A mechanism for sequential replication-coupled destruction of CRL4-Cdt2 substrates (616.1)

The CRL4Cdt2 ubiquitin E3 ligase targets a cohort of key cell cycle regulators during both S phase and DNA repair synthesis. These substrates include the p21 CDK inhibitor, the Cdt1 origin licensing protein, and the Set8 lysine methyltransferase. The intracellular concentrations of each of these proteins has profound consequences for cell proliferation and genome stability. CRL4-Cdt2 only targets substrates if they are bound to DNA-loaded PCNA, coupling substrate destruction to DNA synthesis. PCNA loading does not trigger simultaneous destruction of all CRL4-Cdt2 substrates however; Cdt1 degradation begins nearly instantaneously with PCNA loading whereas p21 degradation is significantly delayed for example. As a consequence, the substrates of CRL4-Cdt2 are destroyed in a stereotypical order, and we have observed this order in multiple cell lines during both DNA repair synthesis and in early S phase. To identify the molecular determinants of delayed vs. immediate destruction, we generated a series of fusio...