This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Morris, S. Articles by Leis, J. Search for Related Content PubMed PubMed Citation Articles by Morris, S. Articles by Leis, J.

 Previous Article  |  Next Article 

Journal of Virology, August 2002, p. 7571-7577, Vol. 76, No. 15
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.15.7571-7577.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Replication of Avian Sarcoma Virus In Vivo Requires an Interaction between the Viral RNA and the TC Loop of the tRNA^Trp Primer

Department of Biochemistry, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106-4935,¹ Department of Microbiology and Immunology, Northwestern University School of Medicine, Chicago, Illinois 60611²

Received 1 February 2002/ Accepted 22 April 2002

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 TC loop of the tRNA^Trp (U5-TC). We now demonstrate that this U5-TC 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-TC 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 TC 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-TC 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 TC loop of tRNA^Pro. These observations confirm the importance of the U5-TC interaction in vivo.

This article has been cited by other articles:

• Wei, M., Cen, S., Niu, M., Guo, F., Kleiman, L. (2005). Defective Replication in Human Immunodeficiency Virus Type 1 When Non-Primers Are Used for Reverse Transcription. J. Virol. 79: 9081-9087 [Abstract] [Full Text]  
• Abbink, T. E. M., Beerens, N., Berkhout, B. (2004). Forced Selection of a Human Immunodeficiency Virus Type 1 Variant That Uses a Non-Self tRNA Primer for Reverse Transcription: Involvement of Viral RNA Sequences and the Reverse Transcriptase Enzyme. J. Virol. 78: 10706-10714 [Abstract] [Full Text]  
• Goldschmidt, V., Paillart, J.-C., Rigourd, M., Ehresmann, B., Aubertin, A.-M., Ehresmann, C., Marquet, R. (2004). Structural Variability of the Initiation Complex of HIV-1 Reverse Transcription. J. Biol. Chem. 279: 35923-35931 [Abstract] [Full Text]  
• Kanevsky, I., Vasilenko, N., Dumay-Odelot, H., Fosse, P. (2003). In vitro characterization of a base pairing interaction between the primer binding site and the minimal packaging signal of avian leukosis virus genomic RNA. Nucleic Acids Res 31: 7070-7082 [Abstract] [Full Text]  
• Rigourd, M., Goldschmidt, V., Brule, F., Morrow, C. D., Ehresmann, B., Ehresmann, C., Marquet, R. (2003). Structure-function relationships of the initiation complex of HIV-1 reverse transcription: the case of mutant viruses using tRNAHis as primer. Nucleic Acids Res 31: 5764-5775 [Abstract] [Full Text]  
• Iwatani, Y., Rosen, A. E., Guo, J., Musier-Forsyth, K., Levin, J. G. (2003). Efficient Initiation of HIV-1 Reverse Transcription in Vitro. REQUIREMENT FOR RNA SEQUENCES DOWNSTREAM OF THE PRIMER BINDING SITE ABROGATED BY NUCLEOCAPSID PROTEIN-DEPENDENT PRIMER-TEMPLATE INTERACTIONS. J. Biol. Chem. 278: 14185-14195 [Abstract] [Full Text]  
• Goldschmidt, V., Ehresmann, C., Ehresmann, B., Marquet, R. (2003). Does the HIV-1 primer activation signal interact with tRNA3Lys during the initiation of reverse transcription?. Nucleic Acids Res 31: 850-859 [Abstract] [Full Text]  
• Kvaratskhelia, M., Miller, J. T., Budihas, S. R., Pannell, L. K., Le Grice, S. F. J. (2002). Identification of specific HIV-1 reverse transcriptase contacts to the viral RNA:tRNA complex by mass spectrometry and a primary amine selective reagent. Proc. Natl. Acad. Sci. USA 99: 15988-15993 [Abstract] [Full Text]