Transcriptional regulation of gene expression represents a means of control or many fundamental cellular processes, such as cell growth and differentiation, and for responses to endogenous and exogenous signals, such as the adaptive/toxic responses to environmental chemicals. As such, transcription factors that mediate gene expression to specific signals are often tightly regulated to ensure physiologically adequate gene transcription and thus, the homeostasis of the cell. The ubiquitin-26S proteasome-mediated proteolysis has been implicated in the regulation of various types of cellular proteins. In this study, we analyzed the mechanism of agonist-induced degradation of the Ah receptor and its functional impact on AhR-mediated gene transcription to 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD). Biochemical and genetic data reveal that TCDD incudes ubiquitination of AhR and shortening of the half-life of the protein through the 26S-proteasome mediated protein degradation. The agonist induced AhR degradation can be blocked by using inhibitors of the 26S proteasomes (MG123 and lactacystin) or cycloheximide, a potent inhibitor of protein synthesis; these data implicate a labile factor in controlling the degradation of AhR, which we designated Ah receptor degradation promoting factor (ADPF). Furthermore, inhibition of the AhR degradation by inhibitors of the 26S proteasomes or protein synthesis markedly enhances the induction of CYP1A1 by TCDD, a phenomenon termed superinduction; these findings suggest that the agonist-induced, ADPF-mediated degradation of AhR serves as a mechanism by which gene transcription by AhR is nagatively controlled in cells. This view is suppored by the observation that ingibition of the AhR degradation by either MG132 or cycloheximide superinduces a number of other AhR target genes, including TiPARP, a novel TCDD-inducible poly(ADP-ribose) polymerase. Our findings provide new insights into the control of the activity of agonist activated AhR through a regulated, proteasomal protein degradation pathway. Our current research is aimed at cloning of ADPF.
The Toxicologist. Society of Toxicology 41st Annual Meeting and ToxExpo, March 17-21, 2002, Nashville, Tennessee