R2 retrotransposition on assembled nucleosomes depends on the translational position of the target site.
R2 retrotransposons insert into the 28S rRNA genes of insects. Integration occurs by specific cleavage of the target site and utilization of the released DNA end to prime reverse transcription of the RNA transcript. Specificity of the protein to the target site is dependent upon nucleotide sequence recognition extending from 35 bp upstream to 15 bp downstream of the cleavage site. In this report, we show that sequence recognition and cleavage by the R2 protein can occur while the target site is assembled into nucleosomes. Reconstitution of DNA fragments containing the 28S gene sequence into a set of nucleosomes with different translational frames revealed that the R2 site adopted the same rotational orientation with respect to the histone octamer. Binding and cleavage by the R2 protein were most efficient when the upstream binding site for the R2 protein was near a nucleosome end. Interaction of the R2 protein with the nucleosome disrupted the histone:DNA contacts in the 50 bp region directly bound by R2, but did not modify the remainder of the nucleosome structure.
Ye
Junqiang
J
University of Rochester, Department of Biology, Rochester, NY 14627, USA.
Yang
Zungyoon
Z
Hayes
Jeffrey J
JJ
Eickbush
Thomas H
TH
D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D002843:Chromatin / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D004331:Drosophila melanogaster; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009707:Nucleosomes / Q000378:metabolism*; D011485:Protein Binding; D014176:Protein Biosynthesis; D012313:RNA / Q000378:metabolism*; D012339:RNA, Ribosomal, 28S / Q000378:metabolism; D018626:Retroelements; D013997:Time Factors
D002843:Chromatin; D009707:Nucleosomes; D012339:RNA, Ribosomal, 28S; D018626:Retroelements; D012313:RNA; D004247:DNA; D003850:Deoxyribonuclease I