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Molecular and Cellular Biology, February 2006, p. 1124-1134, Vol. 26, No. 3
0270-7306/06/$08.00+0     doi:10.1128/MCB.26.3.1124-1134.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Essential Elements of a Licensed, Mammalian Plasmid Origin of DNA Synthesis{dagger}

Jindong Wang,{ddagger} Scott E. Lindner,{ddagger} Elizabeth R. Leight,§ and Bill Sugden*

McArdle Laboratory for Cancer Research, University of Wisconsin—Madison, Madison, Wisconsin 53706

Received 22 August 2005/ Returned for modification 17 October 2005/ Accepted 9 November 2005

We developed a mammalian plasmid replicon with a formerly uncharacterized origin of DNA synthesis, 8xRep*. 8xRep* functions efficiently to support once-per-cell-cycle synthesis of plasmid DNA which initiates within Rep*. By characterizing Rep*'s requirements for acting as an origin, we have uncovered several striking properties it shares with DS, the only other well-characterized, licensed, mammalian plasmid origin of DNA synthesis. Rep* contains a pair of previously unrecognized Epstein-Barr nuclear antigen 1 (EBNA1)-binding sites that are both necessary and sufficient in cis for its origin activity. These sites have an essential 21-bp center-to-center spacing, are bent by EBNA1, and recruit the origin recognition complex. The properties shared between DS and Rep* define cis and trans characteristics of a mammalian, extrachromosomal replicon. The role of EBNA1 likely reflects its evolution from cellular factors involved in the assembly of the initiation machinery.

* Corresponding author. Mailing address: 1400 University Ave., Madison, WI 53706. Phone: (608) 262-6697. Fax: (608) 262-2824. E-mail: sugden{at}oncology.wisc.edu.

{dagger} Supplemental material for this article may be found at http://mcb.asm.org/.

{ddagger} J.W. and S.E.L. contributed equally to this study.

§ Present address: Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St. Louis, MO 63110.

Molecular and Cellular Biology, February 2006, p. 1124-1134, Vol. 26, No. 3
0270-7306/06/$08.00+0     doi:10.1128/MCB.26.3.1124-1134.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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