Degradation of p12 Subunit by CRL4Cdt2 E3 Ligase Inhibits Fork Progression after DNA Damage

After acute DNA damage, the cell arrests S-phase progression by inhibiting origin initiation and fork progression to repair damaged DNA. The intra-S-phase checkpoint kinase Chk1 phosphorylates Cdc25A to target the latter for degradation by CRL1 β-TrCP and so inhibit origin firing. The mechanism for inhibiting fork progression, however, has not been identified. Here, we show that degradation of p12, the fourth subunit of DNA polymerase δ, is critical for inhibiting fork progression. CRL4 Cdt2 is an E3 ligase that ubiquitinates and degrades p12 after UV treatment. Cells expressing a stable form of p12 exhibit UV-resistant DNA synthesis. DNA fiber assay and alkaline-sucrose gradient assay demonstrate that the impairment of fork progression after DNA damage requires p12 degradation. These results suggest that ubiquitination of p12 through CRL4 Cdt2 and subsequent degradation form one mechanism by which a cell responds to DNA damage to inhibit fork progression. Background: The mechanism for inhibiting fork progression after DNA damage still remains. Results: CRL4 Cdt2 promotes the degradation of the p12. Cells expressing a stable form of p12 exhibit UV-resistant DNA synthesis and decreased inhibition of fork progression. Conclusion: p12 degradation by CRL4 Cdt2 is critical for inhibiting fork progression. Significance: p12 degradation is one mechanism by which DNA damage in S-phase cells inhibits fork progression.