROS production in promyelocytic leukemia HL-60 cells induced with 12-O-tetradeconylphorbol-13-acetate (TPA). The treatment of HL-60 cells with TPA (0.1 μM) induced cellular differentiation, which was followed after 48 h by a tenfold increase in chemiluminescence from lucigenin and a 2.5-fold increase in the intracellular oxidation of 2′,7′-dicholorofluorescin (DCFH). Whereas higher concentrations (1.0 μM) of TPA did not stimulate further ROS production, repeated stimulation with 0.1 μM TPA of differentiated cells induced a modest (1.2-fold) but rapid (15 min) increase in chemiluminescence. In cells treated with TPA, the burst in ROS at 48 h was preceded by accumulation at 12 h of gp91phox (8.8-fold) and p47phox mRNA (threefold), whereas untreated cells contained steady-state levels of both transcripts. Time-course experiments with actinomycin D to inhibit transcription revealed that TPA did not improve the stability of gp91phox. In transient transfections, luciferase reporter activity directed from a 1.5-kb gp91phox promoter fragment was enhanced threefold upon treatment with TPA for 24 h. We conclude that TPA can commit HL-60 cells to differentiation and elicit transcription from the proximal gp91phox promoter.
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M3 - Article
C2 - 11200061
AN - SCOPUS:0035155135
SN - 0741-5400
VL - 69
SP - 161
EP - 168
JO - Journal of Leukocyte Biology
JF - Journal of Leukocyte Biology
IS - 1
ER -