No abstract

TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) is the most potent tumor promoter ever tested in rodents. Although it is known that most of the effects of TCDD are mediated by binding to the aryl hydrocarbon receptor (AHR), the mechanisms leading to tumor promotion still remain to be elucidated. Loss of contact-inhibition is a characteristic hallmark in tumorigenesis. In WB-F344 cells, TCDD induces a release from contact-inhibition manifested by a 2-to 3-fold increase in DNA-synthesis and the emergence of foci when TCDD (1 nM) is given to confluent cells. We focussed our interest on potential cell membrane proteins mediating contact-inhibition in WB-F344 cells, namely E-cadherin, ␣,-␤,-and ␥-catenin (plakoglobin). Using indirect immunofluorescence, E-cadherin, ␣-, ␤-and ␥-catenin were detected at cell adhesion sites in untreated, confluent cells. After TCDD-exposure, ␥-catenin was exclusively localized in the cytoplasm whereas localization of E-cadherin, ␣-and ␤-catenin remained unaffected. Cytoplasmic ␥-catenin could be extracted by Triton X-100 treatment, demonstrating that ␥-catenin was no longer bound to the actin cytoskeleton. Western blot analysis showed downregulation of ␥-catenin protein levels. This effect was not blocked by pre-incubation with the selective proteasome inhibitor MG-132, indicating that proteolytical degradation of ␥-catenin by the proteasome system was not increased by TCDD. Because mRNA-levels of ␥-catenin were markedly diminished after TCDD-exposure, we conclude that transcriptional downregulation or destabilization of the mRNA contributes to the decrease in ␥-catenin protein levels in response to TCDD. Because ␥-catenin is considered to be a tumor suppressor, our findings might give more insight into the tumor promoting actions of TCDD. © 2002 Wiley-Liss, Inc. Key words: TCDD; contact-inhibition; ␥-catenin; liver tumor promotion2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a widespread environmental contaminant causing a variety of species-and organ-specific biological effects. Epidemiological studies indicate that TCDD is a human carcinogen 1 classified as Group 1 carcinogen by the International Agency for Research on Cancer. 2 In rodent bioassays, TCDD is the most powerful tumor promoter ever tested. 3 One hallmark of TCDD action is tumor promotion in 2-stage rat-liver carcinogenesis. The underlying mechanisms of liver tumor promotion by TCDD, however, still remain to be elucidated.Loss of contact-inhibition is 1 characteristic feature of transformed cells. In WB-F344 cells, 4 TCDD induces a release from contact-inhibition. [5][6][7] This is manifested by a 2-fold increase in cell number when TCDD is given to confluent, but not to exponentially growing or serum-deprived WB-F344 cells. [5][6][7] We have demonstrated recently that TCDD-treatment of confluent cells leads to an increase in cyclin A protein levels, which shows integration of the TCDD-response in the cell cycle regulatory machinery and explains loss of G1-arrest. 7 Up-stream effects of TCDD, especially on cell-adhesion molecu...