Decreased p21 levels are required for efficient restart of DNA synthesis after S phase block.
The cyclin-dependent kinase inhibitor p21, a major transcriptional target of the tumor suppressor p53, plays a critical role in cell cycle arrest in G1 and G2 after DNA damage. It was previously shown that in some human cell lines when S phase is arrested, p53 is transcriptionally impaired such that some p53 targets including p21 are only weakly induced. We show here that during S phase arrest proteasome-mediated turnover of p21 is significantly increased in a manner that is independent of p53. It is well established that p21 can interact both with cyclin-dependent kinase complexes and with proliferating cell nuclear antigen (PCNA). Interestingly, the scant amount of p21 detected during S phase block cannot fully saturate cyclin A-cyclin-dependent kinase 2 complexes and does not interact detectably with PCNA. Importantly, DNA elongation assays in isolated nuclei show that the C terminus of p21 containing the PCNA-binding domain effectively blocks this process. This implies that p21 down-regulation could be an essential requirement for efficient restart of DNA synthesis. In line with this, only cells expressing low levels of p21 immediately progress through the cell cycle upon release from S phase arrest, whereas the remaining few high p21 producing cells move much more slowly through S. Thus, p21 down-regulation is multiply determined and is required for the reversibility of the arrest in S phase.
Gottifredi
Vanesa
V
Department of Biological Sciences, Columbia University, New York, New York 10027, USA.
McKinney
Kristine
K
Poyurovsky
Masha V
MV
Prives
Carol
C
D015152:Blotting, Northern; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D019925:Cyclin A / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis* / Q000378:metabolism; D015536:Down-Regulation; D016193:G1 Phase; D016195:G2 Phase; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D016196:S Phase; D013997:Time Factors; D014158:Transcription, Genetic; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
C493956:CDKN1A protein, human; D019925:Cyclin A; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D009097:Multienzyme Complexes; D018809:Proliferating Cell Nuclear Antigen; D011994:Recombinant Proteins; D016159:Tumor Suppressor Protein p53; D004247:DNA; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D006918:Hydroxyurea