Indole-3-carbinol (13C), present in cruciferous vegetables, and its major in vivo product 3,3'-diindolylmethane (DIM), have been reported to suppress cancer development. However, the responsible molecular mechanisms are not fully understood. Nonsteroidal anti-inflaminatory drug-activated gene-1 (NAG-1) is a TGF-beta superfamily gene associated with pro-apoptotic and anti-tumorigenic activities. The present study was performed to investigate whether 13C and DIM influence NAG-1 expression and to provide the potential molecular mechanism of their effects oil anti-tumorigenesis. The 13C repressed cell proliferation and induced NAG-1 expression in a concentration-dependent manner. In addition, DIM increased the expression of NAG-1 as well as activating transcription factor 3 (ATF3), and the induction of ATF3 was earlier than that of NAG-1. The DIM treatment increased luciferase activity of NAG-1 in HCT-116 cells transfected with NAG-1 promoter construct. The results suggest that 13C represses cell proliferation through up-regulation of NAG-1 and that ATF3 may play a pivotal role in DIM-induced NAG-1 expression ill human colorectal cancer cells. Furthermore, the mixture of 13C with resveratrol enhances NAG-1 expression, suggesting the synergistic effect of these two unrelated compounds on NAG-1 expression. (C) 2005 Elsevier Inc. All rights reserved.