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Open Access Research article

The double-stranded break-forming activity of plant SPO11s and a novel rice SPO11 revealed by a Drosophila bioassay

Yoshinori Shingu12, Takeshi Tokai3, Yasuo Agawa4, Kentaro Toyota3, Selina Ahmed3, Makiko Kawagishi-Kobayashi5, Akira Komatsu5, Tsutomu Mikawa12, Masa-Toshi Yamamoto4, Kyo Wakasa3*, Takehiko Shibata12* and Kohji Kusano4*

Author Affiliations

1 Cellular & Molecular Biology Laboratory, RIKEN Advanced Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan

2 Department of Supramolecular Biology, Graduate School of Nanobioscience, Yokohama City University, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan

3 Department of Agriculture, Tokyo University of Agriculture, Atsugi, Kanagawa 243-0034, Japan

4 Center for Genetic Resource Education & Development, Kyoto Institute of Technology, Saga-Ippongi-cho, Ukyo-ku, Kyoto 616-8354, Japan

5 National Institute of Crop Science, 2-1-8 Kannondai, Tsukuba, Ibaraki 305-8518, Japan

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BMC Molecular Biology 2012, 13:1  doi:10.1186/1471-2199-13-1

Published: 16 January 2012

Abstract

Background

SPO11 is a key protein for promoting meiotic recombination, by generating chromatin locus- and timing-specific DNA double-strand breaks (DSBs). The DSB activity of SPO11 was shown by genetic analyses, but whether SPO11 exerts DSB-forming activity by itself is still an unanswered question. DSB formation by SPO11 has not been detected by biochemical means, probably because of a lack of proper protein-folding, posttranslational modifications, and/or specific SPO11-interacting proteins required for this activity. In addition, plants have multiple SPO11-homologues.

Results

To determine whether SPO11 can cleave DNA by itself, and to identify which plant SPO11 homologue cleaves DNA, we developed a Drosophila bioassay system that detects the DSB signals generated by a plant SPO11 homologue expressed ectopically. We cytologically and genetically demonstrated the DSB activities of Arabidopsis AtSPO11-1 and AtSPO11-2, which are required for meiosis, in the absence of other plant proteins. Using this bioassay, we further found that a novel SPO11-homologue, OsSPO11D, which has no counterpart in Arabidopsis, displays prominent DSB-forming activity. Quantitative analyses of the rice SPO11 transcripts revealed the specific increase in OsSPO11D mRNA in the anthers containing meiotic pollen mother cells.

Conclusions

The Drosophila bioassay system successfully demonstrated that some plant SPO11 orthologues have intrinsic DSB activities. Furthermore, we identified a novel SPO11 homologue, OsSPO11D, with robust DSB activity and a possible meiotic function.