Misregulated expression of the cyclin dependent kinase 2 protein in human fibroblasts is accompanied by the inability to maintain a G2 arrest following DNA damage.
The misregulation of cell cycle checkpoints has been implicated in the onset of neoplasia. To thoroughly understand the differences in checkpoint regulation between normal and transformed cells, we have compared the cell cycle responses of normal and TAg-transformed fibroblasts to DNA damage by gamma-irradiation. Normal cell lines arrest in both G1 and G2 for in excess of 48 h after DNA damage. Surprisingly, both cyclin-dependent kinase 2 (CDK2) and cyclin A proteins were specifically down-regulated within 24 h of DNA damage. In contrast, TAg transformed cells did not down-regulate either cyclin A or CDK2 after DNA damage and showed a significantly shortened G2 arrest. To investigate the role CDK2 down-regulation plays in cell cycle arrests, we generated normal cell lines that constitutively overexpress CDK2. These cells fail to down-regulate both CDK2 protein and CDK2 protein kinase activity after DNA damage and also show a G2 checkpoint defect; although the cells are able to normally initiate both a G1 and a G2 arrest, they prematurely escape the G2-M arrest after DNA damage. The escape from G2 correlates with an illicit activation of cyclin B-associated protein kinase activity in these cells. These results suggest that the misregulation of CDK2 contributes to the failure of checkpoint control by allowing cells to enter mitosis prematurely.
Department of Molecular Cell Biology, Glaxo Research Institute, Research Triangle Park, North Carolina 27709, USA.
D H
DH
Walker
G R
GR
Adami
K M
KM
Dold
L E
LE
Babiss
D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle / Q000235:genetics* / Q000528:radiation effects; D002461:Cell Line, Transformed; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases / Q000096:biosynthesis*; D004249:DNA Damage; D015536:Down-Regulation / Q000235:genetics / Q000528:radiation effects; D005347:Fibroblasts / Q000378:metabolism / Q000528:radiation effects; D016195:G2 Phase / Q000235:genetics* / Q000528:radiation effects; D005720:Gamma Rays; D006801:Humans; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis*
D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases