Reduction in cyclin D1/Cdk4/retinoblastoma protein signaling by CRE-decoy oligonucleotide

We have previously demonstrated that the activation of p53 signaling may co ntribute to tumor growth inhibition by the CRE-decoy oligonucleotide contai ning CRE sequence (5'-TGACGTCA-3') (Lee et al., Biochemistry 39, 4863-4868, 2000). However, growth inhibition by CRE-decoy treatment was also observed in tumor cells containing a mutant p53 (Park et al., J. Biol. Chem. 274, 1 573-1580, 1999), To understand additional mechanisms of the decoy oligonucl eotide, we investigated the effect on cyclin D1 expression and a cyclin D1/ Cdk4/retinoblastoma protein (pRB) signaling pathway. Here we show that in M CF7 breast cancer cells the CRE-decoy competed with cyclin D1-CRE (5'-T (A) under bar ACGTCA-3') for binding transcription factors and reduced cyclin D1 gene expression (in reporter gene assay, Northern blotting and Western b lotting) to modulate cyclin D1/Cdk4/pRB signaling and G1-S progression in a steady state and/or under estrogen stimulation. Decrease of cyclin D1 prot ein level by CRE-decoy treatment was also observed in p53-mutated cancer ce lls. Cyclin D1 expression was also diminished in MCF7 cells stably expressi ng dominant negative mutant CREB indicating that the nonspecific effect of oligonucleotide or its degradation products could be excluded. These data s uggest that inhibition of cyclin D1 expression contributes to the growth in hibition induced by the decoy oligonucleotide in MCF7 cells through a cycli n D1/Cdk4/pRB signaling pathway. Downregulation of cyclin D1 expression als o provides a mechanism of CRE-decoy-induced growth inhibition in tumor cell s having p53 mutation. (C) 2001 Academic Press.