Replication of avian sarcoma virus in vivo requires an interaction between the viral RNA and the TpsiC loop of the tRNA(Trp) primer.

Abstract:

:Reverse transcription in avian sarcoma virus (ASV) initiates from the 3' end of a tRNA(Trp) primer, which anneals near the 5' end of the RNA genome. The region around the primer-binding site (PBS) forms an elaborate stem structure composed of the U5-inverted repeat (U5-IR) stem, the U5-leader stem, and the association of the tRNA primer with the PBS. There is evidence for an additional interaction between the viral U5 RNA and the T psi C loop of the tRNA(Trp) (U5-T psi C). We now demonstrate that this U5-T psi C interaction is necessary for efficient replication of ASV in culture. By randomizing specific biologically relevant regions of the viral RNA, thereby producing a library of mutant viruses, we are able to select, through multiple rounds of infection, those sequences imparting survival fitness to the virus. Randomizing the U5-T psi C interaction region of the viral RNA results in selection of largely wild-type sequences after five rounds of infection. Also recovered are mutant viruses that maintain their ability to base pair with the T psi C loop of the tRNA(Trp). To prove this interaction is specific to the tRNA primer, we constructed a second library, in which we altered the PBS to anneal to tRNA(Pro), while simultaneously randomizing the viral RNA U5-T psi C region. After five rounds of infection, the consensus sequence 5'-GPuPuCPy-3' emerged, which is complementary to the 5'-GGTTC-3' sequence found in the T psi C loop of tRNA(Pro). These observations confirm the importance of the U5-T psi C interaction in vivo.

journal_name

J Virol

journal_title

Journal of virology

authors

Morris S,Johnson M,Stavnezer E,Leis J

doi

10.1128/jvi.76.15.7571-7577.2002

subject

Has Abstract

pub_date

2002-08-01 00:00:00

pages

7571-7

issue

15

eissn

0022-538X

issn

1098-5514

journal_volume

76

pub_type

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