P21^Cip1 induced by Raf is associated with increased Cdk4 activity in hematopoietic cells

To investigate the functions of the different Raf genes in hematopoietic cell proliferation, the capacities of β-estradiol-regulated ΔRaf:ER genes to induce cell cycle regulatory gene expression and cell cycle progression in FDC-P1 cells were examined. Raf activation increased the expression of Cdk2, Cdk4, cyclin A, cyclin D, cyclin E, p21^Cip1 and c-Myc and decreased the expression of p27^Kip1 which are associated with G₁ progression. However only the cell clones with moderate Raf activation, i.e. FD/ΔRaf-1:ER and FD/ΔA-Raf:ER, successfully underwent cell proliferation. The cell clones with the highest ΔRaf activity, FD/ΔB-Raf:ER, underwent apoptosis before cell proliferation. p21^Cip1 induced by Raf activation specifically bound with Cdk4/cyclin D complexes but not Cdk2/cyclin E complexes and this binding was associated with the increased Cdk4 activity. However, no binding of p27^Kip1 with either Cdk2/cyclin E or Cdk4/cyclin D was observed. Thus Raf mediated growth was associated with elevated p21^Cip1 expression, which may specifically bind with and activate Cdk4/cyclin D complexes and with decreased p27^Kip1 expression.