Calmodulin is essential for cyclin-dependent kinase 4 (Cdk4) activity and nuclear accumulation of cyclin D1-Cdk4 during G(1)

Although it is known that calmodulin is involved in G(1) progression, the c almodulin-dependent G(1) events are not well understood. We have analyzed h ere the role of calmodulin in the activity, the expression, and the intrace llular location of proteins involved in G(1) progression. The addition of a nti-calmodulin drugs to normal rat kidney cells in early G(1) inhibited cyc lin-dependent kinase 4 (Cdk4) and Cdk2 activities, as well as retinoblastom a protein phosphorylation. Protein levels of cdk4, cyclin D1, cyclin D2, cy clin E, p21, and p27 were not affected after CaM inhibition, whereas decrea ses in the amount of cyclin A and Cdc2 were observed. The decrease of Cdk4 activity was due neither to changes in its association to cyclin D1 nor to changes in the amount of p21 or p27 bound to cyclin D1-Cdk4 complexes. Calm odulin inhibition also produced a translocation of nuclear cyclin D1 and Cd k4 to the cytoplasm.. This translocation could be responsible for the decre ased Cdk4 activity upon calmodulin inhibition, Immuno:precipitation, calmod ulin affinity chromatography, and direct binding experiments indicated that calmodulin associates with Cdk4 and cyclin D1 through a calmodulin-binding protein. The facts that Hsp90 interacts with Cdk4 and that its inhibition induced Cdk4 and cyclin D1 translocation to the cytoplasm point to Hsp90 as a good candidate for being the calmodulin-binding protein involved in the nuclear accumulation of Cdk4 and cyclin D1.