Nucleic Acids Research Advance Access originally published online on December 20, 2007
Nucleic Acids Research 2008 36(3):984-997; doi:10.1093/nar/gkm1082
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Nucleic Acids Research, 2008, Vol. 36, No. 3 984-997
© 2007 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 |
Targeted induction of meiotic double-strand breaks reveals chromosomal domain-dependent regulation of Spo11 and interactions among potential sites of meiotic recombination
1Shibata Distinguished Senior Scientist Laboratory, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198 and 2Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, 3-8-1 Komaba, Meguro-ku, Tokyo 153-8902, Japan
*To whom correspondence should be addressed. Tel: +81 (0) 48 467 9277; Fax: +81 (0) 48 462 4691; Email: tfukuda{at}riken.jp
Correspondence may also be addressed to Kunihiro Ohta. Tel: +81 (0) 3 5465 8834; Fax: +81 (0) 3 5465 8834; Email: kohta{at}bio.c.u-tokyo.ac.jp
Received October 17, 2007. Revised November 17, 2007. Accepted November 17, 2007.
Meiotic recombination is initiated by programmed DNA double-strand break (DSB) formation mediated by Spo11. DSBs occur with frequency in chromosomal regions called hot domains but are seldom seen in cold domains. To obtain insights into the determinants of the distribution of meiotic DSBs, we examined the effects of inducing targeted DSBs during yeast meiosis using a UAS-directed form of Spo11 (Gal4BD-Spo11) and a meiosis-specific endonuclease, VDE (PI-SceI). Gal4BD-Spo11 cleaved its target sequence (UAS) integrated in hot domains but rarely in cold domains. However, Gal4BD-Spo11 did bind to UAS and VDE efficiently cleaved its recognition sequence in either context, suggesting that a cold domain is not a region of inaccessible or uncleavable chromosome structure. Importantly, self-association of Spo11 occurred at UAS in a hot domain but not in a cold domain, raising the possibility that Spo11 remains in an inactive intermediate state in cold domains. Integration of UAS adjacent to known DSB hotspots allowed us to detect competitive interactions among hotspots for activation. Moreover, the presence of VDE-introduced DSB repressed proximal hotspot activity, implicating DSBs themselves in interactions among hotspots. Thus, potential sites for Spo11-mediated DSB are subject to domain-specific and local competitive regulations during and after DSB formation.