A pure estrogen antagonist inhibits cyclin E-Cdk2 activity in MCF-7 breastcancer cells and induces accumulation of p130-E2F4 complexes characteristic of quiescence

Estrogen antagonists inhibit cell cycle progression in estrogen-responsive cells, but the molecular mechanisms are not fully defined. Antiestrogen-med iated G(0)/G(1) arrest is associated with decreased cyclin D1 gene expressi on, inactivation of cyclin D1-cyclin dependent kinase (Cdk) 4 complexes, an d decreased phosphorylation of the retinoblastoma protein (pRb). We now sho w that treatment of MCF-7 breast cancer cells with the pure estrogen antago nist ICI 182780 results in inhibition of cyclin E-Cdk2 activity prior to a decrease in the G(1) to S phase transition. This decrease was dependent on p21(WAF1/Cip1) since treatment with antisense oligonucleotides to p21 atten uated the effect, Recruitment of p21 to cyclin E-Cdk2 complexes was in turn dependent on decreased cyclin D1 expression since it was apparent followin g treatment with antisense cyclin D1 oligonucleotides, To define where with in the G(0) to S phase continuum antiestrogen-treated cells arrested, we as sessed the relative abundance and phosphorylation state of pocket protein-E 2F complexes. While both pRb and p107 levels were significantly decreased, p130 was increased 4-fold and was accompanied by the formation of p130.E2F4 complexes and the accumulation of hyperphophorylated E2F4, putative marker s of cellular quiescence. Thus, ICI 182780 inhibits both cyclin D1-Cdk4 and cyclin E-Cdk2 activity, resulting in the arrest of MCF-7 cells in a state with characteristics of quiescence (G(0)), as opposed to G(1) arrest.