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Cell Cycle
2016 May 02;159:1213-26. doi: 10.1080/15384101.2015.1106652.
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Characterization of conserved arginine residues on Cdt1 that affect licensing activity and interaction with Geminin or Mcm complex.
You Z
,
Ode KL
,
Shindo M
,
Takisawa H
,
Masai H
.
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All organisms ensure once and only once replication during S phase through a process called replication licensing. Cdt1 is a key component and crucial loading factor of Mcm complex, which is a central component for the eukaryotic replicative helicase. In higher eukaryotes, timely inhibition of Cdt1 by Geminin is essential to prevent rereplication. Here, we address the mechanism of DNA licensing using purified Cdt1, Mcm and Geminin proteins in combination with replication in Xenopus egg extracts. We mutagenized the 223th arginine of mouse Cdt1 (mCdt1) to cysteine or serine (R-S or R-C, respectively) and 342nd and 346th arginines constituting an arginine finger-like structure to alanine (RR-AA). The RR-AA mutant of Cdt1 could not only rescue the DNA replication activity in Cdt1-depleted extracts but also its specific activity for DNA replication and licensing was significantly increased compared to the wild-type protein. In contrast, the R223 mutants were partially defective in rescue of DNA replication and licensing. Biochemical analyses of these mutant Cdt1 proteins indicated that the RR-AA mutation disabled its functional interaction with Geminin, while R223 mutations resulted in ablation in interaction with the Mcm2∼7 complex. Intriguingly, the R223 mutants are more susceptible to the phosphorylation-induced inactivation or chromatin dissociation. Our results show that conserved arginine residues play critical roles in interaction with Geminin and Mcm that are crucial for proper conformation of the complexes and its licensing activity.
Arias,
PCNA functions as a molecular platform to trigger Cdt1 destruction and prevent re-replication.
2006, Pubmed,
Xenbase
Arias,
PCNA functions as a molecular platform to trigger Cdt1 destruction and prevent re-replication.
2006,
Pubmed
,
Xenbase
Ballabeni,
Human geminin promotes pre-RC formation and DNA replication by stabilizing CDT1 in mitosis.
2004,
Pubmed
,
Xenbase
Ballabeni,
Human CDT1 associates with CDC7 and recruits CDC45 to chromatin during S phase.
2009,
Pubmed
Coulombe,
A spontaneous Cdt1 mutation in 129 mouse strains reveals a regulatory domain restraining replication licensing.
2013,
Pubmed
De Marco,
Quaternary structure of the human Cdt1-Geminin complex regulates DNA replication licensing.
2009,
Pubmed
,
Xenbase
Ferenbach,
Functional domains of the Xenopus replication licensing factor Cdt1.
2005,
Pubmed
,
Xenbase
Fujita,
Cell cycle- and chromatin binding state-dependent phosphorylation of human MCM heterohexameric complexes. A role for cdc2 kinase.
1998,
Pubmed
Fujita,
Cdt1 revisited: complex and tight regulation during the cell cycle and consequences of deregulation in mammalian cells.
2006,
Pubmed
Gillespie,
Reconstitution of licensed replication origins on Xenopus sperm nuclei using purified proteins.
2001,
Pubmed
,
Xenbase
Hashimoto,
The phosphorylated C-terminal domain of Xenopus Cut5 directly mediates ATR-dependent activation of Chk1.
2006,
Pubmed
,
Xenbase
Hashimoto,
Xenopus Cut5 is essential for a CDK-dependent process in the initiation of DNA replication.
2003,
Pubmed
,
Xenbase
Jee,
Structure and mutagenesis studies of the C-terminal region of licensing factor Cdt1 enable the identification of key residues for binding to replicative helicase Mcm proteins.
2010,
Pubmed
Kawakami,
Formation of an ATP-DnaA-specific initiation complex requires DnaA Arginine 285, a conserved motif in the AAA+ protein family.
2005,
Pubmed
Kisielewska,
Dynamic interactions of high Cdt1 and geminin levels regulate S phase in early Xenopus embryos.
2012,
Pubmed
,
Xenbase
Kubota,
Licensing of DNA replication by a multi-protein complex of MCM/P1 proteins in Xenopus eggs.
