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Eukaryotic Transposable Elements and Genome Evolution Special Feature
EUKARYOTIC TRANSPOSABLE ELEMENTS AND GENOME EVOLUTION SPECIAL FEATURE / BIOLOGICAL SCIENCES / RESEARCH ARTICLES / BIOCHEMISTRY
RNA from the 5' end of the R2 retrotransposon controls R2 protein binding to and cleavage of its DNA target site
Department of Biology, University of Rochester, Rochester, NY 14627-0211
Edited by Susan R. Wessler, University of Georgia, Athens, GA, and approved August 11, 2006 (received for review June 30, 2006)
Non-LTR retrotransposons insert into eukaryotic genomes by target-primed reverse transcription (TPRT), a process in which cleaved DNA targets are used to prime reverse transcription of the element's RNA transcript. Many of the steps in the integration pathway of these elements can be characterized in vitro for the R2 element because of the rigid sequence specificity of R2 for both its DNA target and its RNA template. R2 retrotransposition involves identical subunits of the R2 protein bound to different DNA sequences upstream and downstream of the insertion site. The key determinant regulating which DNA-binding conformation the protein adopts was found to be a 320-nt RNA sequence from near the 5' end of the R2 element. In the absence of this 5' RNA the R2 protein binds DNA sequences upstream of the insertion site, cleaves the first DNA strand, and conducts TPRT when RNA containing the 3' untranslated region of the R2 transcript is present. In the presence of the 320-nt 5' RNA, the R2 protein binds DNA sequences downstream of the insertion site. Cleavage of the second DNA strand by the downstream subunit does not appear to occur until after the 5' RNA is removed from this subunit. We postulate that the removal of the 5' RNA normally occurs during reverse transcription, and thus provides a critical temporal link to first- and second-strand DNA cleavage in the R2 retrotransposition reaction.
endonuclease | retrotransposition | reverse transcription | RNA–protein interactions
*Present address: Department of Biology, University of Texas, Arlington, TX 76019.
Present address: Department of Cellular and Molecular Biology, Harvard University, Cambridge, MA 02138.
The authors declare no conflict of interest.
This article is a PNAS direct submission.
To whom correspondence should be addressed. E-mail: eick{at}mail.rochester.edu
© 2006 by The National Academy of Sciences of the USA
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