p16INK4A participates in a G1 arrest checkpoint in response to DNA damage.
Members of the INK4 protein family specifically inhibit cyclin-dependent kinase 4 (cdk4) and cdk6-mediated phosphorylation of the retinoblastoma susceptibility gene product (Rb). p16INK4A, a prototypic INK4 protein, has been identified as a tumor suppressor in many human cancers. Inactivation of p16INK4A in tumors expressing wild-type Rb is thought to be required in order for many malignant cell types to enter S phase efficiently or to escape senescence. Here, we demonstrate another mechanism of tumor suppression by implicating p16INK4A in a G1 arrest checkpoint in response to DNA damage. Calu-1 non-small cell lung cancer cells, which retain Rb and lack p53, do not arrest in G1 following DNA damage. However, engineered expression of p16INK4A at levels compatible with cell proliferation restores a G1 arrest checkpoint in response to treatment with gamma-irradiation, topoisomerase I and II inhibitors, and cisplatin. A similar checkpoint can be demonstrated in p53-/- fibroblasts that express p16INK4A. DNA damage-induced G1 arrest, which requires the expression of pocket proteins such as Rb, can be abrogated by overexpression of cdk4, kinase-inactive cdk4 variants capable of sequestering p16INK4A, or a cdk4 variant incapable of binding p16INK4A. After exposure to DNA-damaging agents, there was no change either in overall levels of p16INK4A or in amounts of p16INK4A found in complex with cdks 4 and 6. Nonetheless, p16INK4A expression is required for the reduction in cdk4- and cdk6-mediated Rb kinase activity observed in response to DNA damage. During tumor progression, loss of p16INK4A expression may be necessary for cells with wild-type Rb to bypass this G1 arrest checkpoint and attain a fully transformed phenotype.
Shapiro
G I
GI
Department of Adult Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA.
Edwards
C D
CD
Ewen
M E
ME
Rollins
B J
BJ
D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics*; D004249:DNA Damage; D016193:G1 Phase / Q000235:genetics*; D016147:Genes, Tumor Suppressor; D006801:Humans; D014407:Tumor Cells, Cultured
D019941:Cyclin-Dependent Kinase Inhibitor p16