As part of a comprehensive survey of the impact of the environmental pollutant and hepatocarcinogen 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the proteome of hepatic cells, we have performed a high resolution two-dimensional gel electrophoresis study on the rat hepatoma cell line 5L. 78 protein species corresponding to 73 different proteins were identified as up- or down-regulated following exposure of the cells to 1 nm TCDD for 8 h. There was an overlap of only nine proteins with those detected as altered by TCDD in our recent study using the non-gel-based isotope-coded protein label method (Sarioglu, H., Brandner, S., Jacobsen, C., Meindl, T., Schmidt, A., Kellermann, J., Lottspeich, F., and Andrae, U. (2006) Quantitative analysis of 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced proteome alterations in 5L rat hepatoma cells using isotope-coded protein labels. Proteomics 6, 2407-2421) indicating a strong complementarity of the two approaches. For the majority of the altered proteins, an effect of TCDD on their abundance or posttranslational modifications had not been known before. Several observations suggest that a sizable fraction of the proteins with altered abundance was induced as an adaptive response to TCDD-induced oxidative stress that was demonstrated using the fluorescent probe dihydrorhodamine 123. A prominent group of these proteins comprised various enzymes for which there is evidence that their expression is regulated via the Keap1/Nrf2/antioxidant response element pathway. Other proteins included several involved in the maintenance of mitochondrial energy production and the regulation of the mitochondrial apoptotic pathway. A particularly intriguing finding was the up-regulation of the mitochondrial outer membrane pore protein, voltage-dependent anion channel-selective protein 2 (VDAC2), which was dependent on the presence of a functional aryl hydrocarbon receptor. The regulatability of VDAC2 protein abundance has not been described previously. In view of the recently discovered central role of VDAC2 as an inhibitor of the activation of the proapoptotic protein BAK and the mitochondrial apoptotic pathway, the present data point to a hitherto unrecognized mechanism by which TCDD may affect cellular homeostasis and survival.