Regulation of Cdt1 in the cell cycle replication initiation

1. Regulation of Cdt1 in the cell cycle replication initiation Xiaoming Zhang Supervisor: professor Hui Zhang 2010-3-15

2. outlines • Molecular mechanisms for cell cycle regulation of DNA replication initiation. • Cdt1, a central factor for the cell cycle regulation of replication initiation, is strictly regulated by three ubiquitin ligases.

3. The mammalian cell cycle

4. The pre-RC formation is a dynamic process A pre-replication complex (pre-RC) is a protein complex that forms at the origin of replication during the initiation step of DNA replication. once and only once replication per single cell cycle" is achieved by the periodic assembly and disassembly of pre-replication complexes (pre-RCs) at replication origins .

5. Cdt1 is an essential target of the Cdc10/Sct1 transcription factor: requirement for DNA replication and inhibition of mitosis. If fission yeastcells were deleted with cdt1, All germinating spores contain a 1 N DNA content anddisplay the terminal arrest phonotype or arrested cell cycle progression. Johannes F and David Beach. EMBO J. (1994)13, 425-434

6. hCdt1 accumulates in Early G1 and Disappears after S-phase Onset Hideo Nishitani et al. J.Biol.Chem(2001) 48, 44905–44911

7. Cdt1 is strictly regulated by three ubiquitin ligases

9. During S and G2 phases, Cdt1 is brought to proteolysis by Cdk phosphorylation-dependent SCFSkp2-mediated ubiquitination In vivo interaction of Cdt1 with various cell cycle-regulating proteins and the role of the Cy motif (Arg68, Arg69,Leu70). Nozomi Sugimoto et al. J.Biol.Chem(2004)19, 19691–19697

10. Association of Cdt1 with Skp2 is dependent on phosphorylation by cyclin A-dependent kinases and results in its degradation during S and G2 phases Nozomi Sugimoto et al. J.Biol.Chem(2004)19, 19691–19697

11. SCFSkp2 complex also mediate rapid degradation of Cdt1 upon UV-induced DNA damage Takeshi Kondo et al. J. Biol. Chem (2004) 26, 27315–27319

12. L2DTL interacts with CDT1 and regulates CDT1 protein stability in response to gamma-irradiation DNA Damage in Drosophila Higa LA et al. Cell Cycle (2006)15, 1675-1680

13. Cdt1 degradation in S–G2 phases and after UV-irradiation occurs in the absence of Skp2 Nishitani, H et al, EMBO J (2006). 25, 1126–1136

14. Stabilization of Cdt1 in S–G2 phases after cosilencing of Skp2 and Cul4 Nishitani, H et al, EMBO J (2006). 25, 1126–1136

15. A model showing that Cdt1 is inactivated by multiple pathways during the cell cycle. Nishitani, H et al, EMBO J (2006). 25, 1126–1136

16. APC/CCdh1 Ubiquitinates Cdt1 In Vitro Nozomi Sugimoto et al. Mol. Cell (2008)19, 1007–1021

17. A model for roles of APC/CCdh1 -mediated Cdt1 proteolytic regulation Nozomi Sugimoto et al. Mol. Cell (2008)19, 1007–1021

18. Cdt1, a central factor for the cell cycle regulation of replication initiation is strictly regulated by three ubiquitin ligases. • During S and G2 phases, Cdt1 is brought to proteolysis by Cdk phosphorylation-dependent SCFSkp2-mediated ubiquitination. SCFSkp2 also degradated Cdt1 upon UV-induced DNA damage. • Cdt1 is also regulated by replication-coupled, Cullin4-DDB1Cdt2 ubiquitin ligase-mediated ubiquitination in S phase. Cullin4-DDB1Cdt2 also degradated Cdt1 in response to gamma-irradiation DNA Damage. • Cdt1 can also be rapidly cleared by APC/CCdh1-mediated proteolysis in G1 and G0 phases.

19. References • Nozomi Sugimoto, Yasutoshi Tatsumi, Tatsuya Tsurumi, Akio Matsukage, Tohru Kiyono,Hideo Nishitani, and Masatoshi Fujita. J. Biol. Chem(2004) 19, 19691–19697. • Nozomi Sugimoto, Yasutoshi Tatsumi, Tatsuya Tsurumi, Akio Matsukage, Tohru Kiyono,Hideo Nishitani, and Masatoshi Fujita. Mol. Cell (2008) 19, 1007–1021. • Hideo Nishitani1, Nozomi Sugimoto,Vassilis Roukos, Yohsuke. EMBO J. (2006) 25, 1126–1136. • Hideo Nishitani, Stavros Taraviras, Zoi Lygerou, and Takeharu Nishimoto. EMBO J. (1994) 13,425-434. • Vasiliki Tsakraklides and Stephen P. Bell. J. Biol. Chem (2010) in press.

20. Thanks!