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End-to-end template jumping by the reverse transcriptase encoded by the R2 retrotransposon
Arkadiusz Bibillo1, Thomas H Eickbush
1Department of Biology, University of Rochester, Rochester, New York 14627, USA. eick@mail.rochester.edu
The Journal of Biological Chemistry
|January 31, 2004
Summary
The R2 retrotransposon reverse transcriptase can jump between RNA templates without sequence identity. This occurs by adding non-templated nucleotides to cDNA, which then anneal to acceptor RNA sequences.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Non-long terminal repeat (non-LTR) retrotransposons utilize reverse transcriptase for replication.
- R2 retrotransposon reverse transcriptase mediates template jumping between RNA molecules.
- Template jumping is also observed in RNA viruses with RNA-directed RNA polymerases.
Purpose of the Study:
- To elucidate the mechanism of end-to-end template jumps by the R2 reverse transcriptase.
- To understand how R2 reverse transcriptase achieves template switching in the absence of sequence homology.
Main Methods:
- Enzymatic assays to study R2 reverse transcriptase activity.
- Analysis of cDNA synthesis and annealing to RNA templates.
- Characterization of non-templated nucleotide addition at RNA template ends.
Main Results:
- R2 reverse transcriptase adds non-templated nucleotides to the 3'-end of cDNA when reaching the donor RNA terminus.
- These overhanging nucleotides facilitate annealing to the acceptor RNA's 3'-end.
- Annealing is most efficient with terminal nucleotides but can occur internally up to 5 nucleotides from the 3'-end.
Conclusions:
- The mechanism involves non-templated nucleotide addition and subsequent annealing, enabling template switching.
- This process explains chimeric integration products formed from multiple RNA templates.
- The findings provide insights into retrotransposon integration and viral RNA replication strategies.