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  • The EMBO Journal (2007) 26, 1327 - 1339
  • doi:10.1038/sj.emboj.7601585

Published online: 15 February 2007

Genome-wide localization of pre-RC sites and identification of replication origins in fission yeast

Makoto Hayashi1, Yuki Katou2, Takehiko Itoh3, Mitsutoshi Tazumi1, Yoshiki Yamada1,a, Tatsuro Takahashi1,b, Takuro Nakagawa1, Katsuhiko Shirahige2,4 and Hisao Masukata1

  1. Department of Biology, Graduate School of Science, Osaka University, Toyonaka, Osaka, Japan
  2. Riken Genomic Science Center, Human Genome Research Group, Genome Informatics Team, Tsurumi-ku, Yokohama, Kanagawa, Japan
  3. Research Center for Advanced Science and Technology, Mitsubishi Research Institute Inc., Chiyoda-ku, Tokyo, Japan
  4. Center for Biological Resources and Informatics, Division of Gene Research, and Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Midori-ku, Yokohama, Japan

Correspondence to:

Hisao Masukata, Department of Biology, Graduate School of Science, Osaka University, 1-1, Machikaneyama-cho, Toyonaka, Osaka 560-0043, Japan. Tel.: +81 6 6850 5432; Fax: +81 6 6850 5440; E-mail: masukata@bio.sci.osaka-u.ac.jp

aPresent address: The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA

bPresent address: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA

Received 24 July 2006; Accepted 8 January 2007

DNA replication of eukaryotic chromosomes initiates at a number of discrete loci, called replication origins. Distribution and regulation of origins are important for complete duplication of the genome. Here, we determined locations of Orc1 and Mcm6, components of pre-replicative complex (pre-RC), on the whole genome of Schizosaccharomyces pombe using a high-resolution tiling array. Pre-RC sites were identified in 460 intergenic regions, where Orc1 and Mcm6 colocalized. By mapping of 5-bromo-2'-deoxyuridine (BrdU)-incorporated DNA in the presence of hydroxyurea (HU), 307 pre-RC sites were identified as early-firing origins. In contrast, 153 pre-RC sites without BrdU incorporation were considered to be late and/or inefficient origins. Inactivation of replication checkpoint by Cds1 deletion resulted in BrdU incorporation with HU specifically at the late origins. Early and late origins tend to distribute separately in large chromosome regions. Interestingly, pericentromeric heterochromatin and the silent mating-type locus replicated in the presence of HU, whereas the inner centromere or subtelomeric heterochromatin did not. Notably, MCM did not bind to inner centromeres where origin recognition complex was located. Thus, replication is differentially regulated in chromosome domains.

  • Keywords:

    • centromere,
    • DNA microarray,
    • fission yeast,
    • replication origin,
    • subtelomere