Ty3 reverse transcriptase complexed with an RNA-DNA hybrid shows structural and functional asymmetry

Elżbieta Nowak,Justyna Studnicka,Marion K Bona,Jakub Jurkowski,Stuart F J Le Grice,Jennifer T Miller,Roman H Szczepanowski,Marcin Nowotny

doi:10.1038/nsmb.2785

Abstract

Retrotransposons are a class of mobile genetic elements that replicate by converting their single-stranded RNA intermediate to double-stranded DNA through the combined DNA polymerase and ribonuclease H (RNase H) activities of the element-encoded reverse transcriptase (RT). However, while a wealth of structural information is available for lentiviral and gammaretroviral RTs, equivalent studies on counterpart enzymes of long terminal repeat (LTR)-containing retrotransposons, from which they are evolutionarily derived, is lacking. In this study, we report the first crystal structure of a complex of RT from the Saccharomyces cerevisiae LTR-retrotransposon Ty3 in the presence of its polypurine tract-containing RNA-DNA hybrid. In contrast to its retroviral counterparts, Ty3 RT adopts an asymmetric homodimeric architecture, whose assembly is substrate-dependent. More strikingly, our structure and biochemical data suggest that the RNase H and DNA polymerase activities are contributed by individual subunits of the homodimer.

Highlights

Retrotransposons are mobile genetic elements that replicate through an RNA intermediate
The structural basis for such spatial separation is established for human immunodeficiency virus (HIV)-1 RT16,17, the origin of the shorter distance for Ty3 reverse transcriptase (RT) is difficult to explain from the retroviral structures
Structural similarity between the ribonuclease H (RNase H) subdomain of HIV-1 RT and its connection subdomain originally suggested the former arose through domain duplication, while an alternative theory proposes that the functional RNase H domain was acquired from a source outside the long terminal repeat (LTR) retrotransposons[18]

Summary

Introduction

Retrotransposons are mobile genetic elements that replicate through an RNA intermediate. Structural similarity between the RNase H subdomain of HIV-1 RT and its connection subdomain (which lacks the catalytic carboxylates found in the RNase H domain) originally suggested the former arose through domain duplication, while an alternative theory proposes that the functional RNase H domain was acquired from a source outside the LTR retrotransposons[18]. Another well-characterized LTR element from S. cerevisiae is Ty1 of the Copia-like group, which is more distantly related to retroviruses than Ty3. Modeling of the spatial separation between the DNA polymerase and RNase H active sites, in addition to phenotypic mixing experiments, suggest that DNA polymerase and RNase H catalytic activities reside in separate subunits

Methods

Results

Conclusion