Inhibitory effects of 1α,25-dihydroxyvitamin D3 on the G1-S phase-controlling machinery

The nuclear hormone 1 alpha ,25-dihydroxyvitamin D-3 induces cell cycle arrest, differentiation, or apoptosis depending on target cell type and state. Although the antiproliferative effect of 1 alpha ,25-dihydroxyvitamin D-3 has been known for years, the molecular basis of the cell cycle blockade by 1 alpha ,25-dihydroxyvitamin D-3 remains largely unknown. Here we have investigated the mechanisms underlying the G(1) arrest induced upon 1 alpha ,25-dihydroxyvitamin D-3 treatment of the human breast cancer cell line MCF-7. Twenty-four-hour exposure of exponentially growing MCF-7 cells to 1 alpha ,25-dihydroxyvitamin D-3 impeded proliferation by preventing S phase entry, an effect that correlated with appearance of the growth-suppressing, hypophosphorylated form of the retinoblastoma protein (pRb), and modulation of cyclin-dependent kinase (cdk) activities of cdk-4, -6, and -2. Time course immunochemical and biochemical analyses of the cellular and molecular effects of 1 alpha ,25-dihydroxyvitamin D-3 treatment for up to 6 d revealed a dynamic chain of events, preventing activation of cyclin D1/cdk4, and loss of cyclin D3, which collectively lead to repression of the E2F transcription factors and thus negatively affected cyclin A protein expression. While the observed 10-fold inhibition of cyclin D1/cdk 4-associated kinase activity appeared independent of cdk inhibitors, the activity of cdk 2 decreased about 20-fold, reflecting joint effects of the lower abundance of its cyclin partners and a significant increase of the cdk inhibitor p21(CIP1/)WAF1, which blocked the remaining cyclin A(E)/cdk 2 complexes. Together with a rapid down-modulation of the c-Myc oncoprotein in response to 1 alpha ,25-dihydroxyvitamin D-3, these results demonstrate that 1 alpha ,25-dihydroxyvitamin D-3 inhibits cell proliferation by targeting several key regulators governing the G(1)/S transition.