Cip1 inhibits DNA replication but not PCNA-dependent nucleotide excision-repair.
BACKGROUND
DNA that is damaged by ultraviolet (UV) light is repaired predominantly by nucleotide excision-repair, a process requiring the DNA polymerase auxiliary factor PCNA. UV-irradiation also induces the production of Cip1 protein via activation of p53. Cip1 is an inhibitor of the cyclin-dependent kinases, which are required for the cell cycle to proceed through the G1/S-phase transition and initiate DNA replication. Inhibition by Cip1 probably causes the block to initiation of DNA replication that is seen in irradiated cells. Cip1 also directly inhibits the function of PCNA during DNA synthesis. As nucleotide excision-repair requires PCNA, the physiological relevance of PCNA inhibition by Cip1 is currently unclear.


RESULTS
We show that nucleotide excision-repair of UV-damaged DNA occurs in extracts of Xenopus eggs, and that this reaction is PCNA-dependent. The repair reaction is not inhibited by Cip1, even when the level of PCNA is reduced 100-fold so that it becomes limiting for DNA repair. By contrast, Cip1 strongly suppresses the function of PCNA in replicative DNA synthesis under these conditions.


CONCLUSIONS
Cip1 can potentially inhibit DNA replication in Xenopus egg extracts by inhibiting the cyclin-dependent kinase function required for the initiation of replication forks, and also by inhibiting PCNA function. The inhibition of PCNA is selective for its function in DNA replication, however, as Cip1 does not affect PCNA function in nucleotide excision-repair. The induction of Cip1 in response to DNA damage, therefore, allows repair to continue in the genome under conditions in which replication is severely inhibited.
Shivji
M K
MK
Imperial Cancer Research Fund, Clare Hall Laboratories, South Mimms, Hertfordshire, UK.
Grey
S J
SJ
Strausfeld
U P
UP
Wood
R D
RD
Blow
J J
JJ
D000818:Animals; D001483:Base Sequence; D002457:Cell Extracts; D002460:Cell Line; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000502:physiology*; D004247:DNA / Q000096:biosynthesis / Q000378:metabolism; D004260:DNA Repair / Q000502:physiology*; D004261:DNA Replication / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D009711:Nucleotides / Q000378:metabolism; D010063:Ovum; D018809:Proliferating Cell Nuclear Antigen / Q000502:physiology*; D047428:Protein Kinase Inhibitors; D014981:Xenopus
C493956:CDKN1A protein, human; D002457:Cell Extracts; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D009687:Nuclear Proteins; D009711:Nucleotides; D018809:Proliferating Cell Nuclear Antigen; D047428:Protein Kinase Inhibitors; D004247:DNA