Inhibition of cyclin-dependent kinase 2 by p21 is necessary for retinoblastoma protein-mediated G(1) arrest after gamma-irradiation

In mammalian cells, activation of certain checkpoint pathways as a result o f exposure to genotoxic agents results in cell cycle arrest. The integrity of these arrest pathways is critical to the ability of the cell to repair m utations that otherwise might compromise viability or contribute to deregul ation of cellular growth and proliferation, Here we examine the mechanism t hrough which DNA damaging agents result in a G(1) arrest that depends on th e tumor suppressor p53 and its transcriptional target p21. By using primary cell lines lacking specific cell cycle regulators, we demonstrate that thi s pathway functions through the growth suppressive properties of the retino blastoma protein (pRB) tumor suppressor. Specifically, gamma-irradiation in hibits the phosphorylation of pRB at cyclin-dependent kinase 2-specific, bu t not cyclin-dependent kinase 4-specific, sites in a p21-dependent manner. Most importantly, we show that pRB is a critical component of this DNA dama ge checkpoint. These data indicate that the p53 --> p21 checkpoint pathway uses the normal cell cycle regulatory machinery to induce the accumulation of the growth suppressive form of pRB and suggest that loss of pRB during t he course of tumorigenesis disrupts the function of an important DNA damage checkpoint.