Knowledge regarding the expression of the recently cloned estrogen receptor β (ERβ) in colonic mucosa is limited. In this study, we demonstrated that five human colon cancer cell lines, HT29, Colo320, Lovo, SW480, and HCT116, expressed ERβ mRNA, but lacked ERα mRNA. Results from a cell growth assay demonstrated that these colon cancer cells were not influenced by estrogen, while genistein possessed slight growth inhibitory effects on HT29, Colo320 and Lovo cells at 10 μM, at which concentration is stimulated the growth of ERα-positive human breast cancer MCF-7 cells. Tamoxifen inhibited the growth of HT29 and Colo320 cells, dose-dependently, as well as MCF-7 cells. A transfected reporter plasmid containing a vitellogenin estrogen response element could be activated by estradiol in Colo320 cells. Taken together with previous reports, these data suggest that ERα and ERβ may have different biological functions in colon cells.