1997,
Pubmed
,
Xenbase
Lee,
Structural basis for inhibition of the replication licensing factor Cdt1 by geminin.
2004,
Pubmed
,
Xenbase
Li,
The SCF(Skp2) ubiquitin ligase complex interacts with the human replication licensing factor Cdt1 and regulates Cdt1 degradation.
2003,
Pubmed
Liu,
TopBP1 recruits Brg1/Brm to repress E2F1-induced apoptosis, a novel pRb-independent and E2F1-specific control for cell survival.
2004,
Pubmed
Liu,
Structural insights into the Cdt1-mediated MCM2-7 chromatin loading.
2012,
Pubmed
Lutzmann,
A Cdt1-geminin complex licenses chromatin for DNA replication and prevents rereplication during S phase in Xenopus.
2006,
Pubmed
,
Xenbase
Masai,
Eukaryotic chromosome DNA replication: where, when, and how?
2010,
Pubmed
McGarry,
Geminin, an inhibitor of DNA replication, is degraded during mitosis.
1998,
Pubmed
,
Xenbase
Mimura,
Xenopus Cdc45-dependent loading of DNA polymerase alpha onto chromatin under the control of S-phase Cdk.
1998,
Pubmed
,
Xenbase
Nishitani,
Two E3 ubiquitin ligases, SCF-Skp2 and DDB1-Cul4, target human Cdt1 for proteolysis.
2006,
Pubmed
Nishitani,
DNA replication licensing.
2004,
Pubmed
,
Xenbase
Nishitani,
The human licensing factor for DNA replication Cdt1 accumulates in G1 and is destabilized after initiation of S-phase.
2001,
Pubmed
,
Xenbase
Ode,
Inter-origin cooperativity of geminin action establishes an all-or-none switch for replication origin licensing.
2011,
Pubmed
,
Xenbase
Okorokov,
Molecular structure of human geminin.
2004,
Pubmed
Sugimoto,
Identification of novel human Cdt1-binding proteins by a proteomics approach: proteolytic regulation by APC/CCdh1.
2008,
Pubmed
Sugimoto,
Cdt1 phosphorylation by cyclin A-dependent kinases negatively regulates its function without affecting geminin binding.
2004,
Pubmed
Tada,
Repression of origin assembly in metaphase depends on inhibition of RLF-B/Cdt1 by geminin.
2001,
Pubmed
,
Xenbase
Takara,
Multiple Cdt1 molecules act at each origin to load replication-competent Mcm2-7 helicases.
2011,
Pubmed
Thépaut,
Crystal structure of the coiled-coil dimerization motif of geminin: structural and functional insights on DNA replication regulation.
2004,
Pubmed
Tsuyama,
Repression of nascent strand elongation by deregulated Cdt1 during DNA replication in Xenopus egg extracts.
2009,
Pubmed
,
Xenbase
Waga,
Dynamics of DNA binding of replication initiation proteins during de novo formation of pre-replicative complexes in Xenopus egg extracts.
2006,
Pubmed
,
Xenbase
Wei,
Characterization and structure determination of the Cdt1 binding domain of human minichromosome maintenance (Mcm) 6.
2010,
Pubmed
Whittaker,
Drosophila double parked: a conserved, essential replication protein that colocalizes with the origin recognition complex and links DNA replication with mitosis and the down-regulation of S phase transcripts.
2000,
Pubmed
Wohlschlegel,
Inhibition of eukaryotic DNA replication by geminin binding to Cdt1.
2000,
Pubmed
,
Xenbase
Xouri,
Cdt1 interactions in the licensing process: a model for dynamic spatiotemporal control of licensing.
2007,
Pubmed
Xouri,
Cdt1 associates dynamically with chromatin throughout G1 and recruits Geminin onto chromatin.
2007,
Pubmed
Yanagi,
Mouse geminin inhibits not only Cdt1-MCM6 interactions but also a novel intrinsic Cdt1 DNA binding activity.
2002,
Pubmed
You,
Cdt1 forms a complex with the minichromosome maintenance protein (MCM) and activates its helicase activity.
2008,
Pubmed
You,
The mini-chromosome maintenance (Mcm) complexes interact with DNA polymerase α-primase and stimulate its ability to synthesize RNA primers.
2013,
Pubmed
