Nucleic Acids Research Advance Access first published online on September 19, 2006
This version published online on October 5, 2006
Nucleic Acids Research, doi:10.1093/nar/gkl661
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© 2006 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Molecular Biology |
The spatial arrangement of ORC binding modules determines the functionality of replication origins in budding yeast
Department of Biochemistry, Molecular Biology and Biophysics 6-155 Jackson Hall, 321 Church Street SE, Minneapolis, MN 55455, USA
*To whom correspondence should be addressed. Tel: +1 612 624 2469; Fax: +1 612 625 2163; Email: bieli003{at}umn.edu
Received June 2, 2006. Revised August 22, 2006. Accepted August 28, 2006.
In the quest to define autonomously replicating sequences (ARSs) in eukaryotic cells, an ARS consensus sequence (ACS) has emerged for budding yeast. This ACS is recognized by the replication initiator, the origin recognition complex (ORC). However, not every match to the ACS constitutes a replication origin. Here, we investigated the requirements for ORC binding to origins that carry multiple, redundant ACSs, such as ARS603. Previous studies raised the possibility that these ACSs function as individual ORC binding sites. Detailed mutational analysis of the two ACSs in ARS603 revealed that they function in concert and give rise to an initiation pattern compatible with a single bipartite ORC binding site. Consistent with this notion, deletion of one base pair between the ACS matches abolished ORC binding at ARS603. Importantly, loss of ORC binding in vitro correlated with the loss of ARS activity in vivo. Our results argue that replication origins in yeast are in general comprised of bipartite ORC binding sites that cannot function in random alignment but must conform to a configuration that permits ORC binding. These requirements help to explain why only a limited number of ACS matches in the yeast genome qualify as ORC binding sites.
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