Evolution of the R2 retrotransposon ribozyme and its self-cleavage site.
R2 is a non-long terminal repeat retrotransposon that inserts site-specifically in the tandem 28S rRNA genes of many animals. Previously, R2 RNA from various species of Drosophila was shown to self-cleave from the 28S rRNA/R2 co-transcript by a hepatitis D virus (HDV)-like ribozyme encoded at its 5' end. RNA cleavage was at the precise 5' junction of the element with the 28S gene. Here we report that RNAs encompassing the 5' ends of R2 elements from throughout its species range fold into HDV-like ribozymes. In vitro assays of RNA self-cleavage conducted in many R2 lineages confirmed activity. For many R2s, RNA self-cleavage was not at the 5' end of the element but at 28S rRNA sequences up to 36 nucleotides upstream of the junction. The location of cleavage correlated well with the types of endogenous R2 5' junctions from different species. R2 5' junctions were uniform for most R2s in which RNA cleavage was upstream in the rRNA sequences. The 28S sequences remaining on the first DNA strand synthesized during retrotransposition are postulated to anneal to the target site and uniformly prime second strand DNA synthesis. In species where RNA cleavage occurred at the R2 5' end, the 5' junctions were variable. This junction variation is postulated to result from the priming of second strand DNA synthesis by chance microhomologies between the target site and the first DNA strand. Finally, features of R2 ribozyme evolution, especially changes in cleavage site and convergence on the same active site sequences, are discussed.
Department of Biology, University of Rochester, Rochester, New York, United States of America.
Danna G
DG
Eickbush
William D
WD
Burke
Thomas H
TH
Eickbush
D000818:Animals; D020134:Catalytic Domain; D019143:Evolution, Molecular; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D016337:RNA, Catalytic / Q000235:genetics*; D012339:RNA, Ribosomal, 28S / Q000235:genetics; D018626:Retroelements / Q000235:genetics*
D016337:RNA, Catalytic; D012339:RNA, Ribosomal, 28S; D018626:Retroelements