Indole-3-Carbinol and 3-3′-Diindolylmethane Antiproliferative Signaling Pathways Control Cell-Cycle Gene Transcription in Human Breast Cancer Cells by Regulating Promoter–Sp1 Transcription Factor Interactions

Gary L. Firestone, Leonard F. Bjeldanes
2003 Journal of Nutrition  
Indole-3-carbinol (I3C), a compound that occurs naturally in Brassica vegetables such as cabbage and broccoli, can induce a G1 cell-cycle arrest of human MCF-7 breast cancer cells that is accompanied by the selective inhibition of cyclin-dependent kinase 6 (Cdk6) expression and stimulation of p21 Waf1/Cip1 gene expression. Construction and transfection of a series of promoter-reporter plasmids demonstrate that the indole-regulated changes in Cdk6 and p21 Waf1/Cip1 levels are due to specific
more » ... cts on their corresponding promoters. Mutagenic analysis reveals that I3C signaling targets a composite transcriptional element in the Cdk6 promoter that requires both Sp1 and Ets transcription factors for transactivation function. Analysis of protein-DNA complexes formed with nuclear proteins isolated from I3C-treated and -untreated cells demonstrates that the Sp1 DNA element in the Cdk6 promoter interacts with an I3C-inhibited protein-protein complex that contains the Sp1 transcription factor. In indoletreated cells, a fraction of [ 3 H]I3C was converted into its natural diindole product 3 H-labeled 3-39-diindolylmethane ([ 3 H]DIM), which accumulates in the nucleus; this suggests that DIM may have a role in the transcriptional activities of I3C. Mutagenic analysis of the p21 Waf1/Cip1 promoter reveals that in transfected breast cancer cells, DIM (as well as I3C) stimulates p21 Waf1/Cip1 transcription through an indole-responsive region of the promoter that contains multiple Sp1 consensus sequences. Furthermore, DIM treatment regulates the presence of a nuclear Sp1 DNA-binding activity. Our results demonstrate that both the Cdk6 and p21 Waf1/Cip1 promoters are newly defined downstream targets of the indole-signaling pathway, and that the observed transcriptional effects are due to a combination of the cellular activities of I3C and DIM. J. Nutr. 133: 2448S-2455S, 2003. KEY WORDS: indoles I3C DIM antiproliferative pathway regulated promoter cell-cycle gene Sp1 transcription factor reproductive cancer cell 1 Published in a supplement
doi:10.1093/jn/133.7.2448s pmid:12840223 fatcat:7pzmanmszzgxbciwlr7zpvzv6a