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Nucleic Acids Research 2005 33(6):1993-2002; doi:10.1093/nar/gki347
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Published online 6 April 2005

© The Author 2005. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oupjournals.org

Article

Functional roles of 3'-terminal structures of template RNA during in vivo retrotransposition of non-LTR retrotransposon, R1Bm

Tomohiro Anzai, Mizuko Osanai, Mitsuhiro Hamada and Haruhiko Fujiwara*

Department of Integrated Biosciences, Graduate School of Frontier Sciences, University of Tokyo Bioscience Building 501, Kashiwa, 277-8562, Japan

*To whom correspondence should be addressed. Tel: +81 4 7136 3659; Fax: +81 4 7136 3660; Email: haruh{at}k.u-tokyo.ac.jp

Received March 3, 2005. Revised March 9, 2005. Accepted March 21, 2005.

R1Bm is a non-LTR retrotransposon found specifically within 28S rRNA genes of the silkworm. Different from other non-LTR retrotransposons encoding two open reading frames (ORFs), R1Bm structurally lacks a poly (A) tract at its 3' end. To study how R1Bm initiates reverse transcription from the poly (A)-less template RNA, we established an in vivo retrotransposition system using recombinant baculovirus, and characterized retrotransposition activities of R1Bm. Target-primed reverse transcription (TPRT) of R1Bm occurred from the cleavage site generated by endonuclease (EN). The 147 bp of 3'-untranslated region (3'UTR) was essential for efficient retrotransposition of R1Bm. Even using the complete R1Bm element, however, reverse transcription started from various sites of the template RNA mostly with 5'-UG-3' or 5'-UGU-3' at their 3' ends, which are presumably base-paired with 3' end of the EN-digested 28S rDNA target sequence, 5'-AGTAGATAGGGACA-3'. When the downstream sequence of 28S rDNA target was added to the 3' end of R1 unit, reverse transcription started exactly from the 3' end of 3'UTR and retrotransposition efficiency increased. These results indicate that 3'-terminal structure of template RNA including read-through region interacts with its target rDNA sequences of R1Bm, which plays important roles in initial process of TPRT in vivo.


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