QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online 1 September 2004
doi: 10.1242/dev.01348

This Article Figures Only Full Text Full Text (PDF) All Versions of this Article: dev.01348v1 131/19/4807    most recent Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Related articles in Development Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Thomer, M. Articles by Calvi, B. R. Search for Related Content PubMed PubMed Citation Articles by Thomer, M. Articles by Calvi, B. R.

Drosophila double-parked is sufficient to induce re-replication during development and is regulated by cyclin E/CDK2

Marguerite Thomer^*, Noah R. May^*, Bhagwan D. Aggarwal, Garrick Kwok and Brian R. Calvi

Department of Genetics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6145, USA

Author for correspondence (e-mail: calvi{at}mail.med.upenn.edu)

Accepted 5 July 2004

It is important that chromosomes are duplicated only once per cell cycle. Over-replication is prevented by multiple mechanisms that block the reformation of a pre-replicative complex (pre-RC) onto origins in S and G₂ phase. We have investigated the developmental regulation of Double-parked (Dup) protein, the Drosophila ortholog of Cdt1, a conserved and essential pre-RC component found in human and other organisms. We find that phosphorylation and degradation of Dup protein at G₁/S requires cyclin E/CDK2. The N terminus of Dup, which contains ten potential CDK phosphorylation sites, is necessary and sufficient for Dup degradation during S phase of mitotic cycles and endocycles. Mutation of these ten phosphorylation sites, however, only partially stabilizes the protein, suggesting that multiple mechanisms ensure Dup degradation. This regulation is important because increased Dup protein is sufficient to induce profound rereplication and death of developing cells. Mis-expression has different effects on genomic replication than on developmental amplification from chorion origins. The C terminus alone has no effect on genomic replication, but it is better than full-length protein at stimulating amplification. Mutation of the Dup CDK sites increases genomic re-replication, but is dominant negative for amplification. These two results suggest that phosphorylation regulates Dup activity differently during these developmentally specific types of DNA replication. Moreover, the ability of the CDK site mutant to rapidly inhibit BrdU incorporation suggests that Dup is required for fork elongation during amplification. In the context of findings from human and other cells, our results indicate that stringent regulation of Dup protein is critical to protect genome integrity.

Key words: DNA replication, Cyclin E, Double-parked, Cdt1, Chorion gene amplification

Related articles in Development:

Double-parked for replication

Development 2004 131: e1904. [Full Text]  

This article has been cited by other articles:

				J. R. Hall, H. O. Lee, B. D. Bunker, E. S. Dorn, G. C. Rogers, R. J. Duronio, and J. G. Cook Cdt1 and Cdc6 Are Destabilized by Rereplication-induced DNA Damage J. Biol. Chem., September 12, 2008; 283(37): 25356 - 25363. [Abstract] [Full Text] [PDF]

				S. Y. Park and M. Asano The origin recognition complex is dispensable for endoreplication in Drosophila PNAS, August 26, 2008; 105(34): 12343 - 12348. [Abstract] [Full Text] [PDF]

				A. M. Moses, M. E. Liku, J. J. Li, and R. Durbin Regulatory evolution in proteins by turnover and lineage-specific changes of cyclin-dependent kinase consensus sites PNAS, November 6, 2007; 104(45): 17713 - 17718. [Abstract] [Full Text] [PDF]

				E. A. Park, D. M. MacAlpine, and T. L. Orr-Weaver Inaugural Article: Drosophila follicle cell amplicons as models for metazoan DNA replication: A cyclinE mutant exhibits increased replication fork elongation PNAS, October 23, 2007; 104(43): 16739 - 16746. [Abstract] [Full Text] [PDF]

				E. E. Arias and J. C. Walter Strength in numbers: preventing rereplication via multiple mechanisms in eukaryotic cells Genes & Dev., March 1, 2007; 21(5): 497 - 518. [Abstract] [Full Text] [PDF]

				Y. Kim and E. T. Kipreos The Caenorhabditis elegans Replication Licensing Factor CDT-1 Is Targeted for Degradation by the CUL-4/DDB-1 Complex Mol. Cell. Biol., February 15, 2007; 27(4): 1394 - 1406. [Abstract] [Full Text] [PDF]

				S. Shibutani, L. M. Swanhart, and R. J. Duronio Rbf1-independent termination of E2f1-target gene expression during early Drosophila embryogenesis Development, February 1, 2007; 134(3): 467 - 478. [Abstract] [Full Text] [PDF]

				C. A. Lovejoy, K. Lock, A. Yenamandra, and D. Cortez DDB1 Maintains Genome Integrity through Regulation of Cdt1 Mol. Cell. Biol., November 1, 2006; 26(21): 7977 - 7990. [Abstract] [Full Text] [PDF]

				W. Zhu and A. Dutta An ATR- and BRCA1-Mediated Fanconi Anemia Pathway Is Required for Activating the G2/M Checkpoint and DNA Damage Repair upon Rereplication Mol. Cell. Biol., June 15, 2006; 26(12): 4601 - 4611. [Abstract] [Full Text] [PDF]

				B. M. Green, R. J. Morreale, B. Ozaydin, J. L. DeRisi, and J. J. Li Genome-wide Mapping of DNA Synthesis in Saccharomyces cerevisiae Reveals That Mechanisms Preventing Reinitiation of DNA Replication Are Not Redundant Mol. Biol. Cell, May 1, 2006; 17(5): 2401 - 2414. [Abstract] [Full Text] [PDF]

				D. Remus, M. Blanchette, D. C. Rio, and M. R. Botchan CDK Phosphorylation Inhibits the DNA-binding and ATP-hydrolysis Activities of the Drosophila Origin Recognition Complex J. Biol. Chem., December 2, 2005; 280(48): 39740 - 39751. [Abstract] [Full Text] [PDF]

				M. E. Liku, V. Q. Nguyen, A. W. Rosales, K. Irie, and J. J. Li CDK Phosphorylation of a Novel NLS-NES Module Distributed between Two Subunits of the Mcm2-7 Complex Prevents Chromosomal Rereplication Mol. Biol. Cell, October 1, 2005; 16(10): 5026 - 5039. [Abstract] [Full Text] [PDF]

				N. R. May, M. Thomer, K. F. Murnen, and B. R. Calvi Levels of the origin-binding protein Double parked and its inhibitor Geminin increase in response to replication stress J. Cell Sci., September 15, 2005; 118(18): 4207 - 4217. [Abstract] [Full Text] [PDF]

				D. Y. Takeda, J. D. Parvin, and A. Dutta Degradation of Cdt1 during S Phase Is Skp2-independent and Is Required for Efficient Progression of Mammalian Cells through S Phase J. Biol. Chem., June 17, 2005; 280(24): 23416 - 23423. [Abstract] [Full Text] [PDF]

				C. Raynaud, C. Perennes, C. Reuzeau, O. Catrice, S. Brown, and C. Bergounioux Cell and plastid division are coordinated through the prereplication factor AtCDT1 PNAS, June 7, 2005; 102(23): 8216 - 8221. [Abstract] [Full Text] [PDF]

				E. E. Arias and J. C. Walter Replication-dependent destruction of Cdt1 limits DNA replication to a single round per cell cycle in Xenopus egg extracts Genes & Dev., January 1, 2005; 19(1): 114 - 126. [Abstract] [Full Text] [PDF]