Mechanism of benzo(a)pyrene induction of alpha-human chorionic gonadotropin gene expression in human lung tumor cells.

Abstract

Human lung cells (ChaGo) derived from a bronchogenic carcinoma synthesize and secrete in the culture medium the alpha subunit of the glycoprotein hormone, human chorionic gonadotropin (alpha-hCG). The synthesis of alpha-hCG by ChaGo cells could be further stimulated by treatment with sublethal concentrations of the polycyclic aromatic hydrocarbons (PAHs), benzo(a)pyrene (gaP), or dimethylbenzanthracene. The production of alpha-hCG could be correlated to the levels of alpha-hCG-specific mRNA sequences in control and PAH-treated cells. Further analysis of the RNA species (Northern blot) revealed that the level of the mature (~1.0 kb) and the high molecular weight alpha-hCG specific nuclear RNA sequences (~2.2 and 5 kb) were all greater in PAH-treated cells.Addition of [3H]BaP (0.25 #g/ml) in the culture medium of ChaGo cells led to immediate uptake of the radioactive compound apparently by simple diffusion. SDS PAGE and subsequent fluorography revealed that the radioactive compound interacted and formed covalent complexes with cytoplasmic and nuclear proteins. This covalent interaction of the [3H]BaP molecule with cellular proteins could be significantly inhibited by either inhibiting the activity of the enzyme aryl hydrocarbon hydroxylase with 7,8-benzoflavone or by reducing the cellular concentration of the enzyme by simultaneous incubation with cycloheximide. These results suggested that in ChaGo cells, the observed covalent complexes were formed by the interaction of the BaP metabolites with cellular proteins. The concentrations at which 7,8-benzoflavone or cycloheximide inhibited (a) formation of metabolites from [3H]BaP and (b) their covalent interaction with cell protein did not affect the BaP-induced stimulation of alphahCG gene expression. However, the cytotoxic effects of BaP in ChaGo cells seemed to be exerted by the metabolism of the compounds. Results presented in this report suggest that gap metabolism and the interaction of the metabolites with cell proteins were not essential for the BaP-induced modulation of alpha-hCG gene expression.Polycyclic aromatic hydrocarbons (PAHs) ~ are currently recognized as one of the major classes of environmental carcinogenic pollutants (1). Of the PAHs, benzo(a)pyrene (BaP) is Abbreviations used in this paper: AHH, aryl hydrocarbon hydroxylase; BaP benzo(a)pyrene; BF, 7,3-benzoflavone; ChaGo cells, human lung bronchogenic carcinoma cells; DMBA, dimethylbenzanthracene; alpha-hCG, the alpha subunit of human chorionic gonadotropin; alpha-hCG mRNA, alpha-hCG-specific mRNA sequences; PAHs, polycyclic aromatic hydrocarbons; SSC, 3 M NaC! and 0.3 M sodium acetate. accepted as the prototype (2, 3). Ectopic hormone and inappropriate gene expression are commonly found to be associated with neoplastic transformation of cells. BaP and several other members of the PAHs have been found to induce aryl hydrocarbon hydroxylase (AHH) in different mammalian cell systems (4). These inducers of AHH also enhance the expression of alpha-fetoprotein gene (5); activities of phospho...

Mechanism of benzo(a)pyrene induction of alpha-human chorionic gonadotropin gene expression in human lung tumor cells

“…1 C demonstrate that of high molecular weight alpha-hCG-specific nuclear RNA sequences between control and PAH-treated cells. The observed increased levels of alpha-hCG mRNA sequences in BaP or DMBA-treated ChaGo cells detected by Northern blot analysis not only confirm the previously reported results obtained by dot hybridization analysis (14), but also characterize the molecular sizes of alpha-hCG-specific cytoplasmic and nuclear RNA transcripts. The increased level of alpha-hCG production in PAH-treated cells can thus be correlated to the corresponding elevated levels of different species of cytoplasmic and nuclear alpha-hCG mRNA sequences.…”

“…As stated above, either BaP or DMBA treatment of ChaGo cells increased the levels of cytoplasmic alpha-hCG mRNA sequences. We have previously reported that DMBA stimulation of alpha-hCG synthesis was somewhat higher than that observed after BaP treatment of ChaGo cells (14). This was further substantiated by the results presented in It is believed that the cytotoxic effects of chemical carcinogens are due to their genotoxic/DNA damaging effects caused by reactive metabolites of BaP.…”

“…This was further substantiated by the results presented in It is believed that the cytotoxic effects of chemical carcinogens are due to their genotoxic/DNA damaging effects caused by reactive metabolites of BaP. We have previously reported that BaP is cytotoxic to ChaGo cells at concentrations of these chemicals of 0.3 #g/ml and higher (14). The possible role of BaP metabolism in the cytotoxic effect of the chemical was verified by growing the cells in the presence of BaP with and without the metabolic inhibitor BF.…”

“…We have previously reported that BaP is able to stimulate the level of expression of an ectopically produced giycoprotein hormone, the alpha subunit of human chorionic gonadotropin (alpha-hCG) in an established human lung bronchogenic carcinoma cell line (ChaGo) under culture conditions (14). A correlation has been established between the increased levels of alpha-hCG-specific mRNA sequences (alpha-hCG mRNA) and BaP-induced hypomethylation of the alpha-hCG gene (14).…”

“…A correlation has been established between the increased levels of alpha-hCG-specific mRNA sequences (alpha-hCG mRNA) and BaP-induced hypomethylation of the alpha-hCG gene (14). In this part of the investigation we examined the following parameters: (a) the levels of different species of nuclear and cytoplasmic alpha-hCG mRNA in control and BaP-treated cells; (b) the pattern of BaP metabolism in ChaGo cells; (c) the possible role of the metabolites of BaP on the induction of alpha-hCG gene expression, and fnally (d) the cytotoxic effect of BaP metabolites in ChaGo cells.…”