Activation of immediate‐early gene expression by peroxisome proliferators in vitro

Abstract

The hepatocarcinogenicity of peroxisome proliferators (PPs) in rodents has been attributed both to oxidative DNA damage resulting from excessive leakage of peroxisomal H2O2 and to increased hepatocellular replication that may be independent of peroxisome proliferation. Because of the growing association between tumor promotion and alterations in growth-regulatory signal transduction pathways, we investigated whether PPs can modulate these pathways in a mouse liver epithelial cell line, BNL-CL.2. We tested two PPs that differ markedly in rodent tumorigenicity for their ability to activate immediate-early proto-oncogene expression. 4-Chloro-6-(2,3-xylidino)-2-pyrimidinylthioacetic acid (Wy-14643), a highly tumorigenic PP, was an exceptionally strong inducer of c-fos expression. Wy-14643 was also stronger than DEHP in stimulating c-jun expression, whereas both PPs were fairly strong inducers of jun-B and jun-D. The induction of fos and jun expression by Wy-14643 was specifically inhibited by the protein kinase C inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperizine dihydrochloride (H-7). DEHP-induced gene expression was strongly inhibited by H-7, but was also partially inhibited by an inhibitor of protein kinase A. The activation of fos and jun gene expression by PPs was independent of peroxisome proliferation since it was an immediately-early response not requiring protein synthesis and since the cell lines used in this study do not undergo peroxisome proliferation. Our r results raise the possibility that the carcinogenicity of PPs may be due, in part, to epigenetic modulation of growth-regulatory signal transduction pathways.

Peroxisome Proliferators Activate Extracellular Signal-regulated Kinases in Immortalized Mouse Liver Cells

“…In addition, some PPs have been shown to induce the expression of immediate-early genes, including c-fos, c-jun, junB, and egr-1, and to modulate cell proliferation in mouse liver cell lines (17,18). In this study, we show that several of the PPs activate the MAPK family members ERK1 and ERK2 in an immortalized mouse liver cell line.…”

“…This concentration causes optimal induction of immediate-early gene expression while avoiding toxicity (17,18). Western blotting of whole cell lysates with anti-phosphotyrosine antibodies showed a transient increase in tyrosine phosphorylation of two proteins in response to Wy-14,643 (Fig.…”

“…Activation of the ERKs leads to the phosphorylation of a variety of substrates including transcription factors (28,29) that ultimately regulate gene expression. Induction of immediate-early gene expression by the PPs has been shown previously (17,18). To determine if activation of ERK1 and ERK2 by the PPs is responsible for their induction of immediate-early gene expression, quiescent cells were preincubated with PD98059 followed by PP stimulation and RNA isolation.…”

Induction of Cyclooxygenase-2 Expression by Peroxisome Proliferators and Non-tetradecanoylphorbol 12,13-Myristate-type Tumor Promoters in Immortalized Mouse Liver Cells

“…At doses comparable to those required for COX-2 expression, PPs induce other immediateearly genes, including the proto-oncogenes c-fos, c-jun, junB, and egr-1 (35,36), they mobilize intracellular Ca 2ϩ (52), and they regulate cell-cycle progression (36). Recently, we found that PPs activate the Erk1 and Erk2 mitogen-activated protein kinases with a similar dose-response.…”

“…PPs have been shown to enhance proto-oncogene expression (31)(32)(33) and protein kinase C activity (34) in vivo. In mouse liver cell lines in vitro, PPs are potent activators of signal transduction pathways involving mitogen-activated protein kinases, 2 they strongly induce the expression of immediate-early proto-oncogenes (35,36), and they regulate cell cycle progression during both early G 1 and late G 1 (36). These growth regulatory effects of PPs in vitro are independent of peroxisome proliferation (35,36).…”