Abstract Cancer cells have a consistent cytological feature of nucleolar hypertrophy, enlarged nucleoli, where RNA polymerase III genes (Pol III genes) are transcripted. This implies that transformation in situ is closely linked to the deregulation of Pol III gene transcription, because the size of the nucleolus reflects the levels of rRNA synthesis. Deregulation of Pol III genes is tightly associated cell transformation and tumorgenesis. However, very little is known about whether alcohol consumption affects Pol III gene transcription. By using cell culture model and animal model, we have found, for the first time that alcohol induces Pol III gene transcription in JNK1 and ADH dependent manner. Alcohol increases c-Jun expression in HepG2-ADH cells. To determine whether the alcohol-mediated increases c-jun required to stimulate TBP and Brf1 expression are due to enhance occupancy of c-jun at their promoters, chromatin immunoprecipitation (ChIP) assays were performed. Ethanol induces a marked increase in the recruitment of c-jun to the TBP promoter, while a modest increase in the occupancy of both c-fos and Elk-1 was observed. These results support the idea that alcohol mediates an increase in TBP expression primarily through enhanced expression of c-jun and its recruitment to the TBP promoter. Sequence analysis reveals both putative AP-1 and Elk-1 binding sites in regions 5’ of the transcription start site of the Brf1 gene. ChIP analysis reveals that ethanol induces an increase in the occupancy of c-jun to the region containing an AP-1 binding site, while no significant increase in binding of Elk-1 to the Elk-1 binding site is observed. Interestingly, c-jun, c-fos and Elk-1 are also found to occupy the tRNALeu gene. Upon ethanol treatment, an increase in TBP recruitment to the tRNALeu gene is observed, as well as a marked increase in the occupancy of c-jun. However, alcohol does not induce any apparent change in recruitment of either c-fos or Elk-1 to the tRNA gene. These results suggest that c-jun may function directly at sequences near tRNA genes to induce transcription. Together, these results support the idea that alcohol enhances expression of TBP, Brf1, and tRNA gene by facilitating c-jun recruitment to these promoters. *: This project was supported by NIH grant AA017288 to S.Z. **: Corresponding author: szhong@usc.edu. Tel: 626-628-7693 or 323-442-114 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3685. doi:10.1158/1538-7445.AM2011-3685