Mechanisms of AH receptor-mediated responses

Abstract

The interaction of the nuclear aryl hydrocarbon (Ah) receptor complex with c-fos and c-jun oncoprotein was investigated in wild-type Ah responsive mouse Hepa lclc7 cells. Chloramphenicol acetyl transferase (CAT) activity was induced by 10 riM 2,3,7,8tetrachlorodibenzo-p-dioxin (TCDD) in Hepa lclc7 cells transfected with an Ah-responsive plasmid containing a dioxin-responsive element (DRE) and a bacterial CAT reporter gene. In cells cotranfected with the DRE-CAT construct and a c-fos expression plasmid, there was an increase in basal CAT activity however Ah-responsiveness was unchanged. In contrast, cotransfection of Hepa lclc7 cells with the DRE-CAT construct and a c-jun expression plasmid had no significant effect on basal or inducible CAT activity. Cotransfection of Hepa 1 c I c7 cells with PRNH I I c Ah-responsive plasmid containing the 1142 to-2434 region of the CYPIAI gene and a c-fos expression plasmid also had minimal effect of Ah-responsiveness. Ah-and E2-responsiveness was investigated in five human osteosarcoma cells namely U-2, G292, HOS(TE85), MG-63 and SAOS-2. The cells was transiently transfected with Ah-responsive PRNH II c and CAT activity was not induced. Cells were also transiently transfected the estrogen-responsive VIT-TK-CAT plasmid, and only minimal induction of CAT activity were observed in some bone cells. Nfinimal estrogen-and Ah-responsiveness was observed in these cell lines although relatively high levels of the nuclear Ah-receptor complex was detected in MG-63 cells. Bcl-2 is a key regulator in cell apoptotic pathways, and bcl-2 gene expression by estrogen in ER' breast cancer cell lines. The effects of TCDD on E2-mediated bcl-2 gene expression were investigated in this study. The E2-induced bcl-2 niRNA levels were inhibited by TCDD in MCF-7 cells using RT-PCR method. Ten nM TCDD caused a significant inhibition on E2-induced CAT activity in T47D cells transiently transfected with plasmids containing the bcl-2 promoter subcloned PUCSVOCAT in T47D cells.