Nuclear factor (NF)-kappaB/Rel transcription factors normally exist in non-B cells, such as epithelial cells, in inactive forms sequestered in the cytoplasm with specific inhibitory proteins, termed IkappaBs. Recently, however, we discovered that breast cancer is typified by aberrant constitutive expression of NF-kappaB/Rel factors. Because these factors control genes that regulate cell proliferation, here we analyzed the potential role of NF-kappaB/Rel in the ability of transforming growth factor (TGF)-beta1 to inhibit the growth of breast cancer cells. The decreased growth of Hs578T and MCF7 breast cancer cell lines on TGF-beta1 treatment correlated with a drop in NF-kappaB/Rel binding. This decrease was due to the stabilization of the inhibitory protein IKB-alpha. Ectopic expression of c-Rel in Hs578T cells led to the maintenance of NF-kappaB/Rel binding and resistance to TGF-beta1-mediated inhibition of proliferation. Similarly, expression of the p65 subunit ablated the inhibition of Hs578T cell growth mediated by TGF-beta1. Thus, the inhibition of the aberrantly activated, constitutive NF-kappaB/Rel plays an important role in the arrest of the proliferation of breast cancer cells, which suggests that NF-kappaB/Rel may be a useful target in the treatment of breast cancer